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autoMACS Pro Manual Software User Manual

1. 7048 Cause Plunger movement blocked because of column clogging blocked tubing set or any other cross section constriction during output of the positive fraction The positive fraction has not been eluted completely Positive cells are still remaining in the system Resuspensed sample could not be taken up completely no final air intake detected Leakage of air into system behind bubble sensor Portion of resuspended sample is remaining in tube Resuspensed sample could not be taken up completely no final air intake detected Leakage of air into system behind bubble sensor Portion of resuspended cells are remaining in tube Required volume cannot be provided in given Chill Rack Miscalculation of diluter movement Target position is negative 10 Troubleshooting Possible remedies Touch CONT to discard the remaining positive cells into the waste bottle To output positive fraction again at lower speed exchange positive tube with an empty tube and touch RETRY Check for air inlet into system behind bubble sensor Touch CONT to process currently uptaken sample volume Touch CANCEL to abort Check for leakage of air into system behind bubble sensor Readjust tubing connectors and verify correct fastening of columns Exchange rack with rack able to provide requested volume e g Chill 50 instead of Chill 15 Touch RETRY to try again Touch CANCEL to abort Touch RETRY to use target position 0 inst
2. 1 Open the box and remove the top compartment of the packaging to reveal the instrument and associated packaging 2 Remove boxes containing the MACS MiniSampler and cover and the MACS Cooling Tube Racks Remove the foam packaging from both sides of the autoMACS Pro Separator Figure 3 2 Packing format of the autoMACS Pro Separator and accessories 3 Place the instrument onto a stable worktop surface e g laboratory bench Remove the plastic bag surrounding the instrument Figure 3 3 Positioning the autoMACS Pro Separator securely onto a laboratory bench 3 Assembly and installation 4 Note the positions of the MACS MiniSampler guiding 2 and its 3 corresponding slot 1 located at the front of the instrument Tilt the MiniSampler and slide the guiding into the receiving slot until resistance is met lower the rack to a horizontal position i e the rack is locked in the position illustrated by the figure below Figure 3 4 Location of the MiniSampler guide 2 and receiving slot 1 for the MACS MiniSampler 5 Ensure that the MiniSampler is completely inserted and secure the connection by fastening the MiniSampler screw as shown in the figure below At Figure 3 5 Securing the MiniSampler by fastening the MiniSampler screw 6 Attach the sensor cables to the cable guide at the back of the autoMACS Pro Separator Figure 3 6 Sensor cables are attached to the back of the instrument as shown above
3. Figure 12 17 Unscrew the thumb screw 5 Note the position of the 2D code reader port Figure 12 18 The position of the 2D code reader port 6 Guide the cable of the 2D code reader underneath the lower Sn front access cover at the left hand side Make sure that 2D code reader cable is located in between the peristaltic pump tubes Figure 12 19 Make sure that 2D code reader cable is located in between the peristaltic pump tubes 7 Insert 2D code reader pin into corresponding opening 12 Hardware and software installation 209 Figure 12 20 Inserting the 2D code reader 8 Fix the connection by fastening the headless screw in the tapped bore hand tight with help of a 2 mm allen key Note Please check if headless screw is already present in the tapped bore Otherwise use headless screw provided with autolabeling package Figure 12 21 Fastening with the allen key 9 Reinstall the washing station Make sure that the cable is installed straight and is guided between the peristaltic pump tubing 210 12 Hardware and software installation 12 Figure 12 22 Reinstall washing station 10 Remove the black plastic square from the 2D code reader window of the Washing Station Cover not shown Snap in black optical frame Reinstall and close the washing station cover 11 Guide the 2D code reader cable underneath the instrument Connect the 2D code reader plug with the corresponding outlet at the back of the
4. Figure 7 13 Store program is underway 2 Install the column substitutes as described by the section 3 3 4 3 Select Done 7 Maintenance 153 154 Note It is recommended to exchange valves once a year Note If wear of a valve is suspected use the Check_up program to analyze functionality 4 Switch OFF the autoMACS Pro Separator using the main power switch 7 4 Planned maintenance 7 41 Valve exchange The fluidic system of the autoMACS Pro Separator is regulated by six valves The valve that is connected with the pump syringe is referred to as dilutor valve If a valve of the autoMACS Pro Separator starts to leak an exchange of the valve should be performed Exchange one valve at a time according to the following instructions 1 Select User settings from the Options menu Highlight Valve_ ex and press Run to start the program Select valves which have to be exchanged Valves will automatically be turned to exchange position 2 Switch OFF and unplug the autoMACS Pro Separator Open the front cover For exchange of the lower valves remove the bottom cover by pulling firmly Figure 7 14 The bottom cover must be removed for access to the lower valves 3 Using the supplied wrench unscrew the valve cover screw which releases the tubing 4 Detach all tubing and valve port locks or blind screws from the respective valve 7 Maintenance 5 Loosen the two valve screws using the screwdriver from th
5. in the Labeling submenu Change the sample volume as recommended in the data sheet For manual labeling it is recommended to dilute cells to a volume of 500 ul 108 total cells refer to corresponding data sheet for further information AT Figure 6 26 Only a positive cell separation will be performed on sample 1 7 View and recheck the sample setup 8 Select Run to start 9 Check buffer levels as instructed by a popup dialog box Click Continue to proceed 6 Cell separation using the autoMACS Pro Separator 139 140 10 Itis recommended to monitor the process from the Status menu Click the Status menu An RE 1 7 Figure 6 27 Sample processing is underway as instructed 6 7 Monitoring the autoMACS Pro Separator during cell separation The autoMACS Pro Separator is a sensor controlled instrument that allows easy monitoring during operation 6 7 1 Status menu before separation Refer to section 4 3 for an overview of how to monitor the instrument status using the Status menu prior to performing a cell separation Nevertheless a brief summary is included below Note Refer to section 4 3 Status of fluid containers for more information for The status of fluid containers is indicated by color coded graphic clarification of the Status symbols and by a text table menus Container Symbol Symbol color and user action Running Buffer Green no action required Red refill container Gray connect bottle sensor
6. 10 2 4 Low recovery of isolated cells 1 Low recoveries and low purities may be due to similar problems Go to section 10 2 3 steps 1 4 2 Low recoveries can also be caused by partial column blockage Run the Safe program and exchange the autoMACS Columns refer to sections 4 7 and 3 3 4 3 Performa test run using PBS as a mock sample and check output volumes For correct volumes refer to table 6 2 in section 6 3 3 If volumes are not correct go to section 10 2 1 4 Ifthe problem persists contact Technical Support Especially when using autolabeling 1 Check whether the uptake needle is positioned in the middle of the sample tube during the mixing procedure If not mixing will be inadequate 2 Check whether the reagent volume contents of reagent vial is sufficient for labeling 3 Check that the recommended maximal sample processing time of 90 minutes was not exceeded In the event of an elevated room temperature the cooling capacity of the Chill Racks may be compromised Higher labeling temperatures could cause unspecific binding which may lead to lower purities when using positive selection kits 10 2 5 Sample not or only partly taken up 1 If the sample has been taken up only partly check if the sample contains clumps larger than 1 mm Continue with the separation of this sample As soon as the separation is finished continue with step 2 Before processing any remaining samples use a Pre Separation Filter 130 041 407 t
7. 13 Note the position of the power socket on the rear panel of the autoMACS Pro Separator figure 3 7 Ensure that the main power switch is in position 0 14 Connect the power cord 15 Elevate the touchscreen and insert the memory card as shown below 3 Assembly and installation 83 84 gt Figure 3 10 Inserting the software memory card 16 Replace the fluid containers Refer to section 3 2 for full details 17 Switch on the autoMACS Pro Separator 18 Secure the MACS Reagent Rack 4 onto the MiniSampler into the left recess The engagement hook has to snap into the undercut si TER Figure 3 11 Securing the MACS Reagent Rack 4 onto the MACS MiniSampler 3 Assembly and installation Note Racks can be pre cooled for 3 4 hours at 2 8 C Do not cool below 0 C since samples may freeze If recognition of the tube rack fails the instrument will display a screen for manual selection of the tube rack Before confirming the choice ensure that the rack is placed correctly into the recess the MACS Triangle should face towards the user Note The 70 ethanol does not contain electrolytes Therefore the filling status of the ethanol container cannot be determined 19 Set a cool tube rack e g Chill Rack 5 onto the MiniSampler into 3 the right recess ensuring that the rack barcode is facing the autoMACS Pro Separator A AGO _ di 4 a d _ AES Figure 3 12 Posit
8. 2 3 Acolumn based high gradient magnetic cell separation The MACS Separation Unit consists of a powerful permanent magnet If a MACS Column is placed into the magnetic field of the Separation Unit the small ferromagnetic structures of the column matrix disturb this homogenous magnetic field and thereby produce high magnetic gradients In their immediate neighborhood the ferromagnetic structures generate magnetic forces allowing to retain target cells labeled with minimal amounts of MACS MicroBeads Additionally when the columns are removed from the separator this strong magnetic field is significantly reduced making the recovery of the retained cells easy and gentle This method allows separation of viable cells from samples containing a few hundred cells up to 10 total cells Within approximately 15 minutes the cells are labeled with MACS MicroBeads nano sized superparamagnetic particles coupled to specific antibodies The actual separation of cells over the column is completed within minutes 2 4 The autoMACS Pro Separator The autoMACS Pro Separator is a benchtop magnetic cell sorter that allows gentle sorting of more than 10 million cells per second from a sample of up to 4x10 total cells The instrument is designed for use with any MACS Cell Separation Reagent for research applications Thus it is possible to choose between different cell separation strategies according to the respective experimental design from positi
9. 3 Assembly and installation 81 82 7 Place the MiniSampler cable underneath the autoMACS Pro Separator and connect it to the socket labeled External CAN at the rear panel of the instrument Use the screw driver to fasten the screws of the connecter 8 Guide the 2D code reader cable underneath the instrument Connect the 2D code reader plug with the corresponding outlet at the back of the instrument labeled RS232 BCR Use the screw driver to fasten the screws of the connecter 9 Attach the sensor cable plug to the socket for sensor cables at the back of the autoMACS Pro Separator labeled Bottle Sensor and fasten securely Use the screw driver to fasten the screws of the connecter Sensor cable plug Code reader plug MACS MiniSampler plug Power socket Figure 3 7 Location of the MACS MiniSampler Sensor Cable Code Reader and power sockets at the back of the instrument 10 Carefully remove the uptake port needle from the foam packaging Bats Figure 3 8 Close up of the autoMACS Pro Separator the needle arm and underside of the touch screen are supported by foam 3 Assembly and installation Note Ensure that the tubing connected to the uptake port needle can move freely when the needle arm extends or when the needle moves into the sample uptake position 11 Place the uptake port needle into its guiding at the needle arm 3 Figure 3 9 The needle port is positioned as illustrated above
10. 5 The decontamination procedure is performed automatically Note In case of spillage it is recommended to use a 6 Optional Wash fluid containers and bottle closures using disinfectant that is detergent 1 hypochlorite or 70 ethanol Rinse thoroughly appropriate for the using deionized water potential pathogen e g bleach isopropyl alcohol 7 Upon completion of the process replace fluid containers and or 70 ethanol to reconnect all tubings decontaminate surfaces with tissue or swabs 8 Select OK The system is rinsed automatically 9 Wipe down the ports of the automated arm and the surface of the instrument using 70 alcohol swaps Note Dispose tissues and 7 3 5 Instrument long term storage swabs appropriately It is The program Store should be applied to prepare the instrument for recommended to wear long term storage Upon completion of the Store program the protective gloves fluidic system contains 70 ethanol protective clothing and safety glasses to prevent To store the autoMACS Pro Separator for a period longer than two contact with skin and weeks the tubing system should be cleaned and the columns should eyes Switch off and be replaced with column substitutes The Store program unplug the instrument automatically performs the cleaning procedure and prompts the beforehand user to install column substitutes 1 Select the Option Special and Store Select Run The system will be rinsed automatically l
11. If object blocked movement remove object Switch off instrument Wait 5 seconds Switch on instrument Otherwise or if error is displayed again call Technical Support Check bar code on rack or try different rack Make sure ambient or direct sun light does not hit sensor Clean sensor to remove salt or dust using a cloth soaked in 70 ethanol or double distilled water and wipe dry 187 188 Error code 264 769 775 777 Cause Unable to read rack bar code correctly Diluter plunger could not be initialized Syringe not mounted correctly syringe damaged or diluter valve not positioned correctly Diluter is not initialized After a diluter malfunction an initialization of the module is necessary This might also be a subsequent error if another error has been displayed shortly before Plunger movement blocked because of column clogging blocked tubing set or any other cross section constriction 6006 Airintake during sample uptake although needle did not yet hit bottom of tube Leakage of air into system in front of bubble sensor liquid level has been overestimated or needle did not move to bottom as fast as neccessary 6009 Unexpected air in system during sample uptake 6216 The reagent designated for the 7001 current labeling process is not assigned a position in the reagent rack Needle could not be retreated completely 10 Troubleshooting Possible remedies Che
12. Purity negative cells in negative non labeled fraction 9 3 Viability of the cells Different dyes are available to discriminate between live and dead cells The most common method to discriminate between live and dead cells is based on trypan blue staining and analysis by light microscopy Trypan blue crosses the cell membrane of dead cells and stains the cells Live cells are not stained Propidium iodide 5 0 g mL is most often used for flow cytometry and fluorescence microscopy It crosses the permeable cell membrane of dead cells enters the nucleus and interacts with DNA Therefore the nucleus of dead cells is fluorescently stained Other fluorescent dyes for example DAPI 4 6 diamidino 2 phenylindole can be used depending on the properties of the flow cytometer i e its excitation wavelength capabilities particularly in the UV range When working with fixed cells it is recommended to use the Fixation and Dead Cell Discrimination Kit 130 091 163 for both the cell fixation as well as the discrimination of dead cells The viability can be calculated as follows No of live cells Viability 100 x No of live cells live amp dead 9 Quality control of separations performed with the autoMACS Pro Separator Troubleshooting Note It is recommended to use the program check_up if general hardware errors occur Select the Options menu from the upper navigation bar Then select User settings
13. crasement et de cisaillement Risque d crasement et de cisaillement des membres corporels d a des dangers m caniques Rayonnement laser Danger de graves l sions oculaires et cutan es Champ magn tique puissant Le champ magn tique interf re avec les objets magn tisables et les appareils lectroniques ou endommage les supports d information magn tiques Risque de grave blessure corporelle pour les per sonnes portant un pacemaker ou des implants m dicaux lectron iques Risque de contamination en cas d utilisation de mati res nocives pour l organisme Indique le danger de mort le risque de blessure grave pour l op rateur de l appareil ou le risque de d t rioration de l quipement d des mati res biologiques potentiellement dangereuses Indique le danger de mort le risque de blessure grave pour l op rateur de l appareil en raison de la tension dangereuse Borne pour conducteur de protection Le symbole est appos l int rieur de l quipement Avertissement destin au personnel de maintenance Alimentation lectrique activ e Alimentation lectrique d sactiv e La notice d utilisation doit tre consult e avant de proc der l installation et l utilisation du syst me e0 gt gt gt gt TE 28 1 Important information 1 2 Avertissements et pr cautions Le s parateur autoMACS Pro fonctionne avec une technologie de pointe Il s agit d un appareil command par ordinateur con
14. since they do not bind free biotin Anti Immunoglobulin MicroBeads against human mouse rat or rabbit immunoglobulins can be used in combination with unconjugated antibodies 1 Prepare a single cell suspension and determine the cell number 2 Centrifuge cell suspension at 300xg for 10 minutes Aspirate supernatant completely 3 Resuspend cell pellet and incubate with the primary antibody according to the manufacturer s recommendations For MACS Fluorochrome conjugated or Biotinylated Antibodies resuspend up to 10 total cells in 100 uL of buffer and add 10 uL of the respective antibodies 4 Mix well and incubate in the refrigerator for 5 10 minutes 2 8 C or according to the manufacturer s recommendations If fluorochrome conjugated antibodies are used incubate in the dark 5 Wash cells to remove unbound primary antibody by adding 1 2 mL of buffer per 10 cells and centrifuge at 300xg for 10 minutes Aspirate supernatant completely 6 Optional Repeat washing step 7 Resuspend cell pellet in 80 uL of buffer per 10 total cells and add 20 uL of indirect MicroBeads per 10 total cells 8 Mix well and incubate for 15 minutes in the refrigerator 2 8 C 9 Optional When using unconjugated or biotinylated primary antibodies cells can be stained with a fluorochrome conjugated antibody at this point 10 Wash cells by adding 1 2 mL of buffer per 10 cells and centrifuge at 300xg for 10 minutes Aspirate supe
15. Advertencias y precauciones Precauciones generales Peligro de descarga el ctrica y propagacion de fuego Campo magn tico intenso Peligro de aplastamiento y corte Radiacion laser Instalaci n segura Accesorios de montaje Circulaci n de aire Agua y humedad Producto conectado a tierra Fuentes el ctricas Chapter Chapter Chapter Content 2 3 4 70 70 71 71 71 72 72 72 73 73 75 75 75 76 75 79 79 85 87 87 87 88 90 92 93 94 95 95 1 4 6 1 4 7 1 5 1 5 1 1 5 2 1 5 3 1 5 4 1 5 5 1 5 6 1 5 7 2 2 2 3 2 4 3 2 3 3 3 3 1 3 3 2 3 3 3 3 3 4 3 3 5 3 3 6 3 3 7 4 1 Accesibilidad Aparatos perif ricos Manejo mantenimiento transporte y eliminaci n seguros Manejo seguro Revision t cnica Limpieza Sustancias peligrosas Transport Eliminacion del instrumento equipo Compatibilidad electromagn tica Introduction MACS Technology the gold standard in cell separation Super paramagnetic MACS MicroBeads A column based high gradient magnetic cell separation The autoMACS Pro Separator Assembly and installation Unpacking and installing the autoMACS Pro Separator Replacing fluid containers and the connection of fluid sensor cables Running the autoMACS Pro Separator for the first time performing a test calibration Switch on the autoMACS Pro Separator Check the date and time Perform a
16. Indica una situaci n de peligro que si no se impide puede producir lesiones leves o moderadas Indica una situaci n de peligro que si no se impide puede producir la muerte o lesiones graves Atenci n consulte el manual de usuario para obtener m s infor maci n y proceda con precauci n Las advertencias incluyen el riesgo de da ar el equipo las lesiones personales graves o la muerte Peligro de aplastamiento o corte Riesgo de que se aplaste o corte alguna parte del cuerpo debido a peligros mec nicos Radiaci n l ser Riesgo de graves lesiones oculares y de piel Campo magn tico intenso El campo magn tico puede interferir en objetos magnetizables e instrumentos electr nicos o da ar portadores magn ticos de informaci n Riesgo de producir lesiones personales graves a personas que lleven un marcapasos o implantes m dicos electr nicos Riesgo de contaminaci n si se usan sustancias peligrosas biol gica mente Indica el riesgo de muerte lesiones graves al operario del aparato o da os al equipo debido a sustancias biol gicas potencial mente peligrosas Indica el riesgo de muerte o lesiones graves al operario del aparato debido a un voltaje peligroso Corriente continua El equipo est marcado con el tipo de suministro corriente continua Encendido suministro Apagado suministro La documentaci n del equipo debe ser consultada antes de proceder con la instalaci n y operaci n del sistema 1 Importan
17. Red Error Check screen for error detection Purple Program Sleep is completed Switch off autoMACS Pro Separator Blinking Action required Check screen for required action Table 6 7 Various bottle illumination statuses 6 Cell separation using the autoMACS Pro Separator The bottle illumination can be switched ON OFF 6 1 Select the Option menu and User settings 2 Highlight O_led and press Run The bottle illumination can now be enabled or disabled 6 8 Shut down the autoMACS Pro Separator 6 8 1 Sleep as the final wash program 1 Combine a separation program and the program Sleep for the last position in the programming field 2 Upon completion of the Sleep program switch off the autoMACS Pro Separator using the main power switch Note If Sleep is chosen 6 8 2 Store the program for long term storage as a wash program the To store the autoMACS Pro Separator for a period longer than two autoMACS Pro Separator weeks the tubing system should be cleaned and the columns should will not allow definition be replaced with column substitutes The Store program of any programs beyond automatically performs the cleaning procedure and prompts the this position user to install column substitutes 1 Select Option and Special The autoMACS Pro Separator automatically 2 Select Store and press Run The system will be rinsed performs a Sleep automatically program if the device is inactive for more than six hours Ensure that e
18. Separator for the first time performing a test calibration 3 3 1 Switch on the autoMACS Pro Separator The main power switch is located on the right side of the instrument in front of the container baskets I indicates on O indicates off Note Ensure that the Switch on the autoMACS Pro Separator autoMACS Pro Separator has been correctly installed section 3 1 and that the empty fluid containers used for transport purposes have been replaced by fresh fluid containers section 3 2 on off switch A Figure 3 13 Location of the on off switch 3 3 2 Checkthe date and time 1 To set the time and date perform the following steps Select Option User settings and Set_time Select Run Highlight either the time or date fields by touching the display Use the keypad to change the date or time accordingly RE o Figure 3 14 Select the desired field and use the numeric keypad to modify the date and or time Select OK to return to the Option menu The procedure can be cancelled at anytime by selecting Cancel 3 Assembly and installation 87 88 3 3 3 Perform atest calibration 1 Onthe touchscreen click Option Calibr_1 Run and Test E Figure 3 15 Instructing the instrument to perform a test calibration 2 Test calibration Chill Rack Ensure that a Chill Rack 5 is placed on the MACS MiniSampler Click Done to continue test calibration 3 Test calibration Reagent Rack Ens
19. Washing Solution Green no action required Red refill container Gray connect bottle sensor Storage solution Gray no liquid detection visually check volume Waste Green no action required Red empty waste Gray connect bottle sensor Table 6 6 Status of fluid containers displayed in the Status menu 6 Cell separation using the autoMACS Pro Separator Column status 6 If the column symbol is green no action is required If the symbol is red the columns must be exchanged The level of the green fill on the column symbol indicates the remaining service life of the autoMACS Columns Rack detection Rack detection only occurs upon starting the separation process MACS MiniSampler detection If the MACS MiniSampler has been detected correctly a rectangular symbol is displayed If it has not been detected a hand symbol will be displayed in the same field 6 7 2 Status menu during cell separation Ss It is recommended to monitor the instrument s status during cell separation using the Status menu Programs yet to be processed appear in yellow fields Programs currently undergoing autolabeling appear in lilac Programs in progress in which no autolabeling is being performed switch to orange completed programs switch to white The current action is always displayed in the status bar located below the lower navigation bar The status bar also displays the overall progress in minutes min Frung a Figure 6 28
20. el uso del instrumento ya no es seguro Apague inmediatamente el separador autoMACS Pro desench felo y p ngase en contacto con el proveedor de servicios de Miltenyi Biotec autorizado o con el servicio de atenci n al cliente de Miltenyi Biotec 1 5 2 Revisi n t cnica IMPORTANTE Salvo que el presente manual de usuario u otra docu mentaci n de Miltenyi Biotec especifique lo contrario no revise usted mismo el separador autoMACS Pro Las revisiones y reparaciones deben ser llevadas a cabo por personal cualificado Las revisiones y reparaciones del separador autoMACS Pro incorrectamente realizadas pueden poner en peligro a sus usuarios producir resultados impre decibles derivar en un mal funcionamiento del aparato o que ste sufra da os as como causar un desgaste prematuro y reducir el tiempo de vida del instrumento pudiendo anular su garant a Pregunte a su representante de Miltenyi Biotec por los contratos de servicios de asistencia t cnica de Miltenyi Biotec o consulte www miltenyibiotec com support IMPORTANTE Cuando se necesiten piezas de recambio o de repuesto aseg rese de que el proveedor de servicios utiliza exclusivamente piezas originales de Miltenyi Biotec o de otros fabricantes especifica dos y recomendados por Miltenyi Biotec El uso de piezas de recam bio o de repuesto no autorizadas puede producir un mal funcionamiento del aparato y alterar los resultados de la separaci n celular Miltenyi Biotec no cubrir
21. gekennzeichnet Gleichstrom Stromversorgung AN Stromversorgung AUS Vor Inbetriebnahme und Betrieb des Ger ts ist die Dokumentation zu beachten 1 Important information 1 2 Warn und Sicherheitshinweise 1 Der autoMACS Pro Separator ist ein hochmodernes computerges teuertes Ger t zur automatischen Separation magnetisch markierter Zellen mittels MACS Technologie Der MACS MiniSampler ist an den autoMACS Pro Separator angeschlossen und damit Teil der Zellsepa rationsvorrichtung autoMACS Pro Separator und MACS MiniSampler gelten bei Einbau und Bedienung durch fachkundiges Personal und Beachtung der allgemein blichen Sicherheitspraktiken und Hinweise in dieser Bedienungsanleitung als sicher Dieses Kapitel erkl rt die im Umgang mit dem Ger t m glichen Gefahren und gibt wichtige Sicherheitshinweise zur Vermeidung dieser Gefahren Halten Sie sich genau an diese Hinweise um sich selbst und das Ger t vor m glichen Gefahren zu sch tzen und ein sicheres Arbeit sumfeld zu garantieren Bei Verwendung des Ger ts unter Missach tung der Herstelleranweisungen ist die Sicherheit nicht gew hrleistet ACHTUNG Bitte lesen und befolgen Sie alle in diesem Benutzerhand buch gegebenen Hinweise zum Betrieb des Ger tes Beachten Sie auch alle auf dem Display erscheinenden Warnhinweise Bewahren Sie dieses Benutzerhandbuch sowie alle weiteren mit diesem Ger t erhaltenen Sicherheits und Betriebsanleitungen an einem f r alle Nutze
22. liti Figure 6 11 Using the 2D code reader two reagent vials were assigned to the reagent rack positions R1 and R2 2 Select Save Template 3 Allocate a name to the template In this example the template was saved as EXPT_2 experiment 2 6 Cell separation using the autoMACS Pro Separator 127 128 RE 0 Figure 6 12 Using the alphanumeric keypad assign a name to the template 4 Select Ok To load a reagent template 1 Select Load Template from the lower navigation bar To scroll through the list of saved templates use the navigation arrows A Y The corresponding template is displayed in the adjacent panel E 0 27 Figure 6 13 Selecting a reagent template Expt_2 was selected which comprises of CD4 h and CD8 m on positions R1 and R2 respectively 2 Select and highlight the desired template in this case Expt_2 3 Select Ok Lesa Figure 6 14 The template Expt_2 was successfully loaded To delete a reagent template 1 Select Load Template from the lower navigation bar 6 Cell separation using the autoMACS Pro Separator Note For a detailed introduction to the separation menu refer to section 4 4 Note option under Labeling denotes that NO autolabeling will be performed Only a cell separation will be performed manual magnetic labeling must be performed prior to placing the sample on the Chill Rack 2 Scroll through the list of saved templa
23. toute d t rioration de l appareil 11 Symboles et niveaux de danger Mise en place des consignes de s curit Example CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard Les consignes de s curit informent l utilisateur des risques poten tiels pouvant survenir si les avertissements et les mesures de pr caution d crits ci dessous ne sont pas respect s Le pictogramme visible gauche d finit le risque En haut le danger mentionn pr c demment est affect un niveau de danger Le niveau le type et la source du danger ainsi que les ventuelles cons quences les interdictions et les mesures prendre sont mentionn s conform ment au tableau ci dessous 1 Important information 27 Symboles et niveaux de danger Le tableau suivant est un glossaire illustr expliquant les symboles utilis s dans ce mode d emploi et sur le s parateur autoMACS Pro Separator Indique une situation dangereuse qui si elle n est pas vit e peut CAUTION z DIE entra ner des blessures mineures ou mod r es Indique une situation dangereuse qui si elle n est pas vit e peut WARNING A entra ner la mort ou des blessures graves Attention consultez le mode d emploi pour obtenir plus de con signes et agissez avec prudence Les avertissements incluent le risque de d t rioration de l quipement de blessure corporelle grave et le danger de mort Risque d
24. Der Hauptstromschalter ebenso wie der Netzstecker f r das Stromka bel sollten leicht zug nglich sein und sich in m glichst unmittelbarer N he zum Bediener des Ger tes befinden Sollte es erforderlich sein die Stromzufuhr zu unterbrechen ziehen Sie den Netzstecker 1 4 7 Peripherieger te An die mit COM beschriftete RS 232 Buchse d rfen nur Peripher ieger te angeschlossen werden die UL 60950 erf llen Die mit nRS232 AUX beschriftete Buchse ist nicht belegt An die mit External CAN CAN1 und CAN2 beschrifteten Buchsen d rfen nur Original autoMACS Pro Ger te angeschlossen werden Die Spannung an diesen Buchsen darf die gef hrlichen Werte von 30 Veff und 42 4 V Spitze bzw 60 V DC nicht bersteigen An die mit Bottle Sensor beschriftete Buchse darf nur das autoMACS Pro Flaschensen 1 Important information sorkabel angeschlossen werden An die mit RS232 BCR beschriftete 1 Buchse darf nur ein von Miltenyi Biotec empfohlener 2D Barcode leser angeschlossen werden Externe Laserger te die an die mit RS232 BCR beschriftete Buchse angeschlossen werden m ssen IEC 60825 1 erf llen Es d rfen nur Anschlusskabel mit einer L nge von lla max 3 Metern verwendet werden 1 5 Sicherer Betrieb Wartung Transport und Entsorgung Beachten Sie die folgenden die folgenden Hinweise bez glich Betriebssicherheit Wartung Transport und Entsorgung Ihres autoMACS Pro Separators WICHTIG rtliche Arbeit
25. Highlight check_up and press Run The program automatically analyzes the functionality of moving hardware components A report is displayed after the analysis of each single component The procedure can be canceled after each step or continued by pressing OK Following hardware components are analyzed dilutor valve valves 1 5 peristaltic waste pump magnet 1 2 needle arm movement along the z axis and the y axis and MACS MiniSampler Furthermore the calibration data is checked 10 In the unlikely event of problems using the autoMACS Pro Separator the following section aims to address any relevant issues If the outcome of a cell separation procedure is deemed unsatisfactory this may either be due to incorrect function of the instrument or to inappropriate sample preparation Both of these factors are dis cussed in this section At the end of this section a list of numerically encoded errors and warning messages are presented along with user actions for troubleshooting This section addresses problems that are not indicated by a warning or error screen but might occur during the separation or rinsing programs Identify the problem and refer to the appropriate section 10 1 Hardware problems not indicated by a warning or error screen 10 1 1 Column leakage 1 Ifa freshly installed autoMACS Column shows signs of leakage check if the column is installed properly The column should be inserted precisely into the c
26. Pin 7 RTS Pin 8 CTS RS232 Interface labeled RS232 Pins 1 4 6 7 8 9 NC AUX Pin 2 RXD Not in use Pin 3 TXD Pin 5 GND RS232 Interface DC Output Pins 4 6 NC labeled RS232 BCR Pin 1 Input Pin 2 RXD Pin 3 TXD Pin 5 GND Pins 7 8 Shorted Pin 9 5 VDC 0 5 A CAN Bus DC Output labeled Pins 1 4 8 NC External CAN Pin 2 CAN L Pins 3 6 GND Pins 5 9 24 VDC 2A Pin 7 CAN H AC Output labeled Bottle Sensor Pins 1 2 3 4 5 5 VAC 10 kQ Pins 6 7 8 14 15 GND Pins 9 10 11 12 13 Input CAN Bus labeled CAN1 or Pins 1 4 5 8 9 NC CAN2 Pin 2 CAN L Pins 3 6 GND Pin 7 CAN H Table 11 2 Pin assignment for autoMACS Pro Separator The autoMACS Pro Separator is labeled as a protection class instrument and must be plugged into a grounded power outlet refer to section 1 The main power supply cord and plug of the instrument shall comply with following specifications USA and Canada only UL listed and KAM cord minimum type SJ minimum 18 AWG 3 conductors Rated for a minimum temperature of 60 C Provided with grounding type NEMA 5 15P attachment plug rated 125 Vac 10 A Opposite end terminates in IEC 320 style connector rated 125 Vac 10 A Conditions of operation 15 30 C with 0 85 humidity non condensing at a maximum altitude of 2000 m Supply voltage fluctuations up to 10 of the nominal voltage Transient overvoltages present on
27. Select Yes from the popup dialog box 7 Maintenance 147 148 Note Ensure enough storage solution is available Do vou remy wer 10 ef Ame he device DR gt Figure 7 3 Shutting down the autoMACS Pro Separator 3 The autoMACS Pro Separator will automatically perform a Sleep program 4 Upon completion of the Sleep program you will be prompted to shut down the instrument The bottle illumination is purple at this point Switch OFF the autoMACS Pro Separator by using the main power switch Store the program for long term storage To store the autoMACS Pro Separator for a period longer than two weeks the tubing system should be cleaned and the columns should be replaced with column substitutes The Store program automatically performs the cleaning procedure and prompts the user to install column substitutes 1 Select Option and Special 2 Select Store and press Run The system will be rinsed automatically Rinsng RS gt gt Figure 7 4 Running the Store program 3 Install the column substitutes refer to section 3 3 4 for details on column exchange 4 Select Done 5 Switch OFF the autoMACS Pro Separator using the main power switch 7 Maintenance 7 3 Periodic maintenance 7 3 1 Column exchange autoMACS Pro Seperator Columns must be exchanged every two weeks or after 100 separations whichever comes first Refer to section 3 3 4 for more details 7 3 2 Clean the pump syringe Clean pump s
28. Separator Depletion programs 6 DEPLETE Depletion in standard mode for removal of cells with normal to high antigen expression and results in better target cell yield DEPLETES Depletion in sensitive mode removal of cells with low antigen expression and results in better target cell purity DEPLO5 Depletion in sensitive mode Il removal of cells with low antigen expression and results in stringent depletion of cells DEPLO25 Depletion in sensitive mode Ill removal of cells with low antigen expression and results in stringent depletion of cells A_DEPLO7 Depletion in standard mode via loading of sample in separate 1 mL stages removal of cells with normal to high antigen expression and results in a better target cell yield This special program is disabled by default To enable select Option User settings and O_progs A_DEPLS7 Depletion in sensitive mode via loading of sample in separate 1 mL stages removal of cells with low antigen expression and results in a better target cell purity This special program is disabled by default To enable select Option User settings and O_progs Positive selection programs In programs Possel and Possels the magnetically labeled target cells are retained in the autoMACS Column 1 The unlabeled cells are released into the negative fraction collection tube row B of the tube rack After automated retraction of the magnet the magnetically labeled cells are eluted into the
29. To delete a separation template 1 Select Load Template from the lower navigation bar 2 Scroll through the list of saved templates use the navigation arrows M W 3 Select the template for deletion DRE 45 Figure 4 23 Expt_3 template was selected for deletion 4 Select Delete Template 4 An overview of the autoMACS Pro touchscreen user interface 105 106 4 5 Reagent menu The reagent menu is used to program any reagent vials that are required for automated magnetic labeling and subsequent cell sorting Reagents can be entered using the 2D code reader or manually using the Enter Reagent input panel For convenience it is possible to load and save reagent templates N Figure 4 24 The reagent menu 4 6 Log list menu RE os Figure 4 25 The Log list display The Log list records a complete log of actions performed by the autoMACS Pro Separator An overview of the log list table follows 4 An overview of the autoMACS Pro touchscreen user interface Name Definition Name Definition A Date Indicates the date Program Name of program for the action was example performed Deplete Cell depletion Possel Positive cell selection Qrinse Quick rinse performed Type Description of the Protocol If a protocol is action for example associated with the i Rack Action log its name will be involving rack listed under this Init Initiation of heading for instrument example Wash only
30. Wash CD4 h CD4 only performed MicroBeads human Service Service step were used for performed autolabeling and cell Special wash separation Special wash performed Pos Corresponding Status The status of the log position on sample is depicted as tube rack follows EA successfully completed EA action failed To display further details about an individual log 1 Select Log list tab and highlight a log from the log list 2 Select Details A detailed view of the program status is shown 3 Select Ok to return to the log list screen or select Log Details to view a detailed log of performed actions 4 An overview of the autoMACS Pro touchscreen user interface 107 108 lite Figure 4 26 Viewing log details A A CDA MicroBead positive cell selection was performed on March 26th 2009 B An overview of program actions and corresponding times are displayed C Details of each instrument action are listed 4 7 Option menu eil Figure 4 27 The Option display The Option menu allows maintenance procedures such as exchange of autoMACS Columns or decontamination of the system The menu is divided into two main categories Special and User settings An overview of the functions available under each category are given below 4 An overview of the autoMACS Pro touchscreen user interface 4 7 1 Special A Special options comprise three special programs for column exchange Col_ex instrument decontaminatio
31. air circulation around the autoMACS Pro Separator at least 15 cm on all sides during operation to ensure adequate cooling of the instrument Prevent direct exposure of the instrument to sunlight Slots and openings of the instrument are provided for ventilation and should never be blocked or covered as these ensure reliable operation of the autoMACS Pro Separator and protect the instrument from overheating Never push a foreign object through an opening into the instrument 1 4 3 Water and moisture Do not use the instrument in a wet or damp location Avoid high humidity or condensation and protect the machine against water splashes 1 4 4 Grounded earthed product The instrument is equipped with a three wire electrical grounding type plug that has a third pin for grounding This plug only fits into a grounded power outlet This is a safety feature Do not try to insert the plug into a non grounded power outlet If you cannot insert the plug into the outlet contact your local electrician to replace the outlet 1 4 5 Power sources The instrument should only be operated from a power source indicated on the product s electrical ratings label If you have questions about the type of power source to use contact your authorized Miltenyi Biotec service provider or local power company 1 Important information 21 Do not use extension cords or power strips Do not overload an electrical outlet The overall system load must not exceed 80
32. and thus contaminate the positive fraction Use buffers devoid of Ca and Mg in order to reduce formation of cell aggregates 10 Troubleshooting 181 182 Dead cells in the cell suspension may non specifically bind to MicroBeads and will then be co enriched in the positive fraction Remove dead cells before separation by using the MACS Dead Cell Removal Kit 130 090 101 or by Ficoll Paque density gradient The concentration of MicroBeads or the temperature used for magnetic labeling were too high It is recommended to dilute the MicroBeads according to the data sheet Cells should be incubated with MicroBeads in the refrigerator 2 8 C The total number of cells may have been miscounted leading to an inappropriate ratio of MicroBeads and cells Use a ratio of MicroBeads and cells as stated in the respective MACS Cell Separation Reagent data sheet Incubation time with MicroBeads was too long leading to background labeling Reduce the incubation time to the recommended values refer to individual MACS Cell Separation Reagent data sheet Typically an incubation time of 15 minutes is required when incubating in the refrigerator 2 8 C For exceptions refer to the respective data sheet When target cells are extremely rare lt 5 of total cells few non labeled cells which are retained non specifically may constitute a high portion compared to the target cells Performa second separation without re labeling to remove n
33. and accessory hardware during operation Keep fingers etc away from all moving parts of the autoMACS Pro Separator and accessory hardware to avoid crushing or shearing injuries or damage to the instrument Do not touch fluid pumps or adjust the tubing while the instrument is in operation Always switch off the instrument before adjusting any part of the fluidic system Always stop or abort a procedure before handling accessory hardware e g MACS MiniSampler or loading removing tubes from the tube rack placed on the sampler Do not circumvent any safety measures or devices Figure 1 2 Open circle shows warning sign for hazard of crushing and shearing 1 3 4 Laser radiation WARNING The instrument is equipped with four vertical cavity surface emitting lasers VCSELs for automated rack detection Class 1M The radiation is not visible Do not view directly with optical instruments e g lenses magnifying glasses and microscopes Viewing the VCSEL port within 100 mm distance with optical instruments could be hazardous to the eye The instrument is also equipped with a 2D Code Reader which uses a visible semiconductor laser as a target pointer for adjusting the reading position and powerful light emitting diodes LEDs for illuminating the reading area Do not look directly at laser or LED radiation or reflected laser or LED radiation from a mirrored surface Otherwise eye injury may result Do not intentionally direct the
34. cleaned every 1 3 months Appropriate maintenance and long term storage assures that no salt deposits accumulate in the pump syringe Salt deposits may cause wear of the pump seal and thus may lead to leakage The pump syringe should not run dry at any time This can damage the pump seal and thereby may lead to leakage of the pump syringe Note To prevent the formation of salt deposits wipe the outlet ports with a tissue soaked with distilled or deionized water before each Sleep program 10 1 4 Pump syringe is filled with air during operation 1 O 1 If there is any air inlet into the pump syringe during operation the correct proceeding of a separation will be impaired 2 Check all tubings that are connected to the fluid bottles Make sure that all tubings are fastened properly If a screw thread is damaged order and install new tubing 3 Check if the hydrophobic air filters connected to the bottles are clogged Clogging may cause positive or negative pressure in the fluid bottles which can lead to pressure problems in the fluidic system If filters are clogged replace them with new hydrophobic air filters refer to section 7 4 5 4 Check if the connections and pump syringe are leaky refer to section 10 1 3 5 Check if the uptake port needle is connected correctly and no air inlet is possible If not unscrew and check screw threads If they are undamaged reinsert precisely and fasten Then use the wrench t
35. como hardware accesorios deber n ser retirados Unicamente por personal autorizado Se debe tener especial cuidado cuando se manejen liquidos Limpie inmediatamente los liquidos vertidos Impida que stos accedan al interior del aparato Desenchufe el cable de electricidad antes de proceder a limpiar manualmente el separador autoMACS Pro Existe un riesgo potencial en caso de usar separador autoMACS Pro abierto que se haya caido o que est averiado si se han derramado liquidos en el equipo si se ha colado algun objeto por las ranuras de ventilaci n o si ha caido algo dentro del equipo Si salen llamas o humo apague inmediatamente el separador autoMACS Pro des ench felo y p ngase en contacto con un proveedor de servicios de Miltenyi Biotec autorizado o con el servicio t cnico de Miltenyi Biotec Esta expresamente prohibido utilizar un instrumento estropeado o cuyo cable de corriente est da ado 1 3 2 Campo magn tico intenso WARNING Extremely strong magnetic fields Risk of severe personal injury to persons carrying pacemakers or electronical medical implants stay clear Keep all magnetically sensitive objects away from device WARNING El separador autoMACS Pro Separator esta equipado con un iman extremadamente potente Mantenga todos los portadores de informaci n magn tica tarjetas de cr dito cintas magn ticas y medios de almacenamiento y todos los equipos electr nicos aud fonos marcapasos instrumentos de
36. fluids with ethanol or disinfectant 7 Maintenance 11 Carefully remove the waste distributor from its position by pulling in an upward direction and remove the draining tube from the old diluter valve Figure 7 22 The waste distributor 12 Mount the new draining tube and finger tighten Take care to clean spilled fluids with ethanol or disinfectant bio Figure 7 23 The draining tube is attached 13 Install the waste distributor back to its former position and reinstall the washing station 14 Install the syringe 15 Switch ON the autoMACS Pro Separator 16 Check for correct function by running the program Rinse 7 Maintenance 159 160 Note The pump syringe should be exchanged once a year Note The peristaltic pump head should be exchanged once a year 17 Run the program Calibr_2 to calibrate the fluidic volume control of the instrument 7 4 3 Exchange of the peristaltic pump head 1 Switch OFF and unplug the instrument 2 Remove the washing station as described in section 7 3 3 steps 1 6 3 Press the tubing clamp on the left hand side of the waste distributor and remove the tubing 4 Pullout the bottom cover as depicted above 5 Press clamps on both sides of the pump and pull out the pump head 6 This will uncover a pin that protrudes from the instrument Clean the pin using 70 ethanol but do not attempt to pull it out The pin drives the pump during operation 7 Re
37. for needle retreat Check that cover of the washing station is properly closed Remove any objects hindering the movement of the needle If needle hits rim of tube or top of rack calibrate needle arm Touch RETRY to try again Check that cover of the washing station is properly closed Remove any objects hindering the movement of the needle If needle hits rim of tube or top of rack calibrate needle arm Touch RETRY to try again Do not use sample volumes exceeding the maximum volume specified for rack or program type If volumes are in specified range but error is displayed anyway please call Technical Support Please clean outside of needle manually If this error is thrown frequently please call Technical Support Check if tubes are correctly positioned in rack correspond ing to template programming Check MACS MiniSampler connection in front of autoMACS Pro Separator 189 190 Error code 7010 7011 7012 7013 7014 7015 7018 7021 7022 7023 7024 7026 Cause Not certain if liquid surface of sample has been detected correctly Restart of the device is required Calibration data not found Calibration data not found Calibration data not found Columns are not installed Calibration data not found A separation program has been started but system had not been rinsed properly Columns are overdue Plunger movement blocked because of column clogging blocked
38. montaje No coloque el separador autoMACS Pro sobre una mesa carrito estante tripode o soporte inestable puesto que podria caerse causando lesiones corporales graves y o da os graves al equipo Utilice exclusivamente una mesa carrito estante tripode o soporte recomendado por Miltenyi Biotec o vendido junto con el instru mento No coloque el separador autoMACS Pro en un mueble empotrado o un espacio reducido como por ejemplo una estanteria a menos que el mismo haya sido dise ado espec ficamente para dar cabida al instrumento que provea una ventilaci n adecuada y que se hayan seguido las instrucciones de montaje del equipo 1 4 2 Circulaci n de aire El instrumento no deberia ser colocado cerca de radiadores rejillas de calor estufas o cualquier otra pieza de equipamiento incluidos 1 Important information 69 70 los amplificadores que produzca calor Permita que circule suficiente aire alrededor del separador autoMACS Pro deje al menos 15 cm de separaci n en todas direcciones mientras esta en funcionamiento para garantizar que el instrumento se enfrie adecuadamente Evite la exposici n directa a la luz solar Las ranuras y aberturas del instru mento sirven para que se ventile y no deben ser bloqueadas o cubiertas puesto que contribuyen a un funcionamiento seguro del separador autoMACS Pro evitando que se recaliente No introduzca ning n cuerpo extra o por las aberturas del instrumento 1 4 3 Agua y humedad No use
39. needle arm should almost touch the bottom of a MACS Reagent vial If the needle is correctly positioned click Done Ver ty reagent rack postion 3 Assembly and installation 89 90 Note If test calibration 1 fails performa calibration 1 Refer to section 12 2 for details Note When delivered the autoMACS Pro fluidic system is filled with double distilled water The autoMACS Columns are reusable for two weeks or up to 100 separations whichever comes first The instrument automatically records the date of the column exchange and can display the due date for the next column replacement if the program Column exchange Col_ex has been used for column installation Columns do not have atop or bottom and do not require special orientation in the column holder Figure 3 18 Test calibration Control position of needle in vial 7 Test calibration is completed 3 3 4 Installation of the autoMACS Columns Remove the column substitutes and install the autoMACS Columns according to the following instructions 1 Open the front door and note the position of the tubing and autoMACS Column slots in the magnet cover column 1 to the left column 2 to the right 3 a 2 Ensure that the fluid containers are filled with solutions 3 Using the touch screen select Option and Special from the Main panel 4 Select Col_ex column exchange from the Detail panel RS 3 0 Figu
40. of the sample rack This program is used for very strong depletion of those cells that weakly express the antigens used for magnetic labeling or to achieve the highest purity of the untouched cell fraction However choosing Depl025 might result in a reduced recovery of the target cell fraction FITC Fluorescein isothiocyanate Fluid container 1 5 L bottles holding the fluids for operational use of the autoMACS Pro Separator Fluid sensors monitor the fluid levels in the containers for Running Buffer Washing Solution and waste via electrolyte conductivity The fluid level in the container for the 70 ethanol cannot be monitored as no electrolytes are present in the solvent Fluid sensors This sensor type measures electrolyte conductivity and is an integral part of the bottle closures of the fluid containers for waste Running Buffer and Washing Solution Fluid sensor cable Cable connecting the fluid sensor to the autoMACS Pro Separator The sensor cable connectors on the bottle closures are color coded red for waste green for Washing Solution and blue for Running Buffer 15 Glossary Fraction collection tube 5 mL 15 mL or 50 mL plastic sample 1 5 tubes to collect the positive and negative fractions The process has been optimized using tubes from BD Falcon Front door The front door opens sidewards giving access to the autoMACS Columns pumps valves washing station and tubings MACS Technology Technology developed b
41. of untouched human monocytes using the Monocyte Isolation Kit Il human 130 091 153 autolabeling program MO IK h separation program Deplete Sample 2 Direct labeling and positive cell enrichment of CD4 cells using CD4 MicroBeads human 130 045 101 autolabeling program CD4 h separation program Possel 2 Select Save Template 3 Allocate a name to the template In this example the template was saved as EXPT_2A experiment 2a 4 An overview of the autoMACS Pro touchscreen user interface 103 Note The sample separation template was loaded however the reagent rack has not yet been configured It is necessary to configure the reagent rack before starting the run 104 E 02 Figure 4 20 Using the alphanumeric keypad assign a name to the template 4 Select Ok To load a reagent template mi Select Load Template from the lower navigation bar To scroll through the list of saved templates use the navigation arrows A Y The corresponding template is displayed on the sample template panel EEE 0 gt Figure 4 21 Selecting a separation template Expt_2a was selected 2 Select and highlight the desired template in this case Expt_2a 3 Select Ok E 0 gt Figure 4 22 The template Expt_2a was successfully loaded 4 An overview of the autoMACS Pro touchscreen user interface Currott rack coda be stares Click desa s for nore n crratan rr ED
42. or deactivate an automatic instrument prime on instrument startup 7 2 2 Clean uptake outlet ports It is recommended to clean uptake and outlets daily This should be performed before running the Sleep program shutdown 1 Switch the instrument OFF 2 The ports can be wiped with tissue soaked with 70 ethanol isopropyl alcohol or MACS Bleach Solution followed by distilled or deionized water 7 Maintenance Note To prevent the formation of salt deposits wipe the outlet ports with a tissue soaked with distilled or deionized water before each Sleep program 3 Remove the finger guard that is attached underneath the outlet J port holder by pulling gently Clean the finger guard as described for the ports Figure 7 2 Removing the finger guard for cleaning 4 Wipe the outlet port for the negative fraction as indicated above The port can be flushed by using a syringe 5 To clean the uptake port ensure the instrument is switched OFF Move the needle holder up and down to get access to the entire surface of the needle 6 Push the finger guard back into position 7 Switch the instrument ON 8 Run the Sleep program 7 2 3 Instrument shutdown Make sure to run a Sleep program prior to switching the instrument OFF This program ensures most efficient preservation of the fluidic system by rinsing with storage solution Shutdown button 1 Press the shutdown button on the upper right hand corner of the screen D 2
43. removal 10 11 12 13 14 15 Insert one end of a fresh column into the bottom column connector and gently screw in the column by turning it clockwise until you feel a resistance Point the column towards the top of the instrument and screw in the top column connector Align the column with the top column connector sitting on the guiding of the magnet cover Press the column into the slot until you feel the guides click Verify that the column is placed in the center of the magnet cover Repeat steps 8 through 11 to install the second column Ensure that the tubes are securely fastened to the columns and that the tubing is neither pinched nor obstructed Press Done Close the front door The unit is now ready to perform cell separations 3 3 5 Priming the autoMACS Pro Separator Priming implicates the initial cleaning and filling of the autoMACS Pro Separator tubing system before cell separations are performed The autoMACS Pro Separator must be primed each time the instrument is switched on 2 Fill all bottles with the appropriate solutions and empty the waste bottle Switch on the instrument After initialization is completed the Status menu will be displayed Verify that the touchscreen symbols for all fluid containers are green If this is not the case check if the fluid sensor cables are connected to the correct fluid containers The symbol for the ethanol container remains gray fluid level not check
44. safety for more information regarding the installation of the equipment 1 3 General precautions To reduce potential risks associated with operating the autoMACS Pro Separator please observe the following general precautions Failure to observe these precautions could result in fire bodily harm and or damage to the instrument 1 3 1 Hazard of electric shock and spread of fire WARNING Hazardous voltages Risk of loss of live severe personal injury Unplug before cleaning Do not use if device is opened or damaged if liquids have been spilled into device if objects device through ventilation slots WARNING Electrical devices pose the risk of an electric shock To reduce the risk of an electric shock do not open any cover other than the front access covers of the autoMACS Pro Separator nor any other accessory hardware supplied by Miltenyi Biotec 1 Important information 17 All other covers of the instrument and accessory hardware are to be removed by authorized personnel only Special care must be taken while handling fluids Clean up spillages immediately Do not allow fluids to enter the interior of the instrument Unplug the power cord before manually cleaning the autoMACS Pro Separator WARNING A potential risk exists if an opened dropped or damaged autoMACS Pro Separator is used if liquids are spilled into the instrument if an object has entered the instrument through the ventilation slots or if an obj
45. the mains supply category Il The instrument is suitable for rated pollution degree 2 Emission sound pressure level at workstation lt 70 dB A The autoMACS Pro Separator is not specified for use in the cold 11 Technical data and specifications room The autoMACS Pro Separator has been investigated by 1 1 Underwriters Laboratories in accordance with the standards UL 61010 1 CAN CSA C22 2 No 61010 1 and IEC 61010 2 081 and meets the intent of the directive 2004 108 EC electromagnetic compatibility and 2006 95 EC low voltage equipment EMC EN 61326 1 EN 61000 3 2 EN 61000 3 3 Low voltage equipment EN 60825 1 EN 61010 1 EN 61010 2 081 Compliance was demonstrated by conformance to the following standards which have been listed in the Official Journal of the European Communities Compliance was demonstrated by conformance to the following FCC Rules of the Code of Federal Regulations 47 CFR 15 class B 11 3 Technical data and specifications of the MACS MiniSampler Parameter Specification Model MACS MiniSampler Input voltage 24 VDC Current 0 8A Sub D9 interface Pin 1 NC with shielding Pin 2 CAN Pin 3 GND Pin 4 NC Pin 5 24V Pin 6 GND Pin 7 CAN Pin 8 NC Pin 9 24 V Table 11 3 Technical data and pin assignment for MACS MiniSampler 11 Technical data and specifications 199 200 The MACS MiniSampler is labeled as a protection class Ill instrument and must only be plugged into t
46. the prompt to enter a Note Refer to sections 4 4 template name and 4 5 for an overview of the separation and 4 Select Ok and Run reagent tabbed menus respectively To select a particular sample position in the programming field touch it once To deselect it touch it once again If the same combination of separation program and washing program is required for more than one sample highlight all the desired sample positions first and then select the programs Alternatively define the program combination for one position and then highlight the other desired positions to adopt the program combination To erase selected program combinations from the programming field first mark the respective positions and choose Delete selected from the lower navigation bar If positions are not marked the button can be used to delete the entire template In this case the option Delete selected will switch to Delete all Alternatively start the separation from a previously saved separation template Note For autolabeling 1 Select Load template from the lower navigation bar the Reagent Rack configuration must be 2 Select the desired template by using the arrows programmed using the Reagent tab menu Refer 3 Select OK to section 4 5 or 6 3 2 for further details 4 Select Run to start the separation Check that there is enough buffer for the number of programmed separations To obtain brief information on the 5 Select Continue se
47. tion of or to any part of the Product use of accessories spare parts and or consumables other than those recommended by Miltenyi Biotec or normal wear and tear Miltenyi Biotec s product warranty does not cover products sold AS IS or WITH ALL FAULTS or which had its serial number defaced altered or removed or any consumables or parts identified as being supplied by a third party those third party accessories or parts may be covered by a separate warranty from their manufacturer Miltenyi Biotec must be informed immediately if a claim is made under such warranty If a material or manufacturing defect occurs within the warranty period Miltenyi Biotec will take the appropriate steps to restore the full usability of your Product Limitation on damages Miltenyi Biotec shall not be liable for any incidental or conse quential damages for breach of any express or implied warranty or condition on this Product 14 Limited warranty 215 Some states or jurisdictions do not allow the exclusion or limitation of incidental or consequential damages so the above limitations or exclusions may not apply to you This warranty statement gives you specific legal rights and you may have other rights which may vary from county to country or jurisdiction to jurisdiction 216 14 Limited warranty Glossary Air filter Hydrophobic 0 2 um air filter attached to the bottle closure Used to vent the bottle and at the same time prevent contaminants
48. to run program Safe once a month Disinfectant solution It is recommended to clean the ports of the automated arm and the 5 Materials required surface of the instrument of spilled fluid with 70 ethanol or 5 isopropyl alcohol on damp tissue Alternatively use alcohol swabs Distilled water It is recommended to use distilled water on tissue to remove any salt crusts from the instrument 5 2 2 Hardware autoMACS Columns Order no 130 021 101 5x2 columns Capacity of 2x10 magnetically labeled cells from up to 4x10 total cells a Column substitutes Order no 130 090 835 2 pieces For installation prior to storage of the autoMACS Pro Separator for longer than two weeks The instrument is delivered with installed column substitutes Hydrophobic 0 2 um air filters Order no 130 090 385 Hydrophobic air filters are used to vent fluid containers and to prevent aerosols Do not use hydrophilic filters since they are easily blocked upon contact with liquid and thus may cause errors autoMACS Pump Syringe Order no 130 090 339 1 2 required per year Wrench Order no 130 090 378 For tightening tube connections Included in the autoMACS Pro Starting Kit autoMACS Pro Laminar Hood Plate Order no 130 093 246 The metal plate has been developed for stable placement of the autoMACS Pro Separator on potentially bending surfaces for example the surface in a laminar flow hood or safety cabinet The thick metal plate is desig
49. tubing set or any other cross section constriction during output of the negative fraction The negative fraction has not been eluted completely Negative cells are still remaining in the system The number of programmed sample positions exceeds the actual positions of the rack on the MACS MiniSampler The protective cover of the MACS MiniSampler seems to be opened by the moving needle arm hitting the cover 10 Troubleshooting Possible remedies Touch IGNORE to continue without liquid detection Needle will be moved to bottom directly This might result in a subsequent warning 7008 if the liquid level is higher than 60 mm above the tube bottom see above TOUCH retry to continue with liquid detection Ensure that the tubing from the needle arm to bubble sensor can move freely Adjust the tubing if necessary Otherwise call Technical Support Please restart instrument Please calibrate needle arm axes Run program Calibr_1 Please calibrate needle arm axes Run program Calibr_1 Please calibrate tubing Run program Calibr_2 Please install columns Please calibrate MACS MiniSam pler using program Calibr_1 Please rinse system by touching WASH or abort with CANCEL Please install new columns Touch CANCEL to abort and then install columns touch CONTINUE to ignore and use old columns not recommended Touch CONT to discard the remaining negative cells into the waste bottle To output negative fraction again at l
50. u pour la s paration automatique des cellules marquage magn tique l aide de la technologie MACS Le mini chantillonneur MACS est branch au s parateur autoMACS Pro et fait donc partie du dispositif de s paration des cellules Le s parateur autoMACS Pro et le mini chantillonneur MACS sont con us pour fonctionner en toute s curit apr s leur installation s ils sont utilis s par des personnes qualifi es selon les pratiques de s curit g n rales et les consignes mentionn es dans ce mode d emploi Les instructions de ce chapitre expliquent les risques potentiels li s l utilisation de l appareil et fournissent des informations de s curit importantes afin de r duire ces risques Respectez strictement ces consignes pour prot ger l quipement et vous m me des risques potentiels et cr er un environnement de travail s r La protection n est pas garantie si cet appareil n est pas utilis conform ment aux instructions donn es par le fabricant IMPORTANT Veuillez lire et respecter toutes les instructions d utilisation fournies dans ce mode d emploi et observez tous les avertissements affich s sur l appareil Conservez ce mode d emploi et les autres consignes de s curit et d utilisation fournies avec l appareil un endroit accessible pour tous les utilisateurs en vue de les consulter ult rieurement IMPORTANT Le s parateur autoMACS Pro Separator est destin une utilisation int rieure uniquement N u
51. 0 nanometers in diameter being comparable to the size of a virus MicroBeads do not change the scatter properties of the cell in the flow cytometer or influence the light microscopic appearance of the cell They form a stable colloidal suspension and do not precipitate or aggregate in magnetic fields MACS MicroBeads are composed of a biodegradable matrix made of iron oxide and polysaccharide Hence it is not necessary to detach cell bound beads after the separation process saving hands on time Usually MACS MicroBeads do not alter structure function or activity status of labeled cells and they are not known to interfere with subsequent experiments The isolated cells can be used directly for subsequent studies or cell culture MACS Cell Separation Reagents are highly specific Thus it is possible to label for example B cells and T cells as well as rare target cells with frequencies as low as 108 MACS Technology allows to isolate viable antigen specific T cells CD133 hematopoietic stem and progenitor 2 Introduction 75 76 cells subpopulations of CD34 hematopoietic progenitor cells and disseminated carcinoma cells circulating in the peripheral blood Finally MACS MicroBeads offer an extremely flexible tool to isolate many cell types from many species Several hundreds of reagents for the isolation of human mouse rat and non human primate cells as well as reagents for indirect labeling of many other cell types are available
52. 1 is opened sidewards giving access to the washing station the peristaltic pump and the tubing of the autoMACS Pro Washing Station 11 Technical data and specifications 193 194 Fluid containers and fluid container baskets Two baskets figure 11 1 holding two fluid containers each are located at each side of the instrument Fluid containers are con nected to the autoMACS Pro System with color coded tubing and sensors at the bottle closures for fluid level control figure 11 1 Figure 11 3 Right side view of the autoMACS Pro Separator Plugs connections and guidings Sockets for the main plug figure 11 4 the fluid sensor cables the 2D code reader cable and the MACS MiniSampler are installed at the rear of the instrument Additional sockets are implemented for further instrument development The main power switch is located at the right hand side of the instrument figure 11 4 Several guidings at the rear and sides of the instrument ease the safe connection of tubings and sensor cables Figure 11 4 Rear view of the autoMACS Pro Separator 11 Technical data and specifications MACS MiniSampler Chill Racks and MACS Reagent Rack 1 1 The MACS MiniSampler figure 11 5 can be loaded with one of three different cooling tube racks that carry cell samples and fraction collection tubes and the MACS Reagent Rack 4 The upper plate of the MiniSampler moves in an x direction and aligns the tube openings with the port of the
53. 3 and R4 BEE 17 25 Figure 6 5 Position R2 was selected for the reagent vial 2 Select Enter Reagent from the lower navigation bar Enter the reagent specific product order number The order number is located on the product data sheet In the event that the data sheet is misplaced visit www miltenyibiotec com to download a printable PDF of the document 6 Cell separation using the autoMACS Pro Separator Figure 6 6 Reagent information is manually entered using the reagent order number 3 Select OK 4 Ifa correct number is inserted the software will immediately recognize the reagent or kit To confirm select the reagent from the list by using the touch screen REE 0 Figure 6 7 CD4 MicroBeads human 130 045 101 was manually entered 5 Select OK to confirm the identified reagent and reagent vial position R2 6 Cell separation using the autoMACS Pro Separator 125 126 __ Figure 6 8 Reagent vial CD4 MicroBeads human was assigned to the reagent rack position R2 Deleting reagents or reagent lists In this example the following reagents were scanned into the following reagent rack positions R1 FcR Blocking Reagent human component 1 3 of product Monocyte Isolation Kit Il human 130 091 153 R2 Monocyte Antibody Cocktail human component 2 3 of product Monocyte Isolation Kit Il human 130 091 153 R3 Anti Biotin MicroBeads component 3 3 of pro
54. 3 302 130 093 303 130 093 304 130 093 305 130 093 306 Spare part Square Key Valve Blind Screw 5 pcs autoMACS Pro Tube t4 Fuse 5x20 T4A set autoMACS Pro Sensor Cable autoMACS Pro Bottle Closure green autoMACS Pro Bottle Closure blue autoMACS Pro Bottle Closure black Table 7 1 List of available spare parts 7 Maintenance Part order number 130 093 371 130 093 372 130 093 397 130 093 407 130 093 669 130 094 682 130 094 729 Spare part 4 port 3 way Valve autoMACS Pro Tube t31 Uptake Port Needle Guiding needle arm Thumb Screw M5x40 MiniSampler Cover MiniSampler Pump Syringe Hamilton 5mL Cavro XP Dilutor Valve 6 port distribution V2 165 7 9 Accessories list Accessory Accessory Accessory Accessory order order number number 130 091 339 Air Filter Extension 130 093 412 MACS Cooling Tube Set Rack 3xChill 15 130 093 245 autoMACS Pro Angle 130 093 416 MACS Cooling Tube Connector set Rack 3xChill 5 130 094 573 autoMACS Pro 130 093 413 MACS Cooling Tube Autolabeling Rack 3xChill 50 Upgrade Kit 130 093 305 autoMACS Pro Bottle 130 092 952 MACS Cooling Tube Closure blue Rack Chill 15 130 093 304 autoMACS Pro Bottle 130 092 951 MACS Cooling Tube Closure green Rack Chill 5 130 093 246 autoMACS Pro 130 092 953 MACS Cooling Tube Laminar Hood Plate Rack Chill 50 130 094 289 autoMACS Pro 130 092 990 MACS Cooling Tube Laminar Hood Se
55. 47 CFR 15 class B 11 Technical data and specifications Hardware and software installation 12 Note Do not remove or insert a memory card while the instrument is turned ON The memory card must remain in the unit Removing the memory card during operation will abort all running processes 12 1 Installation of new software After installing an autoMACS Pro software update it is necessary to calibrate the instrument as instructed below The autoMACS Pro Separator contains various control boards During initialization of the instrument with a software card all of these components are automatically checked for the currently installed software 12 1 1 Exchanging the software cards 1 Ensure the instrument is switched off before proceeding 2 Note the memory card slot at the right hand side of the touch screen f tt Figure 12 1 Location of the memory card slot 3 Remove the old memory card by pressing the black release button beneath the card slot 4 Insert the new memory card 5 Switch the instrument ON A dialog box will appear prompting the operator to proceed if desired 6 Select Update to confirm that you wish to proceed 12 Hardware and software installation 201 202 Note During the panel software update process the display may turn black or white or may freeze or flicker This process may take several minutes The progress will be indicated by an acoustic signal If warning 7030
56. 5 Transport Le s parateur autoMACS Pro Separator doit tre transport avec soin dans un emballage sp cifi par Miltenyi Biotec Un dommage interne peut survenir si l appareil est soumis a des vibrations excessives ou s il chute Si l appareil doit tre r exp di pour tre remis en tat d contaminez l appareil afin d liminer toute mati re dangereuse avant le transport Si vous avez des questions concernant la propre d contamination ou l exp dition n h sitez pas contacter notre service technique assistance 1 5 6 Elimination de l appareil Si vous souhaitez liminer votre appareil veuillez contacter notre service technique qui vous aidera 1 5 7 Compatibilit lectromagn tique Pour les tats Unis uniquement Toute modification et toute transformation de l quipement sauf si elle a t formellement autoris e par Miltenyi Biotec peut annuler votre droit d utilisation de l quipement conform ment la norme 47 CFR art 15 1 Important information 1 37 38 1 Important information Wichtige Information 1 Bitte lesen vor Inbetriebnahme Bitte lesen Sie vor Inbetriebnahme sorgf ltig alle in diesem Benutzerhandbuch enthaltenen Informationen Werden die folgenden Anleitungen nicht gelesen und beachtet so k nnte dies zu unsachgem er oder unvorschriftsm iger Anwendung Bedienung oder Wartung des Ger tes f hren Auf diese Weise k nnten Anwender gef hrdet Ihr Ger t besch digt oder d
57. 950 pueden ser conectados con el conector RS232 rotulado con COM No se utiliza el conector rotulado con RS232 AUX Adicionalmente deben conectarse exclusivamente equipos originales de autoMACS Pro con los conectores rotulados con External CAN CAN1 y CAN2 Los niveles de voltaje de estos conectores no debe sobre pasar los niveles de voltaje peligroso de 30 V CA y 42 4 V punta o 60 V CC Se debe conectar solamente el cable de sensor de botella MACS Pro con el conector del sensor de botella Con el conector RS232 BCR debe conectarse solamente un lector de c digo 2D recomen dado por Miltenyi Biotec Los dispositivos de l ser externos conecta 1 Important information dos con el conector rotulado con RS232 BCR deben cumplir con la 1 norma IEC 60825 1 Utilice exclusivamente cables de conexi n con un largo menor a 3 m 1 5 Manejo mantenimiento transporte y eliminaci n seguros Observe las siguientes instrucciones para asegurar un manejo mantenimiento transporte y eliminaci n de su separador autoMACS Pro seguros IMPORTANTE En todo momento se debe observar la normativa nacional de seguridad en el trabajo las normas del laboratorio y los est ndares de salud y seguridad en el laboratorio y de prevenci n de accidentes 1 5 1 Manejo seguro Si el instrumento no funciona correctamente y las instrucciones o mensajes que aparecen en la pantalla aconsejan ponerse en contacto con el servicio t cnico
58. Anti Biotin antibodies do not bind to free biotin 10 4 2 Cells are excessively stained 1 High background staining may occur when the concentration of the staining reagents is too high and the incubation time for the staining procedure is too long The use of the pre titrated MACS Fluorochrome conjugated Antibodies is recommended to achieve optimal results 2 Check the staining reagent for its specificity 10 5 Contamination of tubing system 1 Decontaminate the instrument refer to section 7 3 4 for details 2 Ifthe problem persists call Technical Support 10 6 Problems indicated by error or warning screens If errors or warnings are displayed on the screen of the instrument please refer to the following table If the below measures do not clear the fault call Miltenyi Biotec Technical Support To assist in the troubleshooting process please have the instrument serial number and details of the error message at hand i e error number module number file and error line If this is not possible please go to the instrument Log List and view the Log Details taking care to note the displayed parameters exactly To discuss the table below please call Technical Support 10 Troubleshooting Error Cause code 5 Hardware module is not initialized After a module malfunction an initialization of the module is neccessary This might also be a subsequent error if another error has been displayed shortly before Standard valve ini
59. CLs para la detecci n automatizada de reactivos clase 1M La radiaci n no es visible No 1 Important information 67 68 mire directamente con instrumentos pticos por ejemplo gafas lupas y microscopios Puede ser peligroso para el ojo mirar el puerto VSCL con instrumentos pticos a una distancia de 100 mil metros El dispositivo tambi n est equipado de un lector de c digo 2D que utiliza un l ser semiconductor visible como indicador de blanco para ajustar la posici n de lectura y diodos electroluminosos de gran alcance LED para iluminar el rea de la lectura No mirar directamente el l ser radiaci n LED l ser reflejado o radiaci n del LED de una superficie reflejada De otra manera podr a ocasionarse una lesi n ocular No dirigir intencionalmente el rayo l ser a otras personas No desmontar modificar ni quitar el l ser o la fuente de radiaci n LED o sus soportes de montaje El l ser o las fuentes de radiaci n LED no interrumpen la emisi n cuando est n desmontados La radiaci n de unidades desmontadas puede ocasionar lesiones oculares Tener cuidado con la trayectoria del rayo o la reflexi n l ser de una superficie reflejada Durante la instalaci n del autoMACS Pro Separator tener cuidado de que durante la operaci n la trayectoria del rayo l ser no est a la misma altura que el ojo humano No permitir que agua aceite polvo u otras sustancias ajenas se peguen a la ventana de abertura del le
60. Copyright 2010 Miltenyi Biotec All rights reserved No part of this user manual may be reproduced stored in a retrieval system transmitted published or distributed in any form or by any means electronically mechanically by photocopying microfilming recording or otherwise without the prior written consent of Miltenyi Biotec however notwithstanding the foregoing the owners of the autoMACS Pro Separator may make copies solely for purposes of training personnel in the use and servicing of the unit within their business or organization Maximal care has been taken by Miltenyi Biotec in the preparation of this user manual However Miltenyi Biotec shall not be liable for any technical or editorial errors or omissions contained herein or for incidental or consequential damages in connection with the furnishing performance or use of this document The information in this document is provided as is without warranty of any kind and is subject to change without notice autoMACS MACS and the MACS logo are registered trademarks or trademarks of Miltenyi Biotec GmbH or its affiliates in Germany the United States and or other countries Ficoll and Ficoll Paque are trademarks of GE Healthcare companies BD and BD Falcon are trademarks of Becton Dickinson and Company All other trademarks mentioned in this document are the property of their respective owners and are used for identification purposes only Unless otherwise specifically indica
61. Ensure enough reagent is present in all vials for all programmed processes As the residual volume in the vial is about 20 uL please be sure to overfill the vial with reagent accordingly 10 3 Magnetic labeling and separation 10 3 1 Positive selection Positive cells have not been retained on the column 1 Magnetic labeling of the cells is insufficient because the MicroBead concentration was too low This may be due to the fact that either too much buffer was added to the cells or too much buffer was left on the cell pellet after centrifugation before adding the MicroBeads Furthermore the total number of cells 10 Troubleshooting may have been miscounted Use a ratio of MicroBeads and cells as 1 O stated in the respective MACS Cell Separation Reagent data sheet 2 Labeling of cells was ineffective due to too much debris and or dead cells in the sample Debris and dead cells will non specifically bind to all other components present Antibodies and MicroBeads will be captured non specifically and not be able to label the cells in the positive fraction sufficiently any more 3 Labeling of cells was ineffective due to an incubation temperature lower than recommended It is recommended to incubate cells in the refrigerator 2 8 C for labeling 4 The number of magnetically labeled cells exceeds the column capacity Calculate the number of magnetically labeled cells for example by staining with flurochrome conjugated antibodies and sub
62. Monitoring the status during cell separation Sample processing at position 1 is completed white color Sample at position 2 has finished the separation and is now performing a Rinse Interrupting cell separation The cell separation process can be paused at any time from all menu screens by selecting Stop 1 From any menu screen select Stop The autoMACS Pro Separator will then immediately stop operation and will display a pop up warning screen from which the current program can be continued or canceled 6 Cell separation using the autoMACS Pro Separator 141 142 Note Interrupting the process after labeling will prolong the incubation period Paola sie 0 Figure 6 29 Select Stop to pause cell separation To continue cell separation select Continue To cancel the entire procedure select Cancel 2 Select Cancel to cancel the procedure Alternatively select Continue to carry on with cell separation i Figure 6 30 The process has been cancelled Bottle illumination The autoMACS Pro Separator has a bottle illumination that facilitates monitoring of the instrument s status The table below summarizes the color code of the bottle illumination and the respective user action required Code Status User action Green Ready for separation No action required Blue Instrument operating No action required Yellow Not ready for separation Run wash program Rinse or Qrinse before starting a separation
63. N sse Geerdetes Produkt Stromquellen Zug nglichkeit der Stromverbindungen Peripherieger te Sicherer Betrieb Wartung Transport und Entsorgung Sicherer Betrieb des Ger tes Wartung Reinigung Gef hrliches Material Transport Ger teentsorgung Content 51 51 51 52 53 53 54 54 55 56 57 57 58 58 58 58 58 59 59 58 59 60 60 61 61 61 63 63 63 64 65 65 66 66 67 67 69 69 69 70 70 70 1 1 1 2 1 3 1 3 1 1 3 2 1 3 3 1 3 4 1 4 1 4 1 1 4 2 1 4 3 1 4 4 1 4 5 1 4 6 1 4 7 1 5 1 5 1 1 5 2 1 5 3 1 5 4 1 5 5 1 5 6 1 5 7 1 1 1 2 1 3 1 3 1 1 3 2 1 3 3 1 3 4 1 4 1 4 1 1 4 2 1 4 3 1 4 4 1 4 5 Informazioni importanti Simboli e livelli di pericolo Indicazioni di sicurezza Simboli e livelli di pericolo Avvertenze e precauzioni Precauzioni generali Pericolo di scossa elettrica e di incendio Forte campo magnetico Pericolo di schiacciamento e di taglio Radiazioni laser Installazione sicura Accessori di montaggio Circolazione dell aria Acqua e umidita Prodotto con messa terra Fonti di alimentazione Accessibilita Periferiche Funzionamento manutenzione trasporto e smaltimento sicuri Funzionamento sicuro Manutenzione Pulizia Materiale pericoloso Trasporto Smaltimento Compatibilit elettromagnetica Informacion importante Simbolos y niveles de peligro Disposici n de advertencias de seguridad Simbolos y niveles de peligro
64. NTE El separador autoMACS Pro est dise ado para ser usado exclusivamente en interior No utilice el equipo en reas clasificadas como peligrosas como ambientes con alta concentraci n de ox geno P ngase en contacto con la autoridad local competente para el suministro de electricidad construcci n de edificios mantenimiento o seguridad para obtener m s informaci n sobre la instalaci n del equipo Si tiene serias dudas sobre el manejo seguro del equipo p ngase en contacto con su proveedor de servicios de Miltenyi Biotec autorizado O llame al servicio de atenci n al cliente de Miltenyi Biotec 1 3 Precauciones generales Para reducir los potenciales riesgos asociados al manejo del separa dor autoMACS Pro por favor observe las siguientes precauciones generales En caso de no seguir estas precauciones puede pro ducirse un incendio o causar da os corporales o al equipo 1 Important information 65 1 3 1 Peligro de descarga el ctrica y propagaci n de fuego WARNING Hazardous voltages Risk of loss of live severe personal injury Unplug before cleaning Do not use if device is opened or damaged if liquids have been spilled into device if objects device through ventilation slots WARNING Los aparatos el ctricos pueden producir una descarga el ctrica Para reducir este riesgo no abra ninguna cubierta ni abra tampoco ning n otro accesorio de harware suministrado por Miltenyi Biotec Cualquier cubierta asi
65. S il s av re n cessaire de d connecter l alimentation lectrique d branchez le cable 1 4 7 Appareils p riph riques Seuls les appareils p riph riques conformes la norme UL 60950 1 Important information peuvent tre branch s au connecteur RS232 tiquet COM Le 1 connecteur tiquet RS232 AUX n est pas utilis De plus seuls des appareils autoMACS Pro originaux doivent tre raccord s aux connecteurs tiquet s External CAN CAN1 et CAN2 Le niveau de tension mesur sur ces connecteurs ne doit pas d passer lla le niveau de tension dangereux de 30 Veff ainsi qu un pic de 42 4 V ou 60 V DC Seul le cable de capteur de flacon autoMACS Pro peut tre raccord au connecteur tiquet Bottle Sensor Seul un lecteur de code 2D recommand par Miltenyi Biotec peut tre branch au connecteur RS232 BCR Les appareils laser externes raccord s au connecteur tiquet RS232 BCR doivent r pondre a la norme IEC 60825 1 Utilisez uniquement des cables de connexion de 3 m tres de longueur maximum 1 5 S curit d utilisation maintenance transport et limination Respectez les consignes suivantes afin de garantir la s curit d utilisation la maintenance le transport et l limination de votre s parateur autoMACS Pro Separator IMPORTANT Les consignes de s curit concernant la zone de travail locale les bonnes pratiques de laboratoire ainsi que les directive
66. TANT Please do not perform any maintenance procedures other than specifically described in this user manual Any other maintenance procedures must be performed by qualified service personnel Inquire with your local Miltenyi Biotec representative about Miltenyi Biotec s extensive instrument service and support arrangements or refer to www miltenyibiotec com support IMPORTANT When replacement or spare parts are required for maintenance only use genuine Miltenyi Biotec parts or third party parts specified and recommended by Miltenyi Biotec Using unauthorized replacement or spare parts can cause malfunction of the instrument and impair cell separation results Miltenyi Biotec does not honor any warranty or accept any responsibility for instrument failure or damages resulting from the use of inappropriate replacement or spare parts CAUTION During a maintenance procedure potentially contaminated liquid may spill out of the orifice of the washing station and the tubing Therefore wear protective gloves protective clothing and safety glasses to avoid contact with skin and eyes Dispose used gloves and clothing appropriately 7 1 General considerations Please consider that the sample quality has a significant influence on the system s performance It is crucial to use single cell suspensions for cell separation Furthermore dead cells and other small particles potentially derived from the sample preparation should be removed prior to
67. There are two basic approaches to magnetic labeling labeling with direct MACS MicroBeads or indirect labeling with MicroBeads against primary antibodies or their conjugates General instructions for magnetic labeling are described in the following sections Direct magnetic labeling A variety of antibody conjugated MicroBeads targeting human non human primate mouse and rat cell surface markers are available for direct magnetic labeling The general protocol given below is based on a 1 5 dilution of MACS MicroBeads Please note that direct magnetic labeling of mouse cells is typically performed in a 1 10 dilution of MACS MicroBeads For details refer to the specific MACS Cell Separation Reagent data sheet 1 Prepare a single cell suspension and determine the cell number 2 Centrifuge cell suspension at 300xg for 10 minutes Aspirate supernatant completely 3 Resuspend cell pellet in 80 uL of fresh buffer per 10 total cells 4 Add 20 ul of MACS MicroBeads per 10 total cells 5 Mix well and incubate for 15 minutes in the refrigerator 2 8 C longer incubation times may lead to non specific cell labeling Fluorescent labeling can be performed either before or after the separation has been completed 7 Wash cells by adding 1 2 mL of buffer per 10 cells and centrifuge at 300xg for 10 minutes Aspirate supernatant completely 8 Resuspend up to 10 cells in 500 uL of buffer 9 Proceed to magnetic separation refer
68. a The 2D code reader is classified as a Class 1 laser product per standard IEC 60825 1 1993 A1 1997 A2 2001 maximum output 116 uW wavelength 655 nm pulse duration 1 ms Please refer also to section 1 3 4 of the autoMACS Pro Separator user manual for associated warning and precautionary information Refer to section 12 3 for information on installation of the 2D code reader Figure 11 7 Expanded view of the autoMACS Pro Separator 2D code reader barcode reader The technical specifications of the autoMACS Pro Separator and the peripheral devices are as follows 196 11 Technical data and specifications 11 2 Technical data and specifications of the autoMACS Pro Separator Parameter Specification Color Footprint Footprint with MiniSampler Height Weight Input voltage autoMACS Separator Power consumption Fuses Programs Sample volume input Sample volume output Column capacity Blue orange 605 mm x 343 mm w x d 605 mm x 455 mm w x d 392 5 454 mm adjustable touchscreen 25 kg 100 240 VAC 50 60 Hz 200 VA 2xT4A 250 12 preset 0 25 50 mL 0 5 52 mL 4x10 cells sample 2x10 magnetically labeled cells sample 15 mL of whole blood Table 11 1 Technical data for the autoMACS Pro Separator 11 Technical data and specifications 11 197 198 Model autoMACS Pro Separator RS232 Interface labeled COM Pins 1 4 6 9 NC Pin 2 RXD Pin 3 TXD Pin 5 GND
69. a di 20 cm dal coperchio del magnete Questi oggetti potrebbero essere danneggiati o il loro funzionamento essere compromesso dal campo magnetico 1 Important information Figura 1 1 Segnale di avvertenza per forte magnete permanente 1 3 3 Pericolo di schiacciamento e di taglio CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard ATTENZIONE Non aprire i coperchi di accesso anteriore mentre il dispositivo in funzionamento Non impedire il movimento del braccio automatizzato e degli accessori hardware durante il funzionamento Tenere le dita ecc lontano da tutte le parti in movimento del dispositivo autoMACS Pro Separator e degli accessori hardware per evitare lesioni da schiacciamento e ferite da taglio o danni al dispositivo Non toccare le pompe dei liquidi e non spostare i tubi mentre il dispositivo in funzione Spegnere sempre il dispositivo prima di regolare eventuali parti del sistema della fluidica Interrompere o sospendere sempre una procedura prima di maneggiare gli accessori hardware ad es il campionatore MACS MiniSampler o prima di caricare rimuovere le provette dal portaprovette collocato sul campionatore Non bypassare alcun dispositivo o misura di sicurezza Figura 1 2 Il cerchio indica il segnale di avvertenza per il pericolo di schiacciamento e di taglio 1 Important information 55 1 3 4 Radiazioni laser L apparecchio do
70. action collection tubes have been in contact with biohazardous material they should be autoclaved after use Bottle closures can be autoclaved 5 Unscrew the thumb screw that attaches the washing station to the instrument Figure 7 11 Unscrew the thumb screw 6 Pull out the washing station 7 Clean the washing station by soaking it in 10 bleach and 70 ethanol for 15 min each Optionally sonicate it in water Rinse with distilled water 8 Reassemble the unit in reverse order 9 Runa Safe program 7 3 4 Instrument decontamination The Safe program is a disinfectant procedure which uses MACS Bleach Solution for cleaning and decontamination of the autoMACS Pro Separator Depending on the level of use and general instrument maintenance it is recommended to decontaminate the fluidic system every 3 to 6 months using the Safe program If predominantly whole blood bone marrow or tissue samples are separated the Safe procedure can be performed more often without harming the instrument e g every four weeks instead of performing a column exchange Perform instrument decontamination according to the following instructions 1 Select menu Option Special and the program Safe RS 130 Figure 7 12 Safe program is underway 7 Maintenance 2 Press Run Follow the screen prompts 3 Disconnect the tubings from all buffer bottles Select OK 4 Place the end of each tubings into a bottle of MACS Bleach Solution
71. aisser un espace d au moins 20 cm entre le cache de l aimant et les supports information magn tiques cartes de cr dit bandes magn tiques et supports de donn es ainsi que l quipement lectronique appareils auditifs pacemakers dispositifs de mesure et de contr le ordinateurs et montres Ces l ments peuvent tre affect s ou endommag s par le champ magn tique distance d au moins 30 cm par rapport au mat riel magn tique mentionn ci dessus Plus ils sont proches l un de l autre plus la force attractive entre les deux aimants ou entre un aimant et 1 Important information une mati re magn tisable augmente Par cons quent n approchez 1 aucune mati re magn tique y compris tout autre aimant ou cran Figure 1 1 Symbole d avertissement pour aimant puissant permanent 1 3 3 Risque d crasement et de cisaillement CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard Avertissement N ouvrez pas les capots frontaux pendant le fonction nement de l appareil Ne bloquez pas le mouvement du bras automa tique ni des pi ces correspondantes pendant le fonctionnement N approchez pas vos doigts etc des pi ces mobiles du s parateur autoMACS Pro et des pi ces correspondantes afin d viter toute blessure ainsi que toute d t rioration de l appareil due l ecrasement et au cisaillement Ne touchez pas les pompes liquide et ne modifiez
72. al incubation time is 15 min Please refer to the respective data sheet for more details Especially by using autolabeling 1 Check whether the reagent vial has enough reagent to perform magnetic cell labeling Check whether the uptake needle is positioned in the middle of the sample tube during the mixing procedure If not limited mixing could lead to low purity Refer to the respective data sheet for the recommended separation program Recovery of target cells is low Refer to section 10 3 1 Overall recovery is low Refer to section 10 3 1 10 3 3 Indirect MACS MicroBeads Positive cells have not been retained on the column 1 The unbound primary antibody was not completely removed and inhibits magnetic labeling with indirect MACS MicroBeads Wash cells carefully optionally wash twice by adding 1 2 mL of buffer per 10 cells i e 10 20 times the incubation volume after incubation with primary antibody Centrifuge and remove supernatant completely preferably with a pipette or by vacuum The concentration of the primary antibody is too low Therefore magnetic labeling is not sufficient to retain the desired cells on 10 Troubleshooting Note Generally MACS Fluorochrome conjugated Antibodies are used in a 1 11 dilution resuspend up to 107 cells in 100 uL of buffer and add 10 uL of MACS Fluorochrome conjugated Antibodies For details or exceptions refer to the MACS Reagent data sheet If other
73. as Risiko eines Stromschlags Um diese Gefahr zu minimieren ffnen Sie weder das Geh use des autoMACS Pro Separators noch anderes Zubeh r Alle Abdeckungen sowie das Ger te Zubeh r d rfen nur von geschultem Personal entfernt werden Besondere Vorsicht ist geboten beim Umgang mit Fl ssigkeiten Beseitigen Sie ausgetretene oder versch ttete Fl ssigkeit sofort Es darf unter keinen Umst nden Fl ssigkeit in das Innere des Ger tes eindringen Ziehen Sie den Netzstecker vor manueller Reinigung des autoMACS Pro Separators Eine potentielle Gefahrenquelle liegt auch im Betrieb eines ge ff neten zu Boden gefallenen oder besch digten Ger tes Ebenfalls sollte unbedingt vermieden werden dass Fl ssigkeit in das Ger t gelangt Fremdk rper durch die Bel ftungs ffnungen eindringen oder von au en in das Instrument hinein gelangen Bei Auftreten von Flammen oder Rauchentwicklung schalten Sie das Ger t sofort aus trennen es von der Stromzufuhr und kontaktieren einen autorisierten Miltenyi Biotec Service Provider oder das Miltenyi Biotec Customer Support Team Der Betrieb eines besch digten Ger tes oder eines Ger tes mit schadhaftem Stromkabel ist ausdr cklich verboten 1 3 2 Starkes Magnetfeld WARNING Extremely strong magnetic fields Risk of severe personal injury to persons carrying pacemakers or electronical medical implants stay clear Keep all magnetically sensitive objects away from device WARNUNG Der autoMACS P
74. automated arm The guiding of the MiniSampler is directly attached to the corresponding slot below the washing station When attaching the MiniSampler sensor cable to the corresponding socket at the rear of the instrument the MiniSam pler will be automatically detected The type of tube rack carried by the MiniSampler is automatically recognized by the rack detector after starting the separation process During operation the tube rack should be covered with the MiniSampler lid that is connected to the lid guiding figure 11 5 The MiniSampler can be disconnected from the autoMACS Pro Separator by pulling is up on the front side and pulling it towards the user Figure 11 5 Rear view of MACS MiniSampler with MACS Reagent Rack and Chill Rack 5 The cell separation unit The central part of the cell processing unit consists of a magnet figure 11 6 and two autoMACS Columns figure 11 6 Once installed the autoMACS Columns become part of a closed fluidic system and can be used for up to two weeks OR 100 separations whichever comes first Fluids are put through the fluidic system with the help of a syringe pump and five valves 11 Technical data and specifications 195 Figure 11 6 Front of autoMACS Pro Separator with access and bottom covers removed 2D code reader barcode reader The autoMACS Pro Separator is equipped with a 2D code reader that uses lasers and powerful light emitting diodes LEDs for illuminating the reading are
75. available R1 R2 R3 and R4 E Figure 6 2 Position R1 was selected for the reagent vial 2 Activate the reader by selecting Read Reagent and present a reagent vial in front of the 2D code reader Ensure the 2D code is facing the blinking code reader light The optimal reading distance is 0 5 2 5 cm from the code reader cover tilt the vial as depicted in figure 6 3 Figure 6 3 Scanning a reagent vial using the 2D code reader In this example the vial CD4 Microbeads human was scanned 3 The vial is automatically recognized by the software The next reagent rack position will be automatically highlighted R2 6 Cell separation using the autoMACS Pro Separator 123 124 Note If the reagent vial cannot be identified by the 2D code reader please enter the reagent information manually refer to Entering reagents manually below ici Figure 6 4 CD4 MicroBeads human 130 045 101 was identified by the 2D code reader software and assigned to the reagent rack position R1 4 Using the same procedure another reagent vial can be scanned using the 2D code reader The reagent rack position R3 is automatically assigned Having entered the desired reagent s click the Separation tab to proceed with programming a cell separation Entering reagents manually 1 Select Reagent tab and highlight the position where the vial will be placed on the reagent rack Four positions are available R1 R2 R
76. azioni relative all installazione della strumentazione contattare le autorit locali competenti per la fornitura di energia elettrica in materia di edilizia manutenzione o sicurezza Qualora abbiate serie preoccupazioni in merito all impiego sicuro dello strumento contattare il rivenditore Miltenyi Biotec autorizzato o chiamare il servizio di assistenza clienti Miltenyi Biotec 1 3 Precauzioni generali Per ridurre i potenziali rischi associati all utilizzo di autoMACS Pro Separator osservare le precauzioni generali di seguito elencate Il mancato rispetto di tali precauzioni pu comportare il rischio di incendi lesioni personali e o danni allo strumento 1 Important information 53 1 3 1 Pericolo di scossa elettrica e di incendio WARNING Hazardous voltages Risk of loss of live severe personal injury Unplug before cleaning Do not use if device is opened or damaged if liquids have been spilled into device if objects device through ventilation slots ATTENZIONE dispositivi elettrici presentano il rischio di scosse elettriche Per ridurre i rischi di scossa elettrica non aprire alcun coperchio tranne i coperchi di accesso anteriore di autoMACS Pro Separator n altri accessori hardware forniti da Miltenyi Biotec Tutti gli altri coperchi dello strumento e gli accessori hardware possono essere rimossi esclusivamente da personale autorizzato Prestare particolare attenzione quando si manipolano liquidi Pulire
77. beling and manual labeling Alternatively each sample could be programmed individually 2 Select CD4_MB human under the submenu Labeling By default the software selects a Possel positive cell target cell enrichment separation strategy and a Qrinse quick rinse wash protocol RE lt 5 Figure 6 23 The current display indicates that autolabeling will be performed on both samples 3 Select the Volume submenu to enter the required sample volume Select Enter For this experiment a minimal starting volume of 160 uL is required for autolabeling for CD4 autolabeling 40 uL MicroBeads is added to the entire 160 uL sample volume Refer to the data sheet if this information is unknown Saerple Vohime 11 14 lt Figure 6 24 A volume of 160 uL was entered for both sample positions 6 Cell separation using the autoMACS Pro Separator 4 Highlight the wash mode to be performed before cell separation 6 In this case a quick rinse Qrinse was selected u Deselect sample position 2 by touching the display at this position Sample position 1 must be setup to perform cell separation on a sample that was previously manually labeled with CD4 MicroBeads Note Both samples positions have been setup for autolabeling Note Sleep O can only selected as the last step in the program N o sequence Figure 6 25 Only sample position 1 is selected 6 Deselect autolabeling for sample 1 by selecting
78. bration 2 by selecting program Calibr_2 4 Select Calibrate The calibration is performed automatically 5 Press Save to finish Calibr_2 12 Hardware and software installation 207 208 Note Carefully read the chapter 1 before installing the 2D code reader If the autoMACS Pro Separator is not provided with the optional 2D code reader an autolabeling package is required for installation Please contact Miltenyi Biotec or your local distributor for further information The autolabeling package contains 1x 2D code reader 1x MACS Reagent Rack 4 1x Autolabeling Software Tools for installation a 2 mm allen key a headless screw an optical frame for the 2D code reader window in the washing station cover and a screw driver for connecting the reader to the outlet at the rear of the instrument 12 3 Installation of the 2D code reader barcode reader Figure 12 14 Contents of the autoMACS Pro Separator upgrade kit 1 Switch OFF and unplug instrument 2 Open and remove the washing station cover Figure 12 15 Removing the washing station cover 3 Remove the peristaltic pump tube at the right hand side of the washing station Figure 12 16 Remove the peristaltic pump tube 4 Unscrew the thumb screw Pull to remove washing station from instrument Take care to clean spilled fluids with ethanol or disinfectant Remove the washing station 12 Hardware and software installation 12
79. cell separation For further information on sample preparation refer to section 6 1 7 Maintenance 145 Note Do not use a dishwasher to clean and do not autoclave any of the removable parts unless indicated otherwise Note In case of malfunction during cell separation this feature must be disabled in order to perform a cell rescue procedure The instrument automatically prompts the user to perform a rinse before performing a cell separation 146 7 2 Daily maintenance The following maintenance procedures must be performed ona daily basis If the instrument is not to be used for an extended period of time the store procedure must be performed refer to 7 2 3 for further details 7 2 1 Instrument priming A Rinse program must be performed before performing the first cell separation after the instrument has been switched on Performing an instrument rinse 1 Click the Separation menu and Wash Now from lower menu bar 2 Select Rinse O 3 ClickRun O_init Optional priming of the instrument at startup By using the O_init program the instrument can be instructed to perform an automated rinse sequence at startup The Rinse program is used for efficient washing and equilibration of the fluidic system 1 Select Option User settings and O_init Press Run 2 Follow the prompt on the screen to enable or disable the initial wash hes news Wine a daba Midi Figure 7 1 Initial wash program is used to activate
80. cells was non specific In order to block non specific binding use MACS FcR Blocking Reagent human 130 059 901 or FcR Blocking Reagent mouse 130 092 575 before labeling 2 The concentration of MicroBeads and or the temperature used for magnetic labeling were too high It is recommended to dilute the MicroBeads according to the data sheet Cells should be incubated with MicroBeads in the refrigerator 2 8 C 3 Incubation time with MicroBeads was too long leading to background labeling Reduce the time for incubation to the recommended values refer to the respective MicroBeads data sheet Typically an incubation time of 15 minutes is required at 2 8 C For exceptions refer to the respective MACS Cell Separation Reagent data sheets 4 Dead cells in the cell suspension may bind non specifically to MicroBeads and will then be co enriched in the positive fraction Remove dead cells before separation by using the MACS Dead Cell Removal Kit 130 090 101 or by Ficoll Paque density gradient 5 Incubation temperatures were higher than recommended typically 2 8 C leading to non specific binding 6 Inadequately stringent separation programs were chosen e g DepleteS instead of Deplete Please refer to the respective data sheet for the recommended program Especially when using autolabelling 1 Check that the recommended maximal sample processing time of 90 minutes was not exceeded If the room temperature is too high t
81. cessed If hazardous material has been used or spilled care must been taken to thoroughly decontaminate the system For details refer to section 7 3 4 Always inspect the fluidics system complete tubing set bottles and their closures valves columns diluter and needles before switching on the instrument If leakage has been detected replace all damaged 1 Important information 23 24 parts before switching on the instrument If damaged parts cannot be replaced unplug and do not use the instrument Failure of parts containing biohazardous material or liquids that have been in contact with such material could cause a hazard Columns tubes and any other consumables that were in contact with biohazardous samples shall be autoclaved prior to disposal Liquid waste shall be autoclaved or decontaminated using a disinfectant that is appropriate for the specific pathogen e g 10 bleach isopropyl alcohol or 70 ethanol Waste disposal must be in accordance with any local regulations Figure 1 4 Warning signs for biohazard located on lower facing panel of the autoMACS Pro Separator left and top of autoMACS Pro Separator fluid bottle right Flammable 70 ethanol is used in Sleep and Store programs The solvent is flammable Therefore keep the instrument away from fire 1 5 5 Transport The autoMACS Pro Separator should be transported with care in packaging specified by Miltenyi Biotec Internal damage can occur if it
82. cia t cnica si desea desprenderse de su instrumento 1 5 7 Compatibilidad electromagn tica S lo para EEUU Los cambios y modificaciones del equipo que no est n expresamente aprobados por Miltenyi Biotec pueden invalidar su autorizaci n para operarlo conforme a la parte 15 del t tulo 47 del C digo de Regulaciones Federales CFR 1 Important information 73 74 1 Important information Introduction 2 2 1 MACS Technology the gold standard in cell separation MACS Technology has become the standard method in cell separation It is based on the use of MACS MicroBeads MACS Columns and MACS Separators strong permanent magnets In a first step surface antigens are magnetically labeled in a highly specific manner with monoclonal antibodies coupled to MACS MicroBeads After magnetic labeling the cells are passed over a MACS Column placed in a MACS Separator Non labeled cells flow through and can be collected as fractions labeled cells are retained in the column and can be released after removing the column from the magnet Thus both labeled and non labeled target cells can efficiently be isolated with MACS Technology The entire procedure is fast easy to handle and gentle to cells leading to the purification of viable and functionally active cells that can immediately be used for further experiments 2 2 Super paramagnetic MACS MicroBeads MACS MicroBeads are super paramagnetic particles of approximately 5
83. ck bar code on rack or try different rack Make sure ambient or direct sun light does not hit sensor Check if syringe is fastened correctly Exchange if broken or damaged If error is displayed again call Technical Support Restart instrument Please call Technical Support if error is thrown again Restart instrument and try a Qrinse to wash out clogged material If error persists run Safe program Otherwise exchange diluter valve or standard valves depending on where a constriction is suspected Please call Technical Support if error is displayed again Make sure foam on top of sample is not higher than 5 mm above liquid level Verify that sample was filtered before separation and that is uptake needle is not clogged Check for leakage at point the where the needle and tubing meet and where the tubing to bubble sensor meet Verify that connectors are appropriately fastened Check for leakage or air bubbles in tubing from uptake needle to valve 1 Verify that connectors are appropriately fastened Calibrate needle arm Check for buffer supply Otherwise call Technical Support Provide all necessary reagents in reagent rack und correct reagent rack definition Try reinitialization by touching RETRY Otherwise call Technical Support Error code 7002 7003 7004 7005 7006 7007 7008 7009 Cause Collision of the needle with the bottom of the tube or any other object has been detecte
84. connect the outlet tubing from the waste cap and put the end of the tubing in a 50 mL tube The rescued cell sample will be rinsed out at the waste port 10 Troubleshooting 3 Select the Separation menu and Wash now Select Qrinse and 1 O press Run The cells will be rinsed out with Running Buffer 4 Reconnect waste closure and waste bottle 5 Perform the Rinse program to prime the autoMACS Pro Separa tor 6 The rescued cell sample can now be centrifuged and subse quently be reprocessed with the autoMACS Pro Separator 10 2 Performance errors not indicated by a warning or error screen 10 2 1 Output volumes are not correct 1 Check for air inlet and leakages in the fluidic system by running the Rinse program From the Separation menu select Wash Now lower menu bar Highlight Rinse and click Run 2 Ifthe tubing is leaking go to section 10 1 2 3 Ifthe pump syringe is leaking go to section 10 1 3 4 Ifthe column is leaking go to section 10 1 1 5 If one or more valves are leaking go to section 7 4 1 6 Check whether all tubing connections including column connections bottle connections and connections at the pump syringe are fastened Loose connections may allow air to enter the system and therefore affect the performance 7 If the problem persists run the Calibr_2 program From the Option menu select User settings e Highlight Calibr_2 and click Run If the problem persists contact Miltenyi Biotec technical su
85. container Container for waste fluid The closure is equipped with a fluid sensor The closure the fluid sensor cable and the tubing connector are color coded red Whole Blood MicroBeads MACS MicroBeads developed for isolating target cells directly from whole blood by using the autoMACS Pro Separator Wrench Black wrench used to tighten and loosen tubing connections 15 Glossary 221
86. ctor 2D Code Esto puede causar errores de lectura Aseg rese de parar la emisi n del l ser antes de limpiar el esc ner De no ser as la exposici n al l ser puede ocasionar lesiones oculares Utilizar un pa o suave y seco para limpiar cualquier sustancia del esc ner No utilizar alcohol u otra sustancia de limpieza El autoMACS Pro Separator est clasificado como producto l ser 1M de est ndar IEC 60825 1 1993 A1 1997 A2 2001 ATENCI N El uso de controles el ajuste o la realizaci n de procedimien tos con excepci n de los aqu especificados puede dar lugar a una peligrosa exposici n de radiaci n 1 Important information 1 4 Instalaci n segura El presente capitulo describe los requisitos que debe reunir el lugar donde desea instalar y operar el separador autoMACS Pro Lea las instrucciones de este capitulo y asegurese de que el emplazamiento esta correctamente preparado antes de conectar el instrumento a la corriente el ctrica Cuando planee la distribuci n del emplazamiento y la ubicaci n del equipo tenga en cuenta las precauciones recogidas en este capitulo para evitar averias del instrumento y reducir la posibilidad del cierre de la empresa por razones medioambientales IMPORTANTE En todo momento se debe observar la normativa nacional de seguridad en el trabajo las normas del laboratorio y los est ndares de salud y seguridad en el laboratorio y de prevenci n de accidentes 1 4 1 Accesorios de
87. d but resetting the collision sensor failed although the needle has been lifted Collision of the needle with an unexpected object Resetting the collision sensor failed as the needle could not be lifted already too close to top Collision of the needle with an unexpected object far above expected tube bottom Collision of the needle with an unexpected object far above expected tube bottom Collision of the needle with an unexpected object far above expected tube bottom Sample volume exceeds maximum volume specified for rack or program type Remain ing portion of sample will not be processed The sample volume has been underestimated The needle has been rinsed but might still be contaminated Collision of needle with bottom of tube could not be detected at expected position 10 Troubleshooting Possible remedies 1 0 Push back needle to it s proper position in the needle holder check for smooth running of the needle in its support then touch CONTINUE Otherwise calibrate needle arm especially if using a Chill 15 Rack Check for objects hindering the movement of the needle Remove the uptake needle from the needle holder and verify that there are no physical obstructions Press CONTINUE Check that cover of the washing station is properly closed Remove any objects hindering the movement of the needle If needle hits rim of tube or top of rack calibrate needle arm using Calibr_1 program Touch LIFT
88. d from the magnet Positive selection Process of isolating cells by labeling the target cells with MACS MicroBeads and performing a MACS Separation The labeled target cells are eluted in row C of the tube rack Possel Positive selection program in standard mode using one autoMACS Column The target cells are eluted in row C of the sample rack This program is used for cells with normal to high frequency and with normal antigen expression 15 Glossary 219 220 Possel_s Positive selection program sensitive mode using one autoMACS Column The target cells are eluted in row C of the sample rack This program is used for cells with normal to high frequency which weakly express the antigens used for magnetic labeling or to achieve the highest recovery of the target cells Posseld Positive selection program normal mode using both autoMACS Columns The target cells are eluted in row C of the sample rack in a volume of 0 5 mL This program is used to isolate rare cells or to achieve a higher purity of the target cells Posseld2 Special positive selection program normal mode using both autoMACS Columns The target cells are eluted in row C of the sample rack in a volume of 2 mL This program is used to isolate rare cells from whole blood cord blood or large cell samples and to achieve a higher recovery of the target cells Posselds Positive selection program sensitive mode using both autoMACS Columns The target cells are elute
89. d in row C of the sample rack in a volume of 2 mL This program is used to isolate rare cells that weakly express antigens used for magnetic labeling e g CD133 Posselwb Special positive selection program using both autoMACS Columns for the isolation of cell subsets from of whole blood The target cells are eluted in row C of the sample rack Running Buffer bottle Container for Running Buffer The bottle closure is equipped with a fluid sensor The closure the fluid sensor cable and the tubing connector are color coded blue Safe solution Solution of 1 w v sodium hypochlorite in distilled water used to decontaminate the autoMACS Pro fluidic system with the Safe program The safe solution is fed into the system from a 50 mL tube Upon completion of the Safe program the fluidic system contains Running Buffer Storage solution Solvent used during the Sleep Store Safe and Column Exchange programs to minimize the contamination risk The fluidic system of the autoMACS Pro Instrument is filled with MACS Storage Solution 70 ethanol Storage solution bottle Container for 70 ethanol The bottle closure the fluid sensor cable and the tubing connector are color coded black Store program Prior to long term storage the Store program should be applied During this procedure the autoMACS Columns are replaced with substitutes Upon completion of the Store program the fluidic system contains 70 ethanol 15 Glossary Syringe
90. data sheet Large cell aggregates may interfere with the separation process and may cause pressure variations in the autoMACS Pro fluidic system It is recommended to use Pre Separation Filters 130 041 407 to remove cell clumps that may clog the column 6 Cell separation using the autoMACS Pro Separator Dead cells and cell debris may bind non specifically to MACS 6 MicroBeads antibodies and antibody conjugates To remove dead cells it is recommended to use density gradient centrifugation or the Dead Cell Removal Kit 130 090 101 For specific recommendations please refer to the general protocols in the corresponding section of this user manual or to the respective MACS Cell Separation Reagent data sheet 6 2 2 Reagent volumes and labeling volumes must be adjusted to the total cell number Typically 107 cells are labeled in a total volume of 100 uL When working with higher cell numbers scale up all reagent volumes and total volumes accordingly For example for 2x107 total cells use twice the volume of all indicated reagent volumes and total volumes indicated in the respective data sheet When working with fewer a than 107 cells do NOT scale down the volume use the same volume as indicated For specific recommendations please refer to the respective MACS Cell Separation Reagent data sheet For details concerning sample volumes required for the first autolabeling step refer to table 6 1 below MACS Product Strategy Rea
91. den autoMACS Pro Separator um eine ausreichende K hlung zu gew hrleisten Vermeiden Sie das Ger t direkter Sonneneinstrahlung auszusetzen ffnungen und Schlitze am Ger t sind zur Bel ftung gedacht und sollten niemals bedeckt oder blockiert werden da sie das Ger t vor berhitzung sch tzen und sicheren Betrieb erm glichen F hren Sie niemals Fremdk rper durch ffnungen in das Ger t ein 1 4 3 Wassereintritt und N sse Setzen Sie das Instrument niemals in einer feuchten oder nasskalten Umgebung ein Setzen Sie das Ger t nie hoher Feuchtigkeit oder Kondensation aus und sch tzen Sie es gegen Spritzwasser 1 4 4 Geerdetes Produkt Das Ger t ist ausgestattet mit einem Schutzkontaktstecker und einer dreiadrigen Anschlussleitung F hren Sie den Stecker nicht in eine Steckdose ohne Schutzkontakt ein Wenn Ihre Steckdose keinen Schutzkontakt aufweist bitten Sie einen ortsans ssigen Elektriker diese zu ersetzen 1 4 5 Stromquellen Das Ger t sollte nur von einer Stromquelle aus betrieben werden die den elektrischen Angaben auf dem Typschild entsprechen Sollten Sie Fragen zur Art der Stromversorgung haben wenden Sie sich an einen autorisierten Miltenyi Biotec Service Provider oder Ihren lokalen Stromversorger Benutzen Sie keine Verl ngerungskabel oder Steckdosenleiste berladen Sie eine Steckdose nicht Die Gesamtlast darf 80 der Zweigstromkreisbemessung nicht berschreiten 1 4 6 Zug nglichkeit der Stromverbindungen
92. duct Monocyte Isolation Kit II human 130 091 153 R4 CD4 MicroBeads human 130 045 101 To delete an individual reagent from the reagent rack list 1 Select the reagent name that must be deleted In this example CD4 MicroBeads human on position R4 must be removed 2 Select Delete Selected to remove the highlighted reagent DRE O 2 Figure 6 9 CD4 MicroBeads human was assigned to reagent rack position R4 which was subsequently selected for deletion To delete the entire reagent list 1 Select an unassigned position on the reagent rack In this example position R4 was selected If there are no unassigned positions delete an individual reagent as described above 6 Cell separation using the autoMACS Pro Separator E 0 22 Figure 6 10 To delete all reagents from the reagent rack an unassigned position on the reagent rack must be highlighted In this example the only unassigned position R4 was selected 2 Select Delete All Working with reagent templates For convenience it is possible to load and save reagent templates In this example the following reagents were scanned into the following reagent rack positions R1 CD4 MicroBeads human 130 045 101 R2 CD8a Ly 2 MicroBeads mouse 130 049 401 R3 No reagent R4 No reagent To save a reagent template 1 Assign reagent vials to reagent rack positions as outlined above refer to section Entering reagent information for autolabeling
93. e Air Filter Connector autoMACS Pro Bubble Sensor autoMACS Pro Tube t2 autoMACS Pro Tube t21 autoMACS Pro Tube t27 autoMACS Pro Tube t5 autoMACS Pro Tube t7 autoMACS Pro Tube reservoir t9 t24 autoMACS Pro Uptake Port Needle Column Connector Fuse Holder Hydrophobic Air Filter O Ring for Bottle Closure Plunger Lock Screw Power Cord D Power Cord USA Pump Syringe Part order number 130 093 307 130 093 308 130 093 309 130 093 310 130 093 311 130 093 312 130 093 313 130 093 314 130 093 315 130 093 316 130 093 317 130 093 349 130 093 350 130 093 362 130 093 364 130 093 365 130 093 366 130 093 367 130 093 368 130 093 370 Spare part autoMACS Pro Bottle Closure red Dilutor Valve 6 port distribution autoMACS Pro Tube t32 autoMACS Pro Tube t26 autoMACS Pro Tube t20 autoMACS Pro Tube t12 114 autoMACS Pro Tube t3 autoMACS Pro Tube t18 autoMACS Pro Tube t11 t25 autoMACS Pro Tube t6 t8 t23 autoMACS Pro Tube t10 t16 t17 t19 t22 Washing Station needle arm autoMACS Pro Tube t1 t13 Tube Connector 2x2 Ports 1 4 28 UNF Tube Guiding Ball needle arm Peristaltic Pump Head incl Tube Outlet Port Unit needle arm Guard needle arm Thumb Screw M5x16 Magnet Lock front cover Part order number 130 090 378 130 091 996 130 093 293 130 09
94. e autoMACS Pro Separator Starting Kit and pull out the valve de SE M 6 Check ifthe groove in the valve drive is positioned horizontally and in the lower half of the axis Figure 7 15 Valve plate driving section 1 groove 2 hole for adjustment pins 7 Maintenance 155 156 Note In case the valve exchange program Valve_ex cannot be performed e g valve does not turn any more switch off instrument and turn the valve bracket manually to the position corresponding to the groove in the valve drive 7 Make sure that the bracket of the new valve is positioned horizontally Figure 7 16 Rear view of the autoMACS Pro Separator valve 1 bracket 2 8 adjustment pins Carefully insert the new valve allowing the bracket to find the groove in the drive At first the bracket will slide in only halfway Figure 7 17 Inserting the new valve 9 10 11 12 13 15 Gently rotate the valve The two adjustment pins will slide into their corresponding holes in the valve plate Make sure that the valve is fully inserted into the driving station Fasten valve screws using the screwdriver Connect the tubing with the installed new valve and fasten tubing by hand until finger tight Plug in and switch ON the autoMACS Pro Separator Perform a Rinse program and check the valves visually for leakage and air inlet Refasten tubing connectors using caution If leakage persists call Tec
95. e das Ger t in Betrieb nehmen Ber cksichtigen Sie bei der Aufstellungsplanung die in diesem Abschnitt beschriebenen Vorkehrungen um Ger test rungen zu vermeiden und die Wahrscheinlichkeit umgebungsbedinget Ger teausf lle zu verringern WICHTIG rtliche Arbeitsschutzbestimmungen Laborrichtlinien Sicherheitsnormen und Unfallverh tungsvorschriften m ssen auf jeden Fall beachtet werden 1 41 Montagezubehor Stellen Sie den autoMACS Pro Separator niemals auf einen instabilen Labortisch Laborwagen Untersatz Laborstativ oder Laborkonsole Als Folge davon k nnte das Instrument zu Boden fallen dadurch k nnten schwere Verletzungen und oder Sch den am Ger t verursacht werden Benutzen Sie daher nur von Miltenyi Biotec empfohlenes oder direkt mit dem Ger t erh ltliches Labormobiliar Versuchen Sie nicht den autoMACS Pro Separator in eine Einbaukon struktion zu integrieren oder auf begrenztem Raum z B in einem Laborregal unterzubringen es sei denn es wurde eine spezielle Vorrichtung daf r entwickelt es ist f r ausreichende Bel ftung gesorgt und die Montageanleitung f r das Ger t wurde befolgt 1 Important information 45 46 1 4 2 Bel ftung Das Instrument sollte nicht in der N he von Radiatoren Hei l fter fen oder anderen W rme erzeugenden Ger ten stehen Verst rker eingeschlossen Erm glichen Sie im Betrieb eine ausreichende Luftzirkulation im Abstand von mindestens 15 cm in allen Richtun gen um
96. e not or only partly taken up Reagent vial runs dry Magnetic labeling and separation Positive selection Depletion Indirect MACS MicroBeads Fluorescent staining Cells are poorly stained Cells are excessively stained Contamination of tubing system 1 11 186 Chapter 11 193 193 197 199 Chapter 12 201 201 201 201 208 Chapter 13 213 Chapter 14 215 Chapter 15 217 1 Important information 10 6 11 12 12 1 12 1 1 12 2 12 3 13 14 15 Problems indicated by error or warning screens Technical data and specifications Labeled schematics of the autoMACS Pro Separator Technical data and specifications of the autoMACS Pro Separator Technical data and specifications of the MACS MiniSampler Hardware and software installation Installation of new software Exchanging the software cards Calibration of the autoMACS Pro Separator Installation of the 2D code reader barcode reader Technical support Limited warranty Glossary 1 Important information 13 Thank you for choosing a Miltenyi Biotec product The autoMACS Pro Separator is an innovative instrument for automated sensor controlled multisample labeling and separation of various cells types At the touch of a button target cells are magnetically labeled separated and eluted in a fully automated fashion Purify and progress Important information 1 Please read before use Please read all
97. e specifically indicated in the MACS Cell Separation Reagent data sheet the magnetically labeled cell samples are suspended at 10 total cells per 500 uL when using manual labeling The cell number and volumes for autolabeling are slightly different refer to table 6 4 and table 6 5 below for further details 1 Select the appropriate tube rack according to table 6 4 and table 6 5 2 Recommended Cool down the tube rack for 3 4 hours in a refrigerator 2 8 C or until the coolant becomes solid Do not cool below 0 C as samples may freeze 3 Equip the tube rack with sample tubes and fraction collection tubes 6 Cell separation using the autoMACS Pro Separator 133 134 Note Row A of the tube rack holds sample tubes row B tubes for non labeled fractions row C tubes for labeled fractions Racktype amp Slots symbol Chill 5 24x5 mL Chill 15 15x15 mL 5x5 mL Chill 50 6x50 mL 3x15 mL 3x5 mL Rack type amp symbol Chill 5 24x5 mL Chill 15 15x15 mL 5x5 mL Chill 50 6x50 mL 3x15 mL 3x5 mL Maximum Minimum numberof first samples incubation volume 6 0 2 mL 5 mL tubes 0 25 mL 5 0 2 mL 15 mL tubes 1mL 3 4mL 50 mL tubes Maximum final labeling volume 2 0 mL 1mL 6 5 mL 4mL 8 mL Maximum number of cells per tube 2 0 4x108 depends on cell labeling concentration and column capacity Whole blood only 6 5x108 1 3x10 depends on cell lab
98. e valve to the exchange position for removal Valves may require periodical exchange Refer to section 7 4 for more details 4 An overview of the autoMACS Pro touchscreen user interface 111 112 4 An overview of the autoMACS Pro touchscreen user interface Materials required o This chapter describes the consumables and accessories required for the operation of your autoMACS Pro Separator IMPORTANT Please be advised that the autoMACS Pro Separator is CE specified for use with MACS MicroBeads autoMACS Columns and other genuine Miltenyi Biotec consumables and accessories only Please only use consumables and accessories recommended by Miltenyi Biotec Failure to use recommended consumables and accessories may result in inaccurate results instrument malfunction or damage premature wear and reduced life time of the instrument Miltenyi Biotec does not honor any warranty or accept any responsibility for damages resulting from the use of inappropriate consumables or accessories 5 1 Materials required for operation 5 1 1 Solutions For daily operation the following solutions are required Running Buffer Washing Solution and ethanol Solution Description Color Capacity Order no code Running autoMACS Blue 6x1 5 L for 6x15 130 091 221 Buffer Running Buffer or separations and MACS Separation rinsing cycles Buffer Washing autoMACS Pro Green 6x1 5 Lfor 6x15 130 092 987 Solution Washing Solution rinsing cycles Rinse Sleep Sto
99. ead of negative value for current diluter move Touch CANCEL to abort Please contact the Technical Support in all cases also if the sample has been processed completely after touching RETRY Technical data and specifications 11 11 1 Labeled schematics of the autoMACS Pro Separator Integrated computer for control of cell processing All interactions with the computer are performed with a TFT color touchscreen figure 11 1 A memory card is used to run all programs and to log processes Figure 11 1 Front view of autoMACS Pro Separator Automated arm with ports for sample uptake and release of cell fractions The automated arm figure 11 1 is a computer controlled part of the autoMACS Pro Separator It holds two ports one for sample uptake and release of the magnetically labeled positive cell fraction and one for release of the non labeled negative cell fraction The automated arm moves in y and z directions The ports are automatically washed in the autoMACS Pro Washing Station during and after the separation process to prevent cross contamination between samples Access covers The front door figure 11 1 is opened sideways to allow access to the parts of the fluidic system that require periodic maintenance by the user This includes the autoMACS Columns pump syringe and upper valves The bottom cover figure 11 1 gives access to the lower valves and can be removed by pulling gently The washing station cover figure 11
100. eck the exchange date of the autoMACS Columns Using the autoMACS Columns for longer than two weeks or for more than 100 separations within two weeks may affect purity Check whether the autoMACS Pro Separator has been set to Sleep Store correctly Inappropriate storage or allowing the autoMACS Pro Separator fluidic system to run dry will affect the columns and therefore the purity Make sure that the appropriate Running Buffer has been used Perform the Qrinse program If pumps are filled with air during the process refer to section 10 1 4 Perform a test run using PBS as a mock sample and check output volumes For correct volumes refer to table 6 2 in section 6 3 3 If volumes are not correct go to section 10 2 1 Run the Safe program refer to section 4 7 If the problem persists contact Technical Support 10 Troubleshooting When utilizing the autolabeling feature 1 O 1 Check whether the uptake needle is positioned in the middle of the sample tube during the mixing procedure If not mixing will be inadequate 2 Check whether the reagent volume contents of reagent vial is sufficient for labeling 3 Check that the recommended maximal sample processing time of 90 minutes was not exceeded In the event of an elevated room temperature the cooling capacity of the Chill Racks may be compromised Higher labeling temperatures could cause non specific binding which may lead to lower purities when using positive selection kits
101. ect all tubing at the appropriate positions and reposition up take needle in needle holder 7 7 2 Rescue procedure B If it is not possible to restart the instrument the cells retained on the columns can be recovered 1 Switch OFF the instrument and disconnect from the power supply 2 Prepare two 50 mL tubes in rack and fill two 5 mL syringes with Running Buffer Separation Buffer 3 Open the front cover and place absorbent tissue underneath the columns 4 Pull out the column from the column holder and replace the top connector with a5 mL syringe filled with Running Buffer 5 Undo the bottom connector and flush the column into a 50 mL tube Discard column and syringe appropriately 6 Repeat steps 4 to 5 with the second column 7 Centrifuge tubes at 350xg for 10 minutes 8 Discard the supernatant and perform reseparation on the recovered cells as soon as possible Keep cells on ice until the separation 7 Maintenance 163 164 9 Install new columns or column substitutes in place of the discarded ones 7 8 Spare parts list Part order number 130 022 101 130 090 684 130 090 685 130 090 387 130 093 291 130 093 285 130 093 287 130 093 284 130 093 288 130 093 286 130 093 289 130 093 290 130 090 676 130 090 834 130 090 385 130 090 386 130 090 644 130 090 389 130 090 391 130 090 339 7 Maintenance Spare part Pump Seal 4 port 4 way Valve 4 port Distribution Valv
102. ect has been dropped into the instrument If flames or smoke appear immediately switch off the the autoMACS Pro Separator unplug the instrument from the electrical outlet and contact an authorized Miltenyi Biotec service provider or the Miltenyi Biotec Customer Support team Use of a damaged instrument or an instrument with a damaged power cable is expressly prohibited 1 3 2 Strong magnetic field WARNING Extremely strong magnetic fields Risk of severe personal injury to persons carrying pacemakers or electronical medical implants stay clear Keep all magnetically sensitive objects away from device WARNING The autoMACS Pro Separator is equipped with an extremely powerful magnet Keep any magnetic information carriers such as credit cards magnetic tapes and floppy disks any electronic equipment such as hearing aids pacemakers measuring and control instruments computers and watches and magnetizable tools and objects at a distance of at least 20 cm from the magnet cover These items may be affected or damaged by the magnetic field Figure 1 1 Location of warning sign for strong permanent magnet 1 Important information 1 3 3 Hazard of crushing and shearing 1 CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard CAUTION Do not open the front access covers while the instrument is in operation Do not obstruct the movement of the automated arm
103. ed by system At this point the bottle illumination is yellow 3 Assembly and installation Note When working with biohazardous samples it is recommended to fill the waste container with 100 mL of disinfectant before use e g MACS Bleach Solution For proper disposal please follow local regulations and carefully read the chapter 1 rel Figure 3 23 The status menu The fluid level status and column are green for go A wash or instrument prime has not yet been performed 3 Select menu Separation and Wash Now from the lower menu You now have the option to perform a quick rinse Qrinse or full rinse Rinse Figure 3 24 Select Rinse to prime the autoMACS Pro Separator 4 Select Rinse and Run to start the priming process The progress will be displayed at the bottom of the touchscreen menu 5 When priming is finished the instrument will display Ready for separation in the Status menu The bottles are illuminated green gt Figure 3 25 The instrument has been primed rinsed 3 3 6 Checking the fluid levels The autoMACS Pro Separator automatically ensures that the uptake port is filled with buffer that fluid containers carry enough fluid for 3 Assembly and installation 93 94 Note A cell separation template can already be programmed or reloaded during the priming procedure By selecting Run the user will asked to confirm that enough running buffer is available thereafter pri
104. eed replaced Note 70 ethanol does not contain electrolytes Therefore the fluid level cannot be determined and the symbol for this bottle is always gray Please ensure that the volume of each solution is sufficient for the 4 Ensure that the symbol for the columns is colored green For number of separation and washing programs to be performed Note The autoMACS Pro Separator automatically records the date of the last column exchange and displays the date for column replacement only if the program Column exchange has been used for column installation more details refer to section 4 3 2 Column status If the symbol for the columns is red the columns must be exchanged The fill level on the symbol is an indicator for the remaining operation life of the columns Figure 6 19 The column status graphic Left The columns are ready Right The columns must be changed 6 Cell separation using the autoMACS Pro Separator 135 136 Note The touchscreen displays the type of tube rack upon starting the separation Note If priming of the instrument has not been performed the autoMACS Pro Separator will automatically ask to run a Rinse program before starting the separation The process can be monitored in the menu Status At this point the bottles are illuminated yellow It is possible to include an initial Rinse program that will be performed automatically upon switching on the instrument The
105. el instrumento en un lugar h medo Evite la elevada hume dad o la condensaci n y proteja a la m quina de salpicaduras 1 4 4 Producto conectado a tierra El instrumento est equipado con un tipo de enchufe de tres varillas que tiene un tercer contacto para toma de tierra Este enchufe s lo encaja en una toma conectada a tierra Se trata de una medida de seguridad No intente enchufarlo en una toma de electricidad no conectada a tierra Si el enchufe no encaja p ngase en contacto con un electricista para que reemplace la toma de corriente 1 4 5 Fuentes el ctricas S lo se deber a maniobrar el instrumento utilizando la fuente el ctrica indicada en la etiqueta con los valores el ctricos del producto Si tiene alguna pregunta sobre qu tipo de fuente el ctrica utilizar p ngase en contacto con su proveedor de servicios de Miltenyi Biotec autorizado o con su compa a el ctrica No utilice alargadores o regletas No sobrecargue la toma de corriente La carga total del sistema no debe sobrepasar el 80 de la rama del circuito 1 4 6 Accesibilidad Aseg rese de poder acceder f cilmente tanto al interruptor principal como al conector para el cable de corriente el ctrica y de que stos est n situados tan cerca del operario como sea posible Si es necesario desconectar el suministro el ctrico desenchufe el cable de la toma de corriente 1 4 7 Aparatos perif ricos Solamente los dispositivos perif ricos que cumplen con la UL 60
106. eling concentration and column capacity Whole blood only Whole blood only Table 6 4 MACS Cooling Tube Racks specifications for autolabeling Maximum number of samples 6 5 mL tubes 5 15 mL tubes 3 50 mL tubes Maximum sample volume 2 5mL 12 5 mL 50 mL Maximum number of total cells per tube 5 0x108 2 5x10 4 0x10 Table 6 5 MACS Cooling Tube Racks specifications for manual labeling 6 Cell separation using the autoMACS Pro Separator 6 5 Prime the autoMACS Pro Separator 6 1 Switch on the autoMACS Pro Separator and wait for the instrument to complete initialization Note It is assumed that the MACS MiniSampler has been already installed the waste container is empty and that all fluid containers are filled with recommended solutions refer to section 3 for more details TE Figure 6 17 The initialization screen 2 After initialization the autoMAC Pro Separator will display the menu Status Verify the status of the instrument as outlined in section 4 3 3 Ensure that the symbols for the fluid containers are colored green If they are red check whether the fluid containers are filled the waste container is empty and the color coded bottle sensors are connected to the appropriate container If symbols are gray check the bottle sensors for proper connection Figure 6 18 Fluid container status symbols Left Fluid containers are ready Right Fluid containers n
107. ent should be stable and vibration free dust free sufficiently ventilated and free from sources of electromagnetic radiation Before operating the autoMACS Pro Separator for the first time carefully read the user manual and contact your local Miltenyi Biotec representative for assistance 3 1 Unpacking and installing the autoMACS Pro Separator Read through the following instructions carefully before commencing the installation procedure Before opening the transportation box check for any visible external damage to the box Check also to see if the shock and position indicators if present suggest incorrect transportation of the instrument If there is apparent damage please contact Technical Support for assistance refer to section 13 3 Assembly and installation 79 Note The top compartment holds the autoMACS Pro user manual the short instructions and various bags containing accessories Carefully remove these parts Note Two persons are required to lift the autoMACS Pro Separator The instrument must be gripped at the base of the orange bottle baskets located at both sides of the instrument Note that the instrument is heavier at the front Ensure the front of the instrument is stabilized while lifting it Due care must be taken while lifting the autoMACS Pro Separator Miltenyi Biotec accepts no liability for potential injuries and damages sustained during lifting and or movement of the instrument
108. enu Figure 4 15 Labeling submenu Using the MACS Reagent Rack 4 a total of four reagents may be used at any given time Assigning sample volumes The Volume submenu 3 is used to inform the instrument about the available sample volume To enter or modify a volume 1 Ensure that the appropriate sample template position is highlighted 2 Select the Volume submenu 3 Using the numeric keypad enter the sample volume For autolabeling enter the sample volume for the first labeling step as outlined in the corresponding data sheet e g 160 uL 2x10 cells for labeling with CD4 MicroBeads human or 120 uL 4x10 cells for labeling with Monocyte Isolation Kit II human Figure 4 16 Entering the sample volume in microliters uL for Monocyte Isolation Kit II human 4 Note that the minimum incubation volume after the addition of the first reagents may not be less than 200 uL 5 Select Enter 4 An overview of the autoMACS Pro touchscreen user interface 101 102 Assigning a cell separation program The Separation submenu 4 is used to instruct the autoMACS Pro Separator which cell separation program should be applied to each sample Several sample positions can be highlighted to assign a separation condition for multiple samples Refer to section 6 3 1 fora detailed explanation of the various cell separation strategies To assign or modify a cell separation program 1 Highlight the appropriate sample s usi
109. epls7 3mLper1mLsample 2mLper1 1 mL min volume mL sample volume Possel 2mL samplevolume 2mL 4mL min Possel_s 2mL samplevolume 2mL 1 mL min Posseld 2mL sample volume 0 5 mL 4 mL min column 1 1 mL min column 2 Posseld2 2mL samplevolume 2mL 4mL min columns 1 and 2 Posselds 2mL samplevolume 2mL 1 mL min columns 1 and 2 Posselwb 2mL samplevolume 2mL 4 mL min predilution volume columns 1 and 2 This program must be used with manual labeling only Table 6 2 Output volumes and loading rates of separation programs 6 3 4 autoMACS Pro Separator wash programs and maintenance programs Wash programs for daily operation The autoMACS Pro Separator is equipped with reusable autoMACS Columns After each cell separation a thorough washing procedure rinses the columns of the autoMACS Pro Separator After the wash program is completed columns and tubing system are filled with Running Buffer Program Washing Running Storage MACS Time Solution Buffer solution Bleach Solution Qrinse 48 mL ore gt 1 5 min Rinse 96 mL 48 mL 4 min Sleep 96 mL 48 mL 5 min Store 96 mL 96mL 8 min Col_ex 96 mL 96 mL 6 min Safe 96 mL 96 mL gt 40 mL 21 min Not including the time required for column exchange Not including the time required for disconnecting and reconnecting bottle tubing Table 6 3 Liquid usage and time of wash and maintenance programs 6 Cell separation using the autoMACS Pro Separa
110. er ser manejado dentro de una campana de seguridad si se procesan sustancias peligrosas o desconocidas Si se han utilizado sustancias peligrosas o stas se han derramado se debe velar por una desinfecci n meticu losa del sistema Las columnas placas tubos de ensayo y cualquier otro objeto que haya estado en contacto con las muestras peligrosas deber n ser autoclavados antes de poder volver a ser utilizados Los residuos l quidos deber n ser autoclavados o descontaminados usando un desinfectante industrial apropiado para el pat geno espec fico por ejemplo hipoclorito de sodio al 10 alcohol isoprop lico o etanol al 70 2 Introduction mate ACS Pro Separator B woes hin w Pomar Sa pol Lia RE rd cus use CAN Pas se Figura 1 4 La eliminaci n de los residuos debe cumplir la normativa nacional 1 5 5 Transporte El separador autoMACS Pro debe ser transportado con cuidado en el embalaje especificado por Miltenyi Biotec Se pueden producir da os internos si es expuesto a vibraciones excesivas o si se cae En caso de que sea necesario devolver el instrumento a su fabricante para su revision limpielo y desinf ctelo de cualquier sustancia peligrosa antes de realizar el envio Si tiene preguntas relativas a la descontami naci n o el env o p ngase en contacto con el servicio de asistencia t cnica 1 5 6 Eliminaci n del instrumento equipo P ngase en contacto con el servicio de asisten
111. erface 95 96 Tab Function Reagent To define the position of MACS Reagents on the MACS Reagent Rack 4 Separation To define an autolabeling and or cell separation strategy for up to six samples In addition cell processing procedures can be saved as templates for regular use Status The instrument status is displayed at a glance Log list The log list details completed actions and errors Option Users can perform special procedures such as exchange of MACS Columns instrument calibration and service steps Table 4 1 Feature overview of the upper tabbed menu After instrument initialization the screen automatically displays the Status menu which helps to monitor the instrument during installation and operation Color coded symbols indicate the status of the hardware components Further information on a particular component can be obtained by touching the symbol RS 2 Figure 4 4 The main root menu screen displays instrument Status at startup 4 3 Status menu The autoMACS Pro Separator is a sensor controlled instrument that allows easy monitoring during operation At a glance the instrument status can be determined by viewing the Status menu at any time F Sample status 2 Fluid container filling status B Column status 4 Tube rack status 5 MiniSampler status iii Figure 4 5 Overview of the instrument status panels 4 An overview of the autoMACS Pro touchscreen user interface A de
112. es autoMACS Pro Separator darauf dass sich der optische Weg des Laserstrahls beim Betrieb nicht auf gleicher H he mit dem menschli chen Auge befindet Vermeiden Sie dass Wasser l Staub oder andere Fremdk rper dem ffnungsfenster des 2D Code Lesers anhaften Dies kann zu Lesefehlern f hren Stellen Sie vor der Reinigung des Scanners sicher dass keine Laserstrahlen mehr emittiert werden Der Umgang mit dem Laser kann sonst zu Augenverletzungen f hren Verwenden Sie zum Abwischen von Substanzen auf dem Scanner ein weiches trockenes Tuch Verwenden Sie keinen Alkohol oder andere Reini gungssubstanzen Der autoMACS Pro Separator ist nach Norm IEC 60825 1 1993 A1 1997 A2 2001 als Klasse 1M Laserprodukt eingestuft WARNUNG Die Verwendung von anderen als hierin genannten Bedienungselementen sowie die Anpassung oder Durchf hrung von anderen als hier genannten Verfahren kann gef hrliche Strahlung freisetzen 1 Important information Figure 1 3 Position von Lasern Im Bereich des Rechtecks befinden sich verdeckt montierte Laser zur Erkennung von Gestellen Der sichtbare 2D Barcodeleser ist hier mit einem Kreis gekennzeichnet 1 4 Sichere Ger teinstallation In diesem Abschnitt werden die Anforderungen an einen betriebssi cheren Standort f r Ihren autoMACS Pro Separator beschrieben Lesen Sie die Anleitungen in diesem Abschnitt und stellen Sie sicher dass der Ger testandort entsprechend vorbereitet ist bevor Si
113. es fournis par Miltenyi Biotec Tous les caches de l appareil et des quipements accessoires doivent tre d mont s par le personnel agr uniquement Soyez particuli rement prudent pendant la manipulation de fluides Nettoyez imm diatement si des liquides se sont d vers s Veillez ce que les fluides ne s infiltrent pas l int rieur de l appareil D branchez le c ble lectrique avant de nettoyer manuellement le s parateur autoMACS Pro Separator Un risque potentiel existe si le s parateur autoMACS Pro Separator utilis est ouvert d form ou endommag si des liquides se d versent dans l appareil si un objet est entr dans l appareil par les fentes de ventilation ou si un objet a chut dans l appareil En cas d apparition de flammes ou de fum e d connectez imm diatement le s parateur autoMACS Pro Separator d branchez l appareil et contactez un prestataire de services Miltenyi Biotec agr ou l quipe de support technique de Miltenyi Biotec Il est formellement interdit d utiliser un appareil endommag ou un appareil dont le c ble lectrique est endommag 1 3 2 Champ magn tique puissant WARNING Extremely strong magnetic fields Risk of severe personal injury to persons carrying pacemakers or electronical medical implants stay clear Keep all magnetically sensitive objects away from device AVERTISSEMENT Le s parateur autoMACS Pro est quip d un aimant extr mement puissant Veillez l
114. essen Betrieb beeintr chtigt werden Auch k nnte sich vorzeitiger Verschlei einstellen und somit die Lebensdauer des Ger tes verk rzen Hierdurch k nnten Ihre Garantieanspr che verfallen Bewahren Sie dieses Benutzerhandbuch an einem sicheren Ort auf zug nglich f r jeden Anwender der autoMACS Pro Separa tor Dieses Kapitel beschreibt die Sicherheitsvorschriften und die Anforderungen an den Ger testandort ihres autoMACS Pro Separator Die folgenden Warnhinweise und Vorsichtsma regeln sollen Ihnen dabei behilflich sein Verletzungen f r sich oder Sch den am Ger t zu vermeiden 1 1 Symbole und Gefahrenstufen Warnhinweis Beispiel CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard Die Gefahrenhinweise informieren den Anwender ber Gefahren die bei Missachtung der nachstehend beschriebenen Warnungen und Vorsichtsma nahmen bestehen k nnen Das Gefahrensym bol links versinnbildlicht die Gefahr Die Gefahrenwarnung oben bezeichnet die Schwere der vorliegenden Gefahr wie vorstehend beschrieben Schwere Art und Ursprung der Gefahr sowie 1 Important information 39 40 CAUTION WARNING 00 0b gt Db gt gt lll m gliche Folgen Verbote und Vorsichtsma nahmen werden im Anschluss beschrieben Symbole und Gefahrenebenen Im folgenden tabellarischen Glossar werden alle in diesem Benutzer handbuch oder auf dem autoMACS Pro Separator ve
115. fety and prevention of accidents must be adhered to 1 5 1 Safe operation If the instrument is not working properly and instructions or messages on the display screen advise to contact technical service secure operation is no longer possible Immediately switch off the autoMACS Pro Separator unplug the instrument from the electrical outlet and contact an authorized Miltenyi Biotec service provider or the Miltenyi Biotec Customer Support team 1 5 2 Servicing IMPORTANT Unless otherwise specifically noted in this User Manual or other Miltenyi Biotec documentation do not service the autoMACS Pro Separator yourself Servicing and repair must be performed by qualified service personnel Improper or incorrect servicing or repair 1 Important information of your autoMACS Pro Separator can cause hazards to users lead to 1 unpredictable results instrument malfunction or damage premature wear and reduced life time of the instrument and may void your warranty fe Inquire with your local Miltenyi Biotec representative about Miltenyi Biotec s extensive instrument service and support arrangements or refer to www miltenyibiotec com support IMPORTANT When replacement or spare parts are required make sure that the service provider uses only genuine Miltenyi Biotec parts or third party parts specified and recommended by Miltenyi Biotec Using unauthorized replacement or spare parts can cause malfunction of the instrument and impair cell
116. from entering the fluid bottle Air filter connector Luer to thread connector for attaching the air filter to the threaded bottle closure vent APC Allophycocyanin autoMACS Columns Specifically designed autoMACS Columns reusable for two weeks or for 100 separations within these two weeks autoMACS Column 1 First autoMACS Column in which labeled cells are retained during positive selection and depletion programs The autoMACS Column 1 occupies the left slot of the black magnet cover autoMACS Column 2 The second autoMACS Column is used during double selection programs The autoMACS Column 2 occupies the right slot of the black magnet cover autoMACS Pro Separator Automated magnetic cell separator also referred to as device or instrument autoMACS Running Buffer Sterile and ready to use solution for cell separation and washing programs The tubing connector is color coded blue autoMACS Pro Washing Solution Sterile and ready to use solution for washing and special rinsing programs The tubing connector is color coded green Bottle closure Vented screw on closure with fluid uptake ports canules and distribution tubes The bottle closures contain fluid sensors and are equipped with sensor cable connectors Column Connector Luer to thread connector connecting the autoMACS Columns to the fluidic system Column substitute Column without spheres that replace the autoMACS Columns for long term storage and shipment Colum
117. g of the autoMACS Pro Tubing System 70 ethanol This solvent should be prepared by diluting absolute ethanol with distilled water 5 1 2 Hardware MACS MiniSampler The MACS Mini Sampler can be equipped with three different tube racks for details refer to table below To attach the MiniSampler and the tube racks refer to section 3 The autoMACS Pro Separator will automatically detect the MiniSampler and the type of tube rack Tube racks tubes and reagent racks Three tube racks Chill 5 Chill 15 and Chill 50 designed for 5 mL 15 mL and 50 mL tubes respectively and are available for use with the autoMACS Pro Separator All three racks contain a coolant To cool the racks keep them in a refrigerator for 3 4 hours Do not cool below 0 C to avoid freezing of the samples Use 5 mL 15 mL or 50 mL tubes The process has been optimized using BD Falcon tubes Rack type Tubes Maximum number Maximum sample of samples volume Chill 5 5mL 6 2 5mL Chill 15 15mL 5 12 5 mL Chill 50 50mL 3 Up to 50 mL Reagent Rack4 4 vials Not applicable Not applicable Table 5 2 Cooling tube and reagent racks 5 2 Materials required for maintenance 5 2 1 Solutions MACS Bleach Solution MACS Bleach solution 130 093 663 is used in combination with the Safe program Depending on the level of use it is recommended to run a Safe program at least every 3 to 6 months If material like whole blood or tissue is primarily used it is recommended
118. gents Cell Minimal Minimal concentra volume absolute tion cell number Direct MicroBeads Positive selection 1 107 cells per 160 uL 2 0x107 human or depletion 80 uL rat non human primate Direct MicroBeads Positive selection 1 107 cells per 180 uL 2 0x107 mouse or depletion 90 uL Whole Blood Whole blood or 1 Original 0 25 mL MicroBeads bone marrow volume 8mL Cell Isolation Kits Untouched 2 107 cells per 160 uL 4 0x107 selection 40 uL Cell Isolation Kits Untouched 3 107 cells per 120 uL 4 0x107 selection 30 uL MicroBead Kits Positive selection 2 107 cells per 120 uL 2 0x107 or depletion 60 uL When working with fewer cells than the necessary minimal volume resuspend cells in the stipulated minimal volume Table 6 1 Table shows dilution volumes required for the first autolabeling step 6 Cell separation using the autoMACS Pro Separator 119 120 6 3 Select a cell separation strategy and a suitable magnetic labeling strategy The autoMACS Pro Separator can perform fully automated sample processing i e sample autolabeling labeled cell incubation cell separation and target cell elution The software also offers the choice to perform cell separation on manually labeled cells Thus it is possible to perform automated cell separation with autolabeling automated cell separation without autolabeling or automated cell separation using a combination of both strategies 6 31 Choosing a cell separation strateg
119. gs are correct tip is correct if a sheet of paper can be easily slid 18 If Change was selected see above use the Up and Down between the needlle tip navigation buttons to adjust the position as required and the surface of the tube rack Cole ote bottom detections Fs Figure 12 9 If necessary adjust the z axis of the needle tip using Up and Down navigation tools 19 Select OK and Done to save the current configuration Calibration 1 Calibration of the x axis step 4 5 20 Select Calibrate to proceed with calibration 21 Place a MACS Cooling Tube Rack and a MACS Reagent Rack 4 onto the MACS MiniSampler It is recommended to use a Chill 5 Tube Rack 22 Select Done 12 Hardware and software installation 205 Note It is possible that the stored calibration value is zero In this event step 23 will be skipped Continue with step 24 Note Due to mechanical limits of the MiniSampler the correction potential is limited Exceeding these limits is reported by the message standard range exceeded Calibration outside the standard range is not recommended The outer positions reagent vial and sample row 6 will not be reached properly Labeling in sample position 6 will be prohibited In this event contact Technical Support refer to section 13 23 Select Use to start the calibration at current position recommended Select Reset to use factory settings 24 If necessar
120. hazard situation which if not avoided could result in minor or moderate injury Indicates a hazardous situation which if not avoided could result in death or serious injury Attention consult the User Manual for further instructions and proceed with caution Warnings include the risk of damage to the equipment severe personal injury or loss of life Hazard of crushing and shearing Risk of crushing and shearing of bodily parts due to mechanical hazards Laser radiation Risk of serious eye and skin injuries Strong magnetic field The magnetic field can interfere with magnetizable objects and electronic devices or damage magnetic information carriers Risk of severe personal injury to persons carrying pacemakers or electronic medical implants Risk of contamination if biohazardous material is used Indicates the risk of loss of life severe injury to the instrument operator or equipment damage due to potentially dangerous biological material Indicates the risk of loss of life or severe injury to the instrument operator due to hazardous voltage Protective conductor terminal Symbol is attached on the inside of the instrument Warning for service personnel On supply Off supply Documentation needs to be consulted before proceeding with installation and operation of the system 1 Important information 1 2 Warnings and precautions 1 The autoMACS Pro Separator employs state of the art technology It is a co
121. he basket Place then fasten a standard lid the bottle in its appropriate position remove the cap and fasten i e without sensor the bottle closure Note the color coding and connect each cables onto the bottle sensor cable to the respective bottle closure before removing the waster container from 4 Attach the sensor cable plug to the socket for sensor cables at the basket This is to the back of the autoMACS Pro Separator labeled Bottle Sensor ensure that the contents and fasten securely of waste container are not accidentally split Connect the hydrophobic air filters 0 2 um to the appropriate over the autoMACS connectors on the bottle closures Pro Separator during removal from the basket y Note The correct positioning of each solution container recognizable by the color Ax code and the symbols is crucial in order to perform successful procedures using the autoMACS Pro Separator To keep buffers sterile each bottle closure should be equipped with a hydrophobic air filter Avoid any contact of hydrophobic air filters with fluids as this may cause clogging ofthe filter When working with biohazardous samples itis recommended to fill the waste container with 100 mL of disinfectant before use e g MACS Bleach Solution 130 093 663 For proper disposal please follow local regulations and carefully read the chapter 1 3 Assembly and installation 3 3 Running the autoMACS Pro 3
122. he connector labelled with External CAN of the autoMACS Pro Separator refer to section 1 2 Size without lid 182 mm x 148 mm x 47 mm Size with lid 280 mm x 153 mm x 172 mm Weight 1 5 kg The MACS MiniSampler is designed for operation with three different tube racks and a reagent rack Racktype Slots Maximum Maximum Maximum number of sample number of samples volume cells per tube Chill 5 24x5 mL 6 5mLtubes 2 5 mL 5x10 Chill 15 15x15 mL 5 15 mL 12 5 mL 2 5x 10 5x5 mL tubes Chill 50 6x50 mL 3 50 mL 50 mL 4x 10 3x15 mL tubes 3x 5mL Table 11 4 MACS Cooling Tube Racks Chill Racks 5 15 and 50 Rack type Slots MACS Reagent Rack 4 4x MACS Reagent vials Table 11 5 MACS Reagent Rack 4 Conditions of operation 15 30 C with 0 85 humidity non condensing at a maximum altitude of 2000 m The MACS MiniSampler is not specified for use in the cold room The MACS MiniSampler has been investigated by Underwriters Laboratories in accordance with the standards UL 61010 1 and CAN CSA C22 2 No 61010 1 and meets the intent ofthe directive 2004 108 EC electromagnetic compatibility CE cpus ustep LAB EQUIP 52ZA Compliance was demonstrated by conformance to the following standards which have been listed in the Official Journal of the European Communities EN 61326 1 EN 61000 3 2 EN 61000 3 3 Compliance was demonstrated by conformance to the following FCC rules of the Code of Federal Regulations
123. he cooling capacity of the Chill Racks may be compromised Higher labeling temperatures could cause unspecific binding which may lead to lower purities when using positive selection kits Non labeled fraction shows low purity 1 Magnetic labeling was insufficient due to a low expression level of the surface marker It is recommended to use an indirect magnetic labeling system for example MACS Anti Biotin MicroBeads in order to increase magnetic labeling of the cells 10 Troubleshooting 10 183 184 The number of magnetically labeled cells exceeds the column capacity Calculate the number of expected magnetically labeled cells The number of cells should not exceed 2x108 per sample If necessary split the sample Labeling of cells was ineffective due to an incubation temperature lower than recommended Incubate the cells in the refrigerator 2 8 C for labeling Labeling of cells was ineffective due to too much debris and or dead cells in the sample Debris and dead cells will non specifically bind to all other components present Antibodies and MicroBeads will be captured non specifically and not be able to label the cells in the positive fraction sufficiently any more Cells were not labeled with MicroBeads because the MicroBeads were degraded Check the expiration date Check for sterility of the MicroBeads if the vial has been opened before Labeling of cells was performed for a shorter time than recommended The typic
124. hnical Support Take care not to pinch the tubings at the bottom left of the instrument when closing the bottom cover 7 Maintenance 7 4 2 Exchange of dilutor valve 1 Switch OFF and unplug the instrument 2 Remove the pump syringe as described in section 7 3 2 3 Unscrew the tubing 10 Figure 7 18 Unscrewing tubing connected to the dilutor valve Note The profile of the 4 Unscrew the two hexagonal socket screws using the key that was valve shaft is shaped delivered with the new or exchange valve Take care to note the asymmetrically similar orientation of the valve shaft as shown below to a trapezoid refer to figure 7 19 5 Pull the valve out of the coupling 6 When inserting the new valve into the coupling make sure that the short side of the trapezoid shaped valve shaft points towards the triangle in the coupling Figure 7 19 Inserting a new valve take care to note the orientation of the valve 7 Maintenance 157 158 7 When the valve is properly inserted mount the hexagonal socket screws 8 Connect the tubing according to the positioning of the diluter valve Figure 7 20 Connect tubing to the value as shown above 9 Guide the draining tube towards the washing station as shown below Figure 7 21 Red dashed line indicates the positioning of the draining tube The draining tube connects the valve to the washing station 10 Remove the washing station Take care to clean spilled
125. immediatamente eventuali schizzi Non lasciare che i liquidi penetrino all interno del dispositivo Staccare il cavo dell alimentazione prima di pulire manualmente lo strumento autoMACS Pro Separator Esistono potenziali rischi in caso di utilizzo di uno strumento AutoMACS Pro Separator aperto caduto o danneggiato in caso di infiltrazione di liquidi all interno dello strumento in caso di penetrazione di oggetti nello strumento attraverso le fessure di ventilazione o in caso di inserimento accidentale di oggetti nello strumento In caso di presenza di fiamme o fumo spegnere immediatamente il autoMACS Pro Separator staccare lo strumento dalla presa di corrente e contattare un rivenditore Miltenyi Biotec autorizzato o il servizio di assistenza clienti Miltenyi Biotec severamente vietato utilizzare uno strumento danneggiato o provvisto di cavo dell alimentazione danneggiato 1 3 2 Forte campo magnetico WARNING Extremely strong magnetic fields Risk of severe personal injury to persons carrying pacemakers or electronical medical implants stay clear Keep all magnetically sensitive objects away from device ATTENZIONE autoMACS Pro Separator provvisto di un magnete estremamente potente Tenere i supporti magnetici come carte di credito nastri magnetici e supporti di memorizzazione e i dispositivi elettronici come apparecchi acustici pacemaker strumenti di misurazione e controllo computer e orologi ad una distanza minim
126. information contained in this user manual before use Failure to read and follow these guidelines could lead to improper or incorrect use handling or care of your instrument and could cause hazards to users unpredictable results instrument malfunction or damage premature wear and reduced life time of the instrument and may void your warranty Keep this user manual in a safe place accessible for anyone using the autoMACS Pro Separator This chapter describes the safety instructions and site requirements for your autoMACS Pro Separator The following warnings and cautions are provided to help you prevent injury to yourself or damage to the instrument 1 1 Symbols and hazard levels Setup of safety notices Example CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard The safety notices inform the user about potential risks if warnings and precautions outlined below are not followed The icon on the left side specifies the risk The hazard level at the top classifies the hazard as mentioned above The level type and source of the hazard as well as potential consequences prohibitions and measures are pointed as follows 1 Important information 15 CAUTION WARNING o o OPP Symbols and hazard levels The following chart is an illustrated glossary depicting the symbols that are used in this user manual and on the AutoMACS Pro Separator Indicates a
127. instrument labeled RS232 BCR Use the screw driver to fasten the screws of the connecter Figure 12 23 Location of the cable sockets at the back of the autoMACS Pro Separator 12 Plug in and switch on instrument 13 Select menu Option Highlight User Settings and O_ber Press Run Figure 12 24 Setting up the software to recognize the 2D code reader 12 Hardware and software installation 211 14 Select Change 15 Select Keyence 16 Switch OFF instrument Wait 5 seconds and switch ON instrument again 17 Replace the cover 212 12 Hardware and software installation Technical support 13 Miltenyi Biotec offers a full range of customer technical support options for your autoMACS Pro Separator For support and technical questions or if you think your autoMACS Pro Separator is defective please contact your local Miltenyi Biotec representative or Miltenyi Biotec s technical support team Germany Austria Switzerland Benelux Miltenyi Biotec GmbH Miltenyi Biotec BV Friedrich Ebert Str 68 Postbus 85183 51429 Bergisch Gladbach 3508 AD Utrecht Germany Netherlands Phone 49 2204 8306 830 Customer service Belgium Fax 49 2204 8306 89 Phone 0800 94016 macsTec miltenyibiotec de Fax 0800 99626 macs miltenyibiotec nl la USA Canada Customer service Luxembourg Miltenyi Biotec Inc Phone 800 24971 2303 Lindbergh Street Fax 0800 24984 Auburn CA 95602 USA Phone 1 530 888 8871 Customer service Netherlands Fa
128. ion and operation of your autoMACS Pro Separator Read the instructions in this section and ensure that your site is properly prepared before you connect the instrument to its power source 1 Important information When planning your site layout and equipment locations keep in 1 mind the precautions described in this section to help avoid instrument failures and reduce the possibility of environmentally caused shutdowns IMPORTANT At all times local working area safety instructions laboratory policies and standards regarding laboratory health and safety and prevention of accidents must be adhered to 1 4 1 Mounting accessories Do not place the autoMACS Pro Separator on an unstable table cart stand tripod or bracket As a consequence the instrument might fall down This may cause serious bodily harm and or serious damage to the instrument Use only on a table cart stand tripod or bracket recommended by Miltenyi Biotec or sold with the instrument Do not place the autoMACS Pro Separator within a built in apparatus or a confined space such as a shelf rack unless the apparatus has been specifically designed to accommodate the instrument proper ventilation is provided and the mounting instructions for the instrument have been followed 1 4 2 Air circulation The instrument should not be placed next to radiators heat registers stoves or other pieces of equipment including amplifiers that produce heat Allow sufficient
129. ioning the Chill Rack 5 adjacent to a MACS Reagent Rack 4 on the MACS MiniSampler 20 Perform a system calibration test refer to section 3 3 for more details 3 2 Replacing fluid containers and the connection of fluid sensor cables Operating the autoMACS Pro Separator requires Running Buffer Washing Solution and storage solution absolute ethanol diluted to 70 with double distilled water It is recommended to operate the instrument with ready to use MACS Buffers The autoMACS Pro Separator is delivered with four empty fluid containers which are connected to the instrument with specifically designed bottle closures The bottle closures consist of a fluid uptake port or a fluid outlet port waste container as well as a sensor for measuring electrolyte conductivity The fluid containers bottle closures and fluid sensor cables are color coded for easy handling refer to table 3 1 below Container Symbol Container Symbol Running Storage Buffer solution blue black Washing Waste Solution red green Table 3 1 Symbols and color coding of fluid containers 1 Install one fluid at a time Please note the corresponding color coding refer to table 3 1 3 Assembly and installation 85 2 Unscrew bottle closures counter clockwise and remove the Note When removing the empty container Do not disconnect the color coded tubing waste container unscrew the bottle closure and 3 Do not open afresh bottle until it is placed in t
130. iori ai 3 metri di lunghezza 1 5 Funzionamento manutenzione trasporto e smaltimento sicuri Seguire le istruzioni di seguito riportate onde assicurare che il funzionamento la manutenzione il trasporto e lo smaltimento del vostro autoMACS Pro Separator avvengano in modo corretto e sicuro IMPORTANTE vanno sempre osservate le istruzioni di sicurezza sul luogo di lavoro le procedure di laboratorio e le norme relative alla salute e alla sicurezza del laboratorio nonch alla prevenzione degli infortuni 1 5 1 Funzionamento sicuro Qualora lo strumento non funzioni correttamente e le istruzioni o i messaggi visualizzati sullo schermo invitino a contattare l assistenza tecnica la sicurezza di impiego non pi garantita Spegnere immediatamente il autoMACS Pro Separator staccare lo strumento dalla presa dell alimentazione e contattare un rivenditore Miltenyi Biotec autorizzato o il servizio di assistenza clienti Miltenyi Biotec 1 5 2 Manutenzione IMPORTANTE se non diversamente indicato nel presente manuale d uso o in altri documenti forniti da Miltenyi Biotec non eseguire autonomamente la manutenzione del vostro autoMACS Pro Separator Gli interventi di manutenzione e riparazione devono essere eseguiti da personale qualificato Interventi di manutenzione e riparazione scorretti o impropri del autoMACS Pro Separator possono rappresentare un pericolo per l incolumit degli operatori avere conseguenze imprevedibili causare malfunzi
131. iotec ou consultez le site www miltenyibiotec com support IMPORTANT Si des pi ces de remplacement ou de rechange sont requises assurez vous que le prestataire de services utilise exclusive ment des pi ces Miltenyi Biotec d origine ou des pi ces de fabricants tiers sp cifi es et recommand es par Miltenyi Biotec L utilisation de pi ces de remplacement ou de rechange non autoris es peut entra ner le dysfonctionnement de l appareil et fausser les r sultats de la s paration des cellules Miltenyi Biotec n accorde pas de prestation de garantie ou d cline toute responsabilit pour les pannes et les dommages de l appareil r sultant de l utilisation de pi ces de remplacement ou de rechange inappropri es Une fois les travaux d entretien ou de r paration achev s demandez votre prestataire de services Miltenyi Biotec agr d effectuer tous les contr les de s curit requis par la proc dure de r paration afin de garantir que l appareil est parfaitement op rationnel Utilisez uniquement les options et les mises jour recommand es par Miltenyi Biotec 1 5 3 Nettoyage D branchez le s parateur autoMACS Pro Separator avant le nettoy age N utilisez pas d agents nettoyants liquides ou en a rosol utilisez toujours un chiffon humide 1 5 4 Mati res dangereuses WARNING Risk of loss of life if biohazardous material is used Wear protective gloves protective clothing and safety glasses to prevent contact with
132. is reported confirm by selecting OK If warning 7016 is reported select Ignore Note If warning 7016 Columns are not installed Start column exchange program appears select Ignore Panes software version confe _ Verzion AM_01 02 5 reaured A 15 5 Figure 12 2 The user must confirm any update The current software panel software version will be overwritten 12 2 Calibration of the autoMACS Pro Separator The autoMACS Pro Separator is calibrated by using the two programs Calibr_1 and Calibr_2 Calibration 1 is recommended after installation of new software Calibration 2 is always necessary when the pump syringe the Diluter Valve or the tubing are changed Calibr_1 is used to calibrate the settings of the needle arm x y and z axis i e alignment of the needle arm with the washing station the MACS Cooling Tube Racks and the bottom detection sensor control Calibr_2 automatically calibrates the liquid volume control This is crucial for the correct measurement and processing of the sample volumes The fluidic system must be filled with buffer before commencing this step i e the Rinse program must be performed Calibration 1 Beginning the calibration sequence 1 Select menu Option 2 Highlight User settings and select Calibr_1 3 Select Run 4 Select Calibrate to proceed The first of five 1 5 calibration steps will begin namely calibration of the washing station 12 Hardware and s
133. is subjected to excessive vibration or if it is dropped If the instrument needs to be shipped back to the manufacturer for service decontaminate the instrument from any hazardous material prior to shipment If you have questions regarding proper decontamination or shipment please contact Technical Support for assistance Refer to section 7 3 4 for further information on instrument decontamination 1 5 6 Instrument disposal Please contact Technical Support for assistance if you wish to dispose of your instrument 1 5 7 Electromagnetic compatibility Changes or modifications of the equipment unless expressly approved by Miltenyi Biotec may void your authority to operate the equipment pursuant to 47 CFR 15 1 Important information Note This equipment has 1 been tested and found to comply with the limits for a Class B digital device pursuant to Part 15 of the FCC Rules These limits are designed to provide reasonable protection against harmful inter ference in a residential installation This equip ment generates uses and can radiate radio frequency energy and if not installed and used in accordance with the instructions may cause harmful interference to radio communications However there is no guarantee that interfer ence will not occurina particular installation If this equipment does cause harmful interfer ence to radio or televi sion reception which can be determined by turning the equipment off and on
134. l chi sur une surface de miroir Sinon des l sions oculaires peuvent en r sulter Ne pas diriger intentionnellement le rayon laser sur d autres personnes Ne pas d monter changer ni retirer des sources de rayon laser ou DEL encastr es ni leur support Les sources de rayon laser ou DEL ne cessent pas forc ment d mettre un rayonnement une fois d mon t es Le rayonnement d appareils d mont s peut entra ner des l sions oculaires Prendre garde au chemin optique du rayon laser ou de sa r flexion sur une surface de miroir Veiller lors de l installation de l autoMACS Pro Separator ce que le chemin optique rayon laser ne se trouve pas hauteur des yeux de personnes pendant le service Eviter que de l eau de l huile de la poussi re ou d autres corps tranger adh rent sur la fen tre de lecture du lecteur de code 2D comme ceci risque d entra ner des erreurs de lecture S assurer avant le nettoyage du scanner qu aucun rayon laser n est plus mis La manipulation du laser risque sinon de provoquer des l sions oculaires Utiliser un chiffon doux pour essuyer des substances sur le scanner Ne pas utiliser d alcool ou d autres produits de nettoyage L autoMACS Pro Separator est d sign comme produit laser de la classe 1M selon la norme CEI 60825 1 1993 A1 1997 A2 2001 AVERTISSEMENT l utilisation d organes de commande autres que ceux mentionn s dans ce document ainsi que toute adaptation ou utilisation d autres pr
135. la garant a ni aceptar ninguna responsabilidad por la aver a de aparatos o por los da os resultantes del uso de piezas de recambio o de repuesto inapropiadas Una vez 1 Important information 71 72 completado el servicio o la reparaci n haga que su proveedor de servicios autorizado por Miltenyi Biotec realice todos los controles de seguridad requeridos por el proceso de reparaci n para asegurarse de que el instrumento esta en buenas condiciones de funciona miento Utilice exclusivamente las opciones y actualizaciones recomendadas por Miltenyi Biotec 1 5 3 Limpieza Desenchufe el separador autoMACS Pro de la toma de corriente antes de proceder a su limpieza No emplee productos de limpieza liquidos ni aerosoles y use siempre un trapo humedo 1 5 4 Sustancias peligrosas WARNING Risk of loss of life if biohazardous material is used Wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Operate device in a safety hood Decontaminate device after spilling of biohazardous material Si se utilizan o se han utilizado sustancias biol gicamente peligrosas el operario deber a utilizar el equipo de seguridad que aparece en las se ales de aviso de las sustancias empleadas P ngase guantes ropa y gafas de seguridad para evitar el contacto con la piel y con los ojos Un equipo de seguridad defectuoso o inadecuado puede poner en peligro al operario El separador autoMACS Pro deb
136. laser beam at others 1 Important information 19 20 Do not disassemble modify or remove the installed laser or LED radiation sources or their mounting rackets The laser or LED radiation sources do not automatically stop emitting when disassembled Radiation of disassembled units may lead to eye injuries Be careful of the path of the laser beam or reflection from a mirrored surface Take care during installation of the autoMACS Pro Separator that the path of the laser beam is not at the same height as that of the human eye during operation Do not allow water oil dust or other foreign substances to stick to 2D Code Reader aperture window This may cause read errors Be sure to stop the laser emission before cleaning the scanner Otherwise exposure to the laser may cause eye injury Use a soft dry cloth to wipe any substances from the scanner Do not use alcohol or other cleaning substance The autoMACS Pro Separator is classified as a Class 1M laser product per standard IEC 60825 1 1993 A1 1997 A2 2001 CAUTION Use of controls or adjustments or performance of procedures other than those specified herein may result in hazardous radiation exposure Figure 1 3 Position of lasers Invisible rack detection lasers are located within the rectangle area The 2D code reader visible is located within the open circle 1 4 Secure installation This section describes the requirements your site must meet for safe installat
137. let port manually using a syringe filled with 70 ethanol or double distilled water 10 1 7 MACS MiniSampler does not move properly 1 5 Check whether the guiding of the MACS MiniSampler is connected properly to the connector at the autoMACS Pro Separator labeled External CAN Check whether the bolt below the rack detection protrudes from the instrument If this is the case push it in and turn it clockwise to lock the bayonet mount Check the cable connection between MiniSampler and autoMACS Pro Separator Check for cable damages Check whether the MiniSampler can freely move to both sides and check for any resistance or collision If the problem persists contact Technical Support 10 1 8 Touchscreen remains dark 1 4 Switch OFF the instrument wait 5 seconds and switch ON again If the autoMACS Pro Separator still does not initialize go to step 2 Check if the power cord is plugged in correctly and if the electric power is switched on Replace the fuses refer to section 7 6 Spare fuses are included in the autoMACS Pro Separator Starting Kit If the problem persists contact Technical Support 10 1 9 Disruption of power supply during cell separation In the unexpected event of a power supply failure during the cell separation procedure follow the rescue procedure to recover the cell sample trapped in the autoMACS Pro Separator fluidic system 1 2 Switch ON the autoMACS Pro Separator Dis
138. ley Surrey GU24 9DR Phone 44 1483 799 800 Fax 44 1483 799 811 macs miltenyibiotec co uk Limited warranty 14 Except as stated in a specific warranty statement which may accompany your autoMACS Pro Separator the Product or unless otherwise agreed in writing by an authorized representative of Miltenyi Biotec Miltenyi Biotec s warranty if any with respect to this Product is subject to the terms and conditions of sale the Terms of the company within the Miltenyi Biotec group which supplied the Product The Terms may vary by country and region Copies of these Terms are available on request or at www miltenyibiotec com Nothing in this document should be construed as constituting an additional warranty Miltenyi Biotec s product warranty only covers Product issues caused by defects in material or workmanship encountered during ordinary use as described in the user manual or other documentation provided by Miltenyi Biotec it does not cover Product issues not arising out of defects in material or workmanship including but not limited to Product issues resulting from failure to follow installation operating and or maintenance instructions or environmental conditions prescribed in this user manual or other Product docu mentation misuse abuse neglect mishandling unauthorized or improperly performed maintenance or repairs accident acts of God a limitations of technology electrical current fluctuations modifica
139. ling Tube Rack with sample tubes onto the MiniSampler It is recommended to use a Chill 5 Tube Rack 10 Select Done and then select Use recommended to start the calibration using current settings 11 Check the central positioning of the uptake needle on the bottom of the tube by using the needle navigation buttons Move back Move fwd Height Ensure the needle is positioned at the bottom center of the tube in row A of the tube rack Select Done to continue Figure 12 7 The tube must appear 12 Select Save to store the new settings Calibration 1 Calibration of the bottom detection step 3 5 13 Select Calibrate to proceed with calibration 14 Place a MACS Cooling Tube Rack onto the MACS MiniSampler 15 Select Done 12 Hardware and software installation 16 Select Use to start the calibration at current position 1 2 recommended Select Reset to reset the position to factory settings The needle will position itself a fraction of a millimeter over the surface ofthe Chill Rack at a point equidistant to the tube coordinates A1 A2 B1 B2 It should not make contact with the plastic surface Figure 12 8 Bottom detection calibration In the case of a Chill 5 Tube Rack the needle will be positioned midpoint equidistant to tube coordinates Al Note The uptake needle A2 B1 B2 moves to the surface ofthe tube rack The 17 To adjust the position select Change or select Done if the position of the needle current settin
140. ll report the current status PE PTE ET ETC ZE CS erabied m gt Figure 4 28 O_progs Enabling or disabling special separation protocols Select Disable to disable special separation programs 3 Select OK if the current reported status should not be changed Alternatively select Disable or Enable to change the status Sepcount Displaying the number of performed column separations The Sepcount or Sep counter program is used to display the number of column separations that have been on the autoMACS Pro Separator To view this statistic perform the follows steps 1 Select Option User settings and Sepcount 2 Select Run A dialog box will report the number of separations 4 An overview of the autoMACS Pro touchscreen user interface N gt Figure 4 29 Sepcount viewing the number of performed separations In a this example no separations have been performed using this instrument 3 Select OK to return to the option menu Set_time Setting the time and date To set the time and date perform the following steps 1 Select Option User settings and Set_time Select Run 2 Highlight either the time or date fields by touching the display 3 Use the keypad to change the date or time accordingly 4 Select OK to return to the option menu The procedure can be cancelled at anytime by selecting Cancel Valve_ex Exchanging the instrument values This program is used for valve exchange Use of this program turns th
141. markers are targeted for magnetic separation When using a fluorochrome conjugated antibody or other ligand in combination with the corresponding Anti Fluorochrome MicroBeads the fluorescent staining can be used for flow cytometric analysis Cell isolation directly from whole blood or bone marrow samples MACS Whole Blood MicroBeads are specially developed to isolate human cell subsets directly from human whole blood using the autoMACS Separator or the autoMACS Pro Separator This reagent can also be applied to bone marrow Note that in this case fat must be removed prior to labeling 6 3 2 Choose a magnetic labeling strategy Autolabeling may be performed prior to cell separation For a list of MACS Products optimized for autolabeling please visit www automacspro com Installing reagents for use with autolabeling The reagent menu is used to program any reagent vials that are required for automated magnetic labeling and subsequent cell sorting Reagents can be entered using the 2D code reader or manually using the Enter Reagent input panel I uk Figure 6 1 The reagent menu 6 Cell separation using the autoMACS Pro Separator Entering reagent information for autolabeling 6 Reagents vials can be directly scanned and recognized by the autoMACS Pro Separator 2D code reader Scanning reagents with the 2D code reader 1 Select Reagent tab and highlight the position where the vial will be placed on the reagent rack Four positions are
142. medici n y control ordenadores y relojes a una distancia m nima de 20 cm de la cubierta magn tica El campo magn tico puede alterar o da ar dichos objetos 1 Important information Figura 1 1 Se al de advertencia para iman permanente potente 1 3 3 Peligro de aplastamiento y corte CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard WARNING No abrir las cubiertas de acceso frontal durante el servicio del dispositivo No obstruir el movimiento del brazo automatizado y de los accesorios durante la operaci n Mantener los dedos etc lejos de todas las piezas m viles del separador autoMACS Pro y de los accesorios para evitar aplastamientos lesiones de corte o da os en el dispositivo No tocar las bombas de fluido ni ajustar la tuberia durante la operaci n del dispositivo Desactivar siempre el dispositivo antes de ajustar cualquier parte del sistema de fluidos El procedimiento debe pararse o interrumpirse antes de maniobrar los accesorios p ej el MACS MiniSampler o antes de colocar o retirar los tubos del soporte de tubo colocado en el sampler No omitir ninguna de las medidas o los dispositivos de seguridad Figura 1 2 El c rculo abierto muestra la se al de advertencia para el peligro de aplastamiento o corte 1 3 4 Radiaci n l ser WARNING El dispositivo est equipado de cuatro unidades l ser de emisi n superficial de cavidad vertical VS
143. ment is switched off 7 Run the autoMACS Pro Cell Separation The autoMACS Pro Separator automatically isolates target cells from up to six independent samples in a sensor controlled manner 8 Shutdown the autoMACS Pro Separator A sleep or store program is used for overnight storage or long term storage respectively Sleep or store must be performed before shutting down the autoMACS Pro Separator 6 1 Select a cell labeling strategy The autoMACS Pro Separator is capable of performing autolabeling manual labeling and a combination of both strategies Up to six samples can be processed at one time Autolabeling protocols are being continually developed and optimized by Miltenyi Biotec For a current list of MACS Reagents and Kits that are optimized for cell separations with the autoMACS Pro Separator autolabeling feature please visit www automacspro com 6 2 Prepare cell samples In order to obtain optimal separation results some crucial points must be considered 6 2 1 Prepare single cell suspensions Cell aggregates may contain mixtures of target and non target cells and therefore can impair the separation results To minimize the risk of cell aggregation MACS Separation Buffer MACS Running Buffer should be used during sample handling steps Resuspend cells carefully after centrifugation For specific recommendations please refer to the general protocol section of the user manual or to the respective MACS Cell Separation Reagent
144. ming will begin The autoMACS Pro Separator will display a warning screen if the buffer supply is low or if the waste bottle is full If no wash program has been performed before the first separation a warning screen will ask to rinse the system You may interrupt any program by selecting Stop The Stop button is located at the bottom left hand corner in all menus When priming the instrument for the first time or when the instrument was not serviced for along period of time it is recommended to visually inspect the fluidic system for potential leaks Open the front door after priming the instrument If there is any sign of leakage e g salt deposits tighten the respective tubing connection Close the front door and proceed with the separation one separation and that the waste container can collect fluid from at least one separation If more than one separation is performed ensure that the containers contain sufficient fluid for all the separations and that the waste container is empty To verify the status of the bottles select the menu Status from the upper navigation bar On the left hand side of the menu four symbols display the solution containers and their filling status If the fluid containers are full and the waste container is empty the symbols are green If the solution containers are empty or the waste container is full the respective symbols are red The symbol for the ethanol container remai
145. mputer controlled instrument for the automated separation of magnetically labeled cells using MACS Technology The MACS SS MiniSampler connects to the autoMACS Pro Separator and thus represents a part of the cell separation instrument The autoMACS Pro Separator and the MACS MiniSampler are designed to operate safely after installation and when used by trained personnel according to general safety practices and the instructions set forth in this user manual The guidelines in this section explain the potential risks associated with the operation of the instrument and provide important safety information in order to minimize these risks By carefully following the instructions you can protect yourself and the equipment from potential hazards and create a safe work environment If this instrument is used in a manner not specified by the manufacturer protection may be impaired IMPORTANT Please read and follow all operating instructions in this user manual and pay attention to all warnings displayed on the instrument Retain this user manual and any other safety and operating instructions provided with the instrument in a place accessible to all users for future reference IMPORTANT The autoMACS Pro Separator is intended for indoor use only Do not use the instrument in areas classified as hazardous locations such as oxygen laden environments Contact your local authority governing electrical power supply building constructions maintenance or
146. n Safe and cleaning of the instrument for long term storage Store Safe This is a disinfectant procedure which uses MACS Bleach Solution for cleaning and decontamination of the autoMACS Pro Separator Depending on the level of use and general instrument maintenance it is recommended to decontaminate the fluidic system every 3 to 6 months using the Safe program HE Store The program Store should be applied to prepare the instrument for long term storage Upon completion of the Store program the fluidic system contains 70 ethanol To store the autoMACS Pro Separator for a period longer than two weeks the tubing system should be cleaned and the columns should be replaced with column substitutes The Store program automatically performs the cleaning procedure and prompts the user to install column substitutes 4 7 2 User settings The User settings are for maintenance and setup of the autoMACS Pro Separator Each program is briefly discussed below About About informs the user about the software version serial number of the instrument and other hardware information Calibr_1 Performing calibration 1 of the needle arm positioning This program is used for the calibration of the needle arm to the washing station and tube racks Refer to section 12 2 for more information Calibr_2 Performing calibration 2 of the instrument tubing This program is used for the calibration of the instrument tubing Refer to sec
147. n substitutes cannot be used for cell separations 15 Glossary 15 217 218 Depletion Isolation of target cells by labeling all cells other than the target cells non target cells with MACS MicroBeads and subse quently performing a MACS Separation The non labeled fraction contains the target cells Deplete Depletion program standard mode a normal cell deposi tion rate is used The non labeled cells are eluted in row B of the sample rack This is the optimal program to be used in combination with MACS Cell Isolation Kits or to achieve the highest recovery rate of untouched cells Depletes Depletion program sensitive mode a slow cell deposition rate is used 1 mL min The non labeled cells are eluted in row B of the sample rack This program is optimized for depletion of those cells that weakly express the antigens used for magnetic labeling or to achieve optimal purity of the untouched cell fraction Depl05 Special depletion program the cell deposition rate is 0 5 mL min The non labeled cells are eluted in row B of the sample rack This program is used for strong depletion of those cells that weakly express the antigens used for magnetic labeling or to achieve the highest purity of the untouched cell fraction However choosing Depl05 might result in a reduced recovery of the target cell fraction Depl025 Special depletion program the cell deposition rate is 0 25 mL min The non labeled cells are eluted in row B
148. ned to stabilize the instrument and maintain optimal alignment with the MACS MiniSampler autoMACS Pro Angle Connector Set Order no 130 093 245 The autoMACS Pro Angle Connector Set has been designed for placement of the autoMACS Pro Separator in a location with limited space Cables that connect the back of the instrument i e the fluid sensor cable the MACS MiniSampler cable and the 2D code reader cable are connected at a 900 angle to reduce the total depth of the instrument The set contains three Sub D pin adaptors 5 Materials required 115 116 5 Materials required Cell separation using the 6 autoMACS Pro Separator The following chapter describes how to perform cell separations using the autoMACS Pro Separator IMPORTANT The autoMACS Pro Separator is intended for research applications only and not for diagnostic or therapeutic use The procedure of cell separation has been categorized into eight steps 1 Select a cell labeling strategy Choose if autolabeling manual labeling or a combination of both strategies should is needed 2 Prepare cell samples A single cell suspension devoid of dead cells aggregates and cell debris is the prerequisite for efficient cell separation 3 Select a cell separation strategy and a suitable magnetic labeling strategy Prior to cell separation samples are labeled by magnetic MicroBeads This can be performed manually or automatically using the autoMACS Pro Separator Cell separa
149. ng system Exchange of fuses Rescue procedure Rescue procedure A Rescue procedure B Spare parts and accessories list Accessories list Protocols Direct magnetic labeling Indirect magnetic labeling 169 Chapter 9 171 171 171 172 Chapter 10 173 173 173 174 174 175 175 176 176 176 176 177 177 177 178 179 179 180 180 180 183 184 185 185 186 186 1 Important information 8 3 9 1 9 2 9 3 10 10 1 10 1 1 10 1 2 10 1 3 10 1 4 10 1 5 10 1 6 10 1 7 10 1 8 10 1 9 10 2 10 2 1 10 2 2 10 2 3 10 2 4 10 2 5 10 2 6 10 3 10 3 1 10 3 2 10 3 3 10 4 10 4 1 10 4 2 10 5 Direct magnetic labeling of human cells using MACS Whole Blood MicroBeads Quality control of separations performed with the autoMACS Pro Separator Recovery of cells Purity of isolated cell population Viability of the cells Troubleshooting Hardware problems not indicated by a warning or error screen Column leakage Tubing leakage Pump syringe leakage Pump syringe is filled with air during operation Washing station overflow Outlet port is clogged MACS MiniSampler does not move properly Touchscreen remains dark Disruption of power supply during cell separation Performance errors not indicated by a warning or error screen Output volumes are not correct Low cell viability in final fraction Low purity of isolated cell population Low recovery of isolated cells Sampl
150. ng the sample template 2 Scroll through the Separation submenu using the arrows amp Y 3 Select and highlight a Separation program Figure 4 17 Positive cell separation sensitive mode was selected Assigning a wash program The Wash submenu 5 is used to instruct the autoMACS Pro Separator which wash program should be applied to each sample Three wash programs are available to choose from Qrinse quick rinse Recommended to save time between sample separation steps The tubing and column receive a quick rinse Rinse The system is primed receiving an extensive wash Rinse is recommended between rare cell whole blood cord blood and bone marrow separations Sleep A rinse is performed before the system is shutdown To assign or modify a wash program 1 Highlight the appropriate sample s using the sample template 2 Scroll through the Wash submenu using the arrows A Y 3 Select highlight a Wash program 4 An overview of the autoMACS Pro touchscreen user interface EE 0 00 Figure 4 18 A rinse program was selected Working with separation templates For convenience it is possible to load and save separation templates Separation templates can be used in combination with Reagent templates refer to above To save a separation template 1 Configure the sample template an example follows EE 0 0 Figure 4 19 A sample template was setup to process two samples Sample 1 Isolation
151. ni locali vigenti 1 5 5 Trasporto Lo strumento autoMACS Pro Separator va trasportato con cautela in imballaggi specificati da Miltenyi Biotec In caso di cadute o di vibrazioni eccessive lo strumento pu subire dei danni interni Qualora si renda necessario rispedire lo strumento al produttore per interventi di manutenzione decontaminare lo strumento da eventuali materiali pericolosi prima della spedizione In caso di domande circa la decontaminazione o il trasporto adeguati contattare l assistenza tecnica 1 5 6 Smaltimento Contattare il servizio di assistenza tecnica per lo smaltimento dello strumento 1 5 7 Compatibilit elettromagnetica Soltanto per gli USA Eventuali cambiamenti o modifiche dello strumento non approvati espressamente da Miltenyi Biotec possono inficiare il diritto all uso dello strumento in conformit all art 15 47 CFR 1 Important information 61 62 1 Important information Informac e 7 on mportante 1 Lea la siguiente informacion antes de usar el instrumento equipo Por favor lea integramente la informaci n contenida en este manual de usuario antes de utilizar el instrumento equipo La lectura y o seguimiento incorrectos de estas directrices pueden llevar a un uso manejo o mantenimiento incorrectos del equipo lo cual puede poner en peligro a sus usuarios producir resultados impredecibles derivar en un mal funcionamiento del aparato o que ste sufra da os asi como causar un desgas
152. no action is required If the symbol is red the columns must be exchanged The level of the green fill on the column symbol indicates the remaining operation life of the autoMACS Columns Touch the column symbol to activate the column status popup textbox Ii Figure 4 9 Column status By touching the column symbol a pop up box displays a text report of the column status 4 An overview of the autoMACS Pro touchscreen user interface 4 3 3 Tube rack status A Figure 4 10 Tube rack status graphic a 5 mL tube rack was detected If no tube rack is detected no graphic is displayed a Rack detection occurs prior to starting the separation process Before cell labeling and or cell separation is performed the instrument will not attempt to detect the rack Touch the position of the rack status graphic in order to view the tube rack popup text table Un 1 Figure 4 11 Tube rack status text box No rack is detected until cell separation is performed 4 3 4 MACS MiniSampler status Figure 4 12 MACS MiniSampler status graphic Left The MiniSampler was successfully installed Right No MiniSampler was detected If the MACS MiniSampler has been detected correctly a rectangular symbol is displayed If it has not been detected a hand symbol will be displayed in the same field 4 4 Separation menu The Separation menu schematically represents a sample rack and allows definition of sample processing strategies for each sample
153. nough Storage Solution is in the bottle Rinsino th Figure 6 31 Running the Store program 3 Install the column substitutes refer to section 3 3 4 for details on column exchange 4 Select Done 5 Switch off the autoMACS Pro Separator using the main power switch 6 8 3 Shutdown button 1 Press the shutdown button on the upper right hand corner of the screen W 6 Cell separation using the autoMACS Pro Separator 143 2 Select Yes from the popup dialog box Do you rendy ward 10 seen he device RS 00 Figure 6 32 Shutting down the autoMACS Pro Separator 3 The autoMACS Pro Separator will automatically perform a Sleep program 4 Upon completion of the Sleep program you will be prompted to shut down the instrument The bottle illumination is purple at this point Switch off the autoMACS Pro Separator by using the main power switch Note Ensure enough storage solution is available 144 6 Cell separation using the autoMACS Pro Separator Maintenance 7 Appropriate maintenance of the autoMACS Pro Separator helps to maintain excellent reproducibility of the cell separation results The following section gives you an overview on the procedures required for efficient maintenance of the instrument IMPORTANT Insufficient or improper maintenance of your autoMACS Pro Separator can cause unpredictable results avoidable malfunction and premature failure of the instrument and may void your warranty IMPOR
154. ns gray It is recommended to select the menu Status when the autoMACS Pro Separator is in operation Saas x Figure 3 26 Status of the fluid containers is green for ready 3 3 7 Test of barcode scanner 1 Select Reagent 2 Press Read Reagent 3 Place barcode figure 1 3 in front of barcode scanner 4 CD4 MicroBeads human will appear as scanned reagent 3 Assembly and installation An overview of the autoMACS Pro A touchscreen user interface The autoMACS Pro Separator is operated through a TFT color touchscreen After switching on the instrument the startup screen pre will appear MM Figure 4 1 The autoMACS Pro Separator startup screen 4 1 Working with the lower menu bar Figure 4 2 The lower menu bar of the Separation menu A lower menu bar is accessible from all Menu screens Depending on the status of the instrument and the selected upper tabbed menu option the lower menu buttons switches between an inactive grey background and active state white background state For example Stop can only be selected when the instrument is actively performing a process such as cell separation Figure 4 3 Left Stop button is active and can be selected Right Stop button is inactive and cannot be selected 4 2 The main menu screen Five main menus allow easy interaction with the instrument They are accessed through the upper tabbed menu 4 An overview of the autoMACS Pro touchscreen user int
155. ntenendo uno spazio libero di almeno 15 centimetri su ogni lato durante il funzionamento onde assicurare un adeguato raffreddamento dello strumento Evitare l esposizione diretta ai raggi solari Le fessure e le aperture dello strumento sono necessarie per la ventilazione e non devono essere in alcun caso ostruite o coperte poich garantiscono il funzionamento corretto del autoMACS Pro Separator e proteggono il dispositivo dal surriscaldamento Non inserire mai corpi estranei nelle aperture dello strumento 1 4 3 Acqua e umidit Non utilizzare lo strumento in ambienti bagnati o umidi Evitare ambienti ad elevata umidit o condizioni di condensa e proteggere lo strumento dagli schizzi d acqua 1 4 4 Prodotto con messa terra Lo strumento dotato di un sistema di alimentazione a tre fili ed provvisto di spina con un terzo polo per la messa a terra La spina pu essere inserita unicamente in una presa di corrente con messa a terra per ragioni di sicurezza Non cercare di inserire la spina in una presa sprovvista di messa a terra Qualora non riusciate ad inserire la spina nella presa vi consigliamo di contattare il vostro elettricista di fiducia per sostituire la presa 1 4 5 Fontidi alimentazione Lo strumento deve essere alimentato unicamente dalla fonte indicata sulla targa relativa alle caratteristiche di alimentazione del prodotto In caso di domande sul tipo di alimentazione da usare contattare il rivenditore Miltenyi Biotec au
156. o remove the clumps from the sample 10 Troubleshooting 179 180 2 Remove the sample tube Wipe the uptake port needle with a tissue soaked with 70 ethanol Run the Qrinse program 3 Check for cell clumps in the tubing system If cell clumps are suspected run the Safe program 4 For separating cells that tend to aggregate e g tissue cells it may be helpful to dilute the sample 1 2 5 Remove the sample tube and run the Qrinse program Check tubing system and pump syringe for air inlet 6 Make sure that all tubing are connected properly especially the tubing connected to the columns and to the buffer bottles 7 Verify that the uptake port needle is connected correctly and that no air inlet is possible Tighten if necessary 8 Make sure that both columns are installed correctly 9 Clean pump syringe as described in section 10 1 3 to remove salt deposits Salt crusts might allow air to enter the fluidic system 10 Check whether the hydrophobic air filters connected to the bottles are clogged If filters are clogged replace them with new hydrophobic air filters 11 Run the Check_up program to identify hardware malfunctions 12 If problem persists call Technical Support 10 2 6 Reagent vial runs dry As the liquid levels in the reagent vials is not controlled by the system before uptake of reagent it will not be detected if the reagent volume provided in the vial is sufficient for the current separation process or not
157. o turn an extra quarter turn Do not to overwind the screw 6 Ifthe problem persists contact Technical Support 10 1 5 Washing station overflow 1 Verify that the washing station is not clogged with salt deposits Take out the washing station and clean as indicated in section 7 3 3 2 Reassemble washing station and run a Rinse program From the Separation menu select Wash Now lower menu bar Highlight Rinse and click Run 3 If problem persists follow the steps below 4 Make sure that the peristaltic waste pump works properly Run the check_up program From the Option menu select User settings Highlight Check_up and click Run 5 Ifthe check_up program reports a problem with the waste pump remove the pump head and clean the pin that drives the pump for details refer to section 7 4 4 Clean the washing station refer to section 7 3 3 Reassemble the unit and check whether the problem persists 6 Ifthe problem persists replace the pump head refer to section 7 4 4 10 Troubleshooting 175 176 Note If initial wash Rinse program is enabled press Stop as soon as the Status menu appears on the screen A window opens displaying that the process is paused Press Cancel 10 1 6 Outlet port is clogged 1 If the outlet ports are clogged e g due to salt deposits the elution process might be affected Wipe the outlet port with a tissue soaked with 70 ethanol or double distilled water Flush the out
158. oc d s que ceux mentionn s dans ce docu ment peuvent d gager des rayonnements dangereux Rayonnement laser Danger de graves l sions oculaires et cutan es 1 Important information Figure 1 3 Position des lasers Les lasers invisibles de d tection de rateliers sont situ s dans la zone rectangulaire Le lecteur de code 2D visible est entour d un cercle blanc 1 4 S curit de l installation Ce paragraphe d crit les exigences auxquelles votre site doit r pondre pour garantir l installation et la s curit d utilisation de votre s parateur autoMACS Pro Separator Lisez les consignes de ce paragraphe et assurez vous que votre site a t am nag en cons quence avant de brancher l appareil sa source lectrique Lors de la planification de l am nagement de votre site et de l emplacement de l quipement tenez compte des pr cautions d crites dans ce paragraphe afin d viter que l appareil ne tombe en panne et de r duire le nombre d arr ts pouvant tre caus s par l environnement de l appareil IMPORTANT Les consignes de s curit concernant la zone de travail locale les bonnes pratiques de laboratoire ainsi que les directives relatives la sant la s curit et la pr vention des accidents en laboratoire doivent tre observ es en permanence 1 41 Accessoires de montage Ne placez pas le s parateur autoMACS Pro Separator sur une table un chariot un pied un tr pied ou un support L a
159. odies allows low background and easy optimization of the separation Fluorescent staining using fluorochrome conjugated antibodies can simultaneously be performed for subsequent analysis of the separated fractions by flow cytometry or fluorescence microscopy 6 Cell separation using the autoMACS Pro Separator 121 122 Note Titration and manual incubation with the primary antibody or other ligand is recommended to be done manually If there is no wash step between incubation of the sample with an unknown primary antibody or other ligand followed by application of indirect MicroBeads the procedure cannot be optimized Results may be impaired Indirect magnetic labeling Indirect labeling is recommended when no direct MicroBeads for a particular cell surface marker are available Almost any monoclonal or polyclonal antibody or other specific ligand targeting any cell type from any species can be used for indirect labeling Cells are first incubated with a primary antibody ligand that is unconjugated biotinylated or fluorochrome conjugated In a second step magnetic labeling is performed by using Anti Immunoglobulin Anti Biotin Streptavidin or Anti Fluorochrome MicroBeads respectively A cocktail of antibodies or other ligands can also be used to concurrently isolate or deplete a number of cell types As it results in an amplification of the magnetic label indirect labeling may be the method of choice if weakly expressed
160. of the branch circuit rating 1 4 6 Accessibility Make sure that the main switch as well as the connector for the power cable are easily accessible and located as close to the operator of the instrument as possible If it is necessary to disconnect the power supply unplug the cable from the power outlet 1 4 7 Peripheral devices Only peripheral devices that comply to UL 60950 are allowed to be connected to the RS232 connector labeled COM The connector labeled RS232 AUX is not in use In addition only original autoMACS Pro Equipment should be attached to the connectors labeled External CAN CAN1 and CAN2 The voltage levels on these connectors shall not exceed hazardous voltage levels of 30 V rms and 42 4V peak or 60 Vdc Only the autoMACS Pro Bottle Sensor Cable should be attached to the Bottle Sensor connector Only a 2D code reader recommended by Miltenyi Biotec should be connected to the RS232 BCR connector External laser devices connected to the connector labeled RS232 BCR have to comply with the standard IEC 60825 1 Only use connector cables less than 3 min length 1 5 Secure operation maintenance transport and disposal Observe the following instructions to ensure secure operation maintenance transport and disposal of your autoMACS Pro Separator IMPORTANT At all times local working area safety instructions laboratory policies and standards regarding laboratory health and sa
161. oftware installation RS 0 Figure 12 3 Performing calibration of the needle arm a 5 step calibration process Calibration 1 Calibration of the washing station step 1 5 5 Select Calibrate and then Use to proceed with calibration The needle arm will automatically move towards the washing station and should be located directly above the center of the rear opening of the washing station DAN SEN pra Lire casted cabron vate 2 rec cr reset 4 e Calbesta wadhang taden ION Figure 12 4 Calibration of the washing station To use already saved settings select Use To reset calibration settings not recommended select Reset 6 Select Height and check the central positioning of the uptake needle by using the buttons Move up and Move down Select Position and adjust by using the Move back and Move fwd buttons Select Done after making necessary adjustments Figure 12 5 Test calibration of the needle arm position in the washing station The position of the uptake needle should appear as shown above 12 Hardware and software installation 12 203 204 7 Select Save to save the new configuration Catbration 1 5 Washing eaten Save new valve 3 out 0 Calbrata washing sation O 1 10 Figure 12 6 Saving new settings for calibration ofthe washing station Calibration 1 Calibration of the tube rack step 2 5 8 Select Calibrate to continue with the calibration process 9 Place a MACS Coo
162. ol f r Quetsch und Schergefahr 1 3 4 Laserstrahlung WARNUNG Das Ger t ist zur automatischen Reagenzglasgestellerken nung Klasse 1M mit vier oberfl chenemittierenden Lasern mit vertikalem Resonator VSCLs ausgestattet Die Strahlung ist nicht sichtbar Schauen Sie nicht direkt mit optischen Instrumenten z B 1 Important information 43 Objektiven Vergr erungsgl sern und Mikroskopen hinein Das Hineinschauen mit optischen Instrumenten in die VSCL ffnung innerhalb eines Abstandes von 100 mm kann Ihre Augen sch digen Das Ger t ist auch mit einem 2D Code Leser ausgestattet der einen sichtbaren Halbleiterlaser als Target Pointer Zeiger zur Einstellung der Leseposition sowie leistungsstarken Leuchtdioden LEDs zur Ausleuchtung des Lesefeldes verwendet Schauen Sie weder direkt in die Laser oder LED Strahlung noch in eine durch eine Spiegelfl che reflektierte Laser oder LED Strahlung Dies kann sonst zu Augenverletzungen f hren Richten Sie den Laserstrahl nicht absichtlich auf andere Personen Demontieren wechseln oder entfernen Sie nicht eingebaute Laser oder LED Strahlungsquellen oder deren Halterung Die Laser oder LED Strahlungsquellen h ren bei Demontage nicht automatisch auf Strahlung zu emittieren Strahlung von demontierten Ger ten kann zu Augenverletzungen f hren Achten Sie auf den optischen Weg des Laserstrahls oder der Reflek tion durch eine Spiegeloberfl che Achten Sie bei der Einrichtung d
163. olumn connector and fastened to the point of resistance If this is not the case loosen the column connector insert the column precisely and tighten the connector again 2 Run the Qrinse program select the Separation menu from the upper navigation bar and Wash now from the lower navigation bar Select Qrinse and press Run Check if the leakage persists If so unscrew the column and check if the luer connectors of the columns are damaged If this is the case exchange the leaking column with a new autoMACS Column refer to section 3 3 4 3 Check if the column connector is fastened properly If not use second wrench to counter and tighten another quarter turn 4 Ifthe problem persists contact Technical Support 10 Troubleshooting 173 174 Note Do not remove the connector from the tubing Note Please note that each tubing has a specific length and should be exchanged with the corresponding spare part only 10 1 2 Tubing leakage 1 Identify the location of the leaky tubing by running the Rinse program From the Separation menu select Wash Now lower menu bar e Highlight Rinse and click Run 2 Check whether the tubing is tightened properly If this is not the case tighten the tube connector The connector should be inserted precisely 3 Ifthe problem persists loosen the tube connector and pull back the connector from the tubing 4 Check the ends of the tubing for wear and fissures If necessary replace tubing
164. on specifically retained cells or choose a double column separation like PosselD PosselD2 or PosselDS Especially when using autolabelling 1 Check that the recommended maximal sample processing time of 90 minutes was not exceeded In the event of an elevated room temperature the cooling capacity of the Chill Racks may be compromised Higher labeling temperatures could cause unspecific binding which may lead to lower purities when using positive selection kits Check whether the correct sample volume was programmed Higher sample input volumes will result in an excess concentration of MicroBeads leading to non specific labeling Recovery of magnetically labeled cells is low Refer to section 10 3 1 Positive cells have not been retained on the column Viability of magnetically labeled cells is low Dead cells have been co enriched in the positive fraction due to non specific binding Remove dead cells before separation by using the MACS Dead Cell Removal Kit Refer to section 10 2 2 This is prone to occur if the number of dead cells in the starting material exceeds the number of target cells 10 Troubleshooting Overall recovery is low Cells are often lost during washing steps Determine the number of cells immediately before magnetic separation to check whether the low recovery is due to this or due to problems during the cell separation 10 3 2 Depletion Too many cells are retained on the column 1 Magnetic labeling of the
165. onali o morte Pericolo di schiacciamento o di taglio Rischio di schiacciamento o di taglio di parti del corpo a causa di componenti meccanici Forte campo magnetico Il campo magnetico pu interferire con oggetti magnetizzati e dispositivi elettronici o danneggiare i supporti magnetici per le informazioni Rischio di lesioni personali gravi per soggetti portatori di pacemaker o impianti medici elettronici Rischio di contaminazione in caso di utilizzo di materiale a rischio biologico Indica il rischio di morte lesioni gravi a danno dell operatore o danni alle attrezzature a causa di materiale biologico potenzialmente pericoloso Indica il rischio di morte o di lesioni gravi a danno dell operatore a causa di un livello pericoloso di tensione Terminale per conduttore di protezione Il simbolo affisso all interno dello strumento Avvertenza per il personale di assistenza tecnica On acceso alimentazione Off spento alimentazione E necessario consultare la documentazione prima di procedere con l installazione e l utilizzo dell apparecchio 1 Important information 1 2 Avvertenze e precauzioni 1 Il separatore autoMACS Pro Separator utilizza tecnologie all avanguardia Si tratta di un dispositivo gestito mediante computer per la separazione automatica delle cellule marcate magneticamente BEE mediante la tecnologia MACS Il campionatore MACS MiniSampler collegato al separatore autoMACS Pro Separator e rappresenta pe
166. onamenti o danni provocare l usura prematura e una minore durata di esercizio dello strumento nonch invalidare la garanzia 1 Important information 59 60 Per i contratti di assistenza e manutenzione offerti da Miltenyi Biotec contattare il rappresentante Miltenyi Biotec locale o consultare il sito www miltenyibiotec com support IMPORTANTE qualora si rendano necessari la sostituzione o l impiego di pezzi di ricambio accertarsi che il tecnico utilizzi unicamente pezzi di ricambio originali Miltenyi Biotec o ricambi di altri fabbricanti specificati e raccomandati da Miltenyi Biotec L impiego di pezzi di ricambio non autorizzati pu causare il malfunzionamento del dispositivo e compromettere il risultato della separazione cellulare Miltenyi Biotec non fornisce alcuna garanzia n si assume la responsabilit per eventuali guasti o danni derivanti dall impiego di pezzi di ricambio inappropriati Al termine dell intervento di assistenza o di riparazione chiedere al tecnico Miltenyi Biotec autorizzato di eseguire tutti i controlli di sicurezza previsti dalla procedura di riparazione onde assicurarsi che lo strumento funzioni correttamente Utilizzare esclusivamente le funzioni opzionali e gli aggiornamenti raccomandati da Miltenyi Biotec 1 5 3 Pulizia Staccare il autoMACS Pro Separator dalla presa di corrente prima di eseguire la pulizia Non utilizzare detergenti liquidi o aerosol utilizzare sempre un panno umido 1 5 4 Mate
167. one dello strumento autoMACS Pro Separator seguenti simboli di avvertenza e attenzione servono quale ausilio alla prevenzione di lesioni personali e danni al dispositivo 1 1 Simbolie livelli di pericolo Indicazioni di sicurezza Esempio CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard Le indicazioni di sicurezza informano l operatore dei rischi potenziali derivanti dal mancato rispetto delle avvertenze e precauzioni descritte di seguito L icona sul lato sinistro specifica il tipo di rischio Il livello di rischio in alto classifica il tipo di pericolo come indicato in precedenza Il livello il tipo e la fonte del pericolo nonch le possibili conseguenze i divieti e le misure da adottare vengono indicati come segue 1 Important information 51 52 CAUTION WARNING e0 P p BP Simboli e livelli di pericolo Di seguito viene presentato un glossario illustrato che descrive i simboli utilizzati nel presente manuale d uso e sul autoMACS Pro Separator Indica una situazione pericolosa che potrebbe comportare lesioni di grado lieve o moderato qualora non venga evitata Indica una situazione pericolosa che potrebbe comportare lesioni gravi o morte qualora non venga evitata Attenzione consultare il Manuale d uso per ulteriori istruzioni e procedere con cautela Le avvertenze includono il rischio di danni alle attrezzature gravi lesioni pers
168. ower speed exchange negative tube with an empty tube and touch RETRY Exchange rack with rack holding more samples and touch OK or touch CANCEL to abort and reprogram Check configuration and connection of the protective cover the MACS MiniSampler and the front support at the autoMACS Pro Separator then touch CONTINUE Touch CANCEL to abort Error code 7027 7028 7029 7030 7031 7032 7033 7034 7035 Cause The protective cover of the MACS MiniSampler needs to be closed Bar code on chill rack could not be read MACS MiniSampler is not connected properly Unable to read rack bar code correctly The instrument has been shut down without using SLEEP The given whole blood sample size exceeds the maximum volume specified The sample cannot be diluted sufficiently Air in system during calibration of tubing Date and time is outdated Air intake during sample uptake although needle did not yet hit bottom of tube Leakage of air into system in front of bubble sensor liquid level has been overestimated or needle did not move to bottom as fast as neccessary Air intake during sample uptake although needle did not yet hit bottom of tube Leakage of air into system in front of bubble sensor liquid level has been overestimated or needle did not move to bottom as fast as neccessary Uptaken sample has been processed Portion of sample is remaining in tube 10 Troubleshoo
169. paration and wash programs highlight the The autoMACS Pro Separator automatically detects the type of tube program of interest and rack in use and allows the user to utilize only the number of samples press the information and sample positions the tube rack can handle If the tube rack does button located not match the template definition a warning screen will be displayed between the arrows A upon starting the separation pop up window will be displayed Alternatively 6 6 1 Entering sample separation instructions refer to section 6 3 3 for a workthrough example more information on In the following example two samples were placed on positions separation programs 1 and 2 of a Chill Rack 15 Sample 1 was already manually labeled using CD4 MicroBeads human Sample 2 has not yet been labeled the autoMACS Pro Separator will perform autolabeling using CD4 MicroBeads human 1 Highlight sample positions 1 and 2 on the sample template 6 Cell separation using the autoMACS Pro Separator 137 138 Note The MACS Reagent CD4 MicroBeads human was identified by the 2D code reader barcode reader The vial was placed on position 1 of the MACS Reagent Rack Refer to section 4 5 for full details on using the Reagent menu EE gt Figure 6 22 Sample positions 1 and 2 were selected The purpose of this example is to demonstrate how sample processing conditions can be easily modified and to clarify the differences between autola
170. pas les tuyaux pendant le fonctionnement de l appareil D connectez toujours l appareil avant de modifier une partie du syst me fluidique Stoppez ou annulez toujours le proces sus avant de manipuler les appareils suppl mentaires comme le mini chantillonneur MACS ou de charger retirer les prouvettes du r telier prouvettes place sur l chantillonneur Ne bloquez pas les dispositifs de s curit et respectez les mesures de s curit Figure 1 2 Le symbole d avertissement entour d un cercle blanc indique un risque d crasement et de cisaillement 1 Important information 31 32 1 3 4 Rayonnement laser AVERTISSEMENT L appareil est quip pour la d tection automatique de porte prouvettes classe 1M par quatre diodes laser mission par la surface cavit verticale VCSEL Le rayonnement est invisible Ne pas regarder directement dans ce rayon au travers d instruments optiques p ex lentilles verres grossissants et microscopes Regarder dans l ouverture du VCSEL une distance inf rieure 100 mm au travers d instruments optiques peut endommager les yeux L appareil est galement quip d un lecteur de code 2D utilisant un laser semi conducteur visible comme pointeur pour le r glage de la position de lecture ainsi que de diodes lectroluminescentes DEL haute puissance pour l clairage du champ de lecture Ne diriger le regard ni dans le rayon laser ou DEL direct ni le rayon laser ou DEL r f
171. place pump head with the spare part 8 Reassemble in reverse order Take care not to pinch the tubing at the bottom left when pushing the bottom cover back into place 7 4 4 Exchange of hydrophobic air filter Hydrophobic air filters 0 2 uM are attached to the bottle closures to vent the liquid bottles and to prevent release of aerosols To avoid 7 Maintenance Note The hydrophobic air filters should be exchanged once a year clogging of the filters and to prevent contamination of liquids air J filters should be exchanged if they come into direct contact with any liquid i e become wet They also should be exchanged once a year to avoid clogging through dust deposits 1 Hydrophobic air filters are attached to all fluid containers Figure 7 24 Hydophobic air filter attached to a fluid container 2 Unscrew the hydrophobic air filter from the fluid container Replace the new filter in a similar manner 7 4 5 Exchange of autoMACS Columns The autoMACS Columns should be exchanged after 100 separations or two weeks after the last column exchange whichever comes first The procedure is described in detail in section 3 3 4 7 5 Exchange of the tubing system If there is any leakage in the tubing system the affected tubing part should be exchanged Please note that each tubing has a specific length and should be exchanged exclusively with the corresponding spare part 1 Switch OFF and unplug the instrument 2 Remove
172. pler and tube racks are automatically detected by the autoMACS Pro Separator adding to its operational efficiency The user manual aims at explaining the principles of cell sorting with the autoMACS Pro Separator and to assist you by providing step by step protocols for cell labeling separation and quality control Tips and hints in the troubleshooting section as well as general protocols and special separation protocols are intended to help optimize your magnetic cell separations uw Figure 2 1 Front image ofthe autoMACS Pro Separator the access cover and bottom cover were made transparent for the purpose of illustration 2 Introduction 2 77 78 autoMACS Pro Starting Kit contains autoMACS Pro Separator MACS MiniSampler 130 092 545 MACS Reagent Rack 4 1 MACS Cooling Tube Rack Chill 5 for 5 mL tubes 1 MACS Cooling Tube Rack Chill 15 for 15 mL tubes 1 MACS Cooling Tube Rack Chill 50 for 50 mL tubes Software Barcode reader 5x2 autoMACS Columns 2x2 column substitutes autoMACS Pro Buffer Combination user manual and short instruc tions one year warranty Technical specifications of the MACS Cooling Tube Racks and Reagent Racks MACS Slots Cooling Tube Racks Chill 5 24x5 mL Chill 15 15x15 mL 5x5 mL Chill 50 6x50 mL 3x15 mL 3x5 mL Max number Max sample Max number ofsamples volumeper of cells per tube tube 6 5mL 2 5mL 5x10 tubes 5 15 mL 12 5 mL 2 5x10 tube
173. positive fraction collection tube row C of the tube rack In the double positive selection programs Posseld Posselds Posseld2 and Posselwb the magnetically labeled target cells are first retained in the autoMACS Column 1 The negative fraction containing the non labeled cells is retrieved in the negative fraction collection tube row B of the tube rack Then the magnetically labeled cells are held in a reservoir and loaded onto the autoMACS Column 2 Finally the magnetically labeled cells are eluted into the positive fraction collection tube row C of the tube rack Depletion programs When running any depletion program the magnetically labeled non target cells are retained in the autoMACS Column 1 The non labeled target cells pass through the column and are released into the negative fraction collection tube row B of the tube rack The magnetically labeled fraction containing the unwanted cells is eluted into the positive fraction collection tube row C of the tube rack 6 Cell separation using the autoMACS Pro Separator 131 132 Program Volume of non Volume of Loading labeled fraction labeled rate i e negative fraction fraction i e positive fraction Deplete 2mL samplevolume 2mL 4 mL min Depletes 2mL samplevolume 2mL 1 mL min Depl05 2mL samplevolume 2mL 0 5 mL min Depl025 2mL samplevolume 2mL 0 25 mL min A_Depl07 3mLper1mLsample 2mLper1 4 mL min volume mL sample volume A_D
174. ppareil pourrait sinon chuter provoquant des blessures corporelles graves et ou la d t rioration consid rable l appareil Utilisez le s parateur unique ment sur une table un chariot un pied un tr pied ou un support recommand par Miltenyi Biotec ou vendu avec l appareil Ne placez pas le s parateur autoMACS Pro Separator dans un appareil int gr ou un espace restreint comme un casier sauf si le dispositif a t 1 Important information 34 sp cialement con u pour accueillir l appareil qu une ventilation appropri e est garantie et que les instructions de montage relatives l appareil ont t respect es 1 4 2 Circulation de l air Ne placez pas l appareil proximit de radiateurs de registres de chaleur de fours ou d autres pi ces d quipement amplificateurs qui produisent de la chaleur Veillez ce que suffisamment d air puisse circuler autour du s parateur autoMACS Pro Separator au moins 15 cm de chaque c t pendant le fonctionnement afin de garantir le refroidissement ad quat de l appareil vitez d exposer l appareil un ensoleillement direct Les encoches et les fentes de l appareil sont pr vues pour la ventilation et ne doivent jamais tre bloqu es ou recouvertes car elles assurent le fonctionnement fiable du s parateur autoMACS Pro Separator et prot gent l appareil de la surchauffe N introduisez jamais d objet tranger dans l appareil par l une des fentes 1 4 3 Eau et humidi
175. pport 10 2 2 Low cell viability in final fraction 1 Low cell viability can result from problems during both the cell preparation and the autoMACS Pro Cell Separation Hence also check section 10 3 2 Check the 70 ethanol Make sure that analytical reagent grade ethanol without additives is used to prepare the solvent 10 Troubleshooting 177 178 8 Check the Running Buffer Make sure that the appropriate buffer has been used and that no contamination has occurred Check Washing Solution Make sure that the appropriate solution has been used and no contamination has occurred Check the pump syringe for contamination If contamination is obvious refer to section 10 1 3 and change columns afterwards Check for contamination of the tubing system and refer to the appropriate section Run the Safe program to decontaminate the fluidic system Check the column status The autoMACS Columns should be exchanged after 100 separations or two weeks after the last column exchange whichever comes first The procedure is described in detail in section 3 3 4 If the problem persists contact Technical Support 10 2 3 Low purity of isolated cell population 1 Low purities can be caused by problems during both the cell preparation and the autoMACS Pro Separation Hence also check section 10 3 Low purities can also result from using an inappropriate separation program or labeling approach Please also refer to section 6 Ch
176. pump Computer controlled high precision syringe 1 5 pump with Teflon seal plunger that drives fluids through the autoMACS Pro fluidic system Touchscreen High resolution TFT color touchscreen located on top of the autoMACS Pro Separator The touchscreen is used to operate and monitor the instrument through on screen menus Tubing connector Plastic threaded connector with square nut used to connect the tubings to the bottle closures the columns the pump or valves Tubing system Permanent set of Teflon tubing through which fluids circulate in the autoMACS Pro Separator fluidic system Tube racks Three different acrylic tube racks are provided with the instrument They are designed for optimal positioning of sample tubes and fraction collection tubes at the ports of the automated arm They contain a coolant allowing to pre cool the racks in the refrigerator for cooling of the cells during the separation process The racks have four tube positions for each sample Position A holds the sample tube containing the starting material Position B holds the tube for the non labeled fraction Position C holds the tube for the magnetically labeled fraction Position D is auxiliary For details refer to the table in section 4 1 2 Washing Solution bottle Container for autoMACS Pro Washing Solution The bottle closure is equipped with a fluid sensor The closure the fluid sensor cable and the tubing connector are color coded green Waste
177. r tes entsprechend den f r die verwendeten Substanzen geltenden Warnhinweisen und Schutzbestimmungen eine pers nliche Schutzausr stung tragen Tragen Sie Schutzhand schuhe Schutzkleidung und Schutzbrille um Ber hrung der Gefahrenstoffe mit Haut und Augen zu vermeiden Mangelhafte oder unzureichende Schutzausr stung kann den Bediener des Instru mentes gef hrden Werden biologische Gefahrenstoffe oder unbekannte Substanzen eingesetzt sollten Sie mit dem AutoMACS Pro Separator in einer Sterilbank arbeiten Falls Gefahrstoffe verwen det wurden oder ausgetreten sind achten Sie auf eine sorgf ltige Dekontaminierung des Ger tes S ulen Auffanggef e und alle weiteren Verbrauchsmaterialien die in Kontakt mit biologischen Gefahrenstoffen gelangt sind sollten vor Entsorgung autoklaviert werden Fl ssigabfall sollte autoklaviert oder unter Verwendung eines f r das jeweilige spezifische Pathogen geeigneten Desinfektionsmittels dekontaminiert werden z B 10 Bleichmittel Isopropylalkohol oder 70 Ethanol 1 Important information N mate MACS Pra Separator NA ose EHRE w COTE CE Nema RC rd Venne Patan ria Figure 1 4 Die Entsorgung der Verbrauchsmaterialien muss gem lokal geltender Bestimmungen erfolgen 1 5 5 Transport Der autoMACS Pro Separator sollte vorsichtig gehandhabt in der von Miltenyi Biotec bereit gestellten Verpackung transportiert werden Im Ger t k nnen innere Sch den auftreten falls es g
178. r des Ger ts zug nglichen Ort auf so dass diese k nftig jederzeit darauf zur ckgreifen k nnen ACHTUNG Der autoMACS Pro Separator ist ausschlie lich ausgelegt f r den Betrieb in Innenr umen Bitte benutzen Sie das Ger t nicht in ausgewiesenen Gefahrenzonen wie etwa sauerstoffangereicherten Arbeitsumgebungen Wenden Sie sich an die rtlichen Beh rden und Ihren Stromversorger f r weitere Informationen zur Stromversorgung Geb udeinstalla tionen Wartung und Sicherheit f r die Installation dieses Ger tes Wenn Sie Sicherheitsbedenken in Bezug auf die Ger tenutzung haben setzen Sie sich bitte mit Ihrem autorisierten Miltenyi Biotec Service Provider in Verbindung oder kontaktieren den Miltenyi Biotec Customer Support 1 3 Allgemeine Sicherheitshinweise Um m gliche mit dem Betrieb des autoMACS Pro Separators verbun dene Sicherheitsrisiken zu verringern beachten Sie bitte die folgenden allgemeinen Sicherheitshinweise Eine Nichteinhaltung dieser Vorsichtsma nahmen k nnte Feuer Gesundheitssch den und oder Sch den am Ger t verursachen 1 Important information 41 1 3 1 Gefahr des Stromschlags und der Ausbreitung von Feuer WARNING Hazardous voltages Risk of loss of live severe personal injury Unplug before cleaning Do not use if device is opened or damaged if liquids have been spilled into device if objects device through ventilation slots WARNUNG Der Umgang mit elektrischen Ger ten birgt d
179. rack position For each sample rack position it is possible to define cell labeling cell separation and washing programs 4 An overview of the autoMACS Pro touchscreen user interface 99 100 A single wash program can be performed by selecting Wash Now from the lower navigation bar EE Os 29 Figure 4 13 The separation menu An overview of the separation menu follows H Sample rack template IZ Sample labeling options 3 Sample processing volume H Separation program 5 Wash procedure e Sample rack template The positions in the programming field 7 correspond to the sample positions in the tube rack Positions 1 6 are used in combination with tube rack Chill 5 positions 1 5 with rack Chill 15 and positions 1 3 with rack Chill 50 1 Toselect deselect a sample position touch to highlight the desired position 1 6 2 Select deselect a sample position by touching the display RS gt Figure 4 14 Sample positions 1 3 and 4 were selected 4 An overview of the autoMACS Pro touchscreen user interface Sample labeling options A The Labeling submenu 2 is used to instruct the instrument if autolabeling is to be performed on a sample The default setting is which indicates that NO autolabeling will be performed the type of autolabeling that will be performed A list of product options is only visible if reagents have already been assigned to positions on the reagent rack using the Reagent m
180. re Safe Storage 70 v v ethanolin Black Not available solution distilled water prepared from absolute ethanol Table 5 1 Solutions required for daily operation Solution bottles can be identified by color code and symbols table 3 1 Symbols and color coding of fluid containers For proper operation of the autoMACS Pro Separator solution containers must be filled with a minimum volume of 150 200 mL It is recommended to use ready to use MACS Buffers or fresh 5 Materials required 113 114 Note MACS Running Buffer contains azide preservative A solution without preservative may be prepared by diluting MACS BSA Stock Solution 130 091 376 1 20 with autoMACS Rinsing Solution 130 091 222 Note Do not use denatured ethanol technical ethanol as the autoMACS Columns are not resistant to oxidative compounds Use 100 ethanol analytical reagent grade without additive Note Program Safe can also be used for decontaminating the autoMACS Pro fluidic system For special decontamination procedures contact the technical support team for further advice filter sterilized solutions to prevent potential contamination of the tubing system The autoMACS Pro Washing Solution is a filter sterilized and ready to use solution to rinse the fluidic system after any autoMACS Pro Cell Separation It contains a detergent that dissolves cell aggregates The autoMACS Pro Washing Solution was developed for optimal cleanin
181. re 3 20 Selecting column exchange from the Option tab 3 Assembly and installation 5 Select Run to start program Col_ex A popup dialog offers four 3 options Remove to remove columns Install to remove and install new columns Sep count to inform the software how many separations have been performed Exit to cancel this process Figure 3 21 Select column action dialog box Note Only install one 6 Select Install from the popup dialog box column at a time 7 Wait until the instrument prompts you to exchange the autoMACS Columns before proceeding Select Done 8 Using both hands take the top and bottom of the column one substitute and pull gently but firmly to remove it from its slot j 9 Place a wide mouth container under the column substitute Hold the column substitute in one hand and gently unscrew the bottom column connector counter clockwise Unscrew the top column connector while holding the column over the container to catch any fluids Store the column substitute in the autoMACS Pro Starting Kit box 3 Assembly and installation 91 Note The program will then proceed to wash the columns with autoMACS Running Buffer Check that the column is securely fastened to the column connectors and that no buffer is leaking Note Read the warnings and precautions section before priming the autoMACS Pro Separator for the first time 92 Figure 3 22 Unscrewing the top column connector for column
182. re o altre sostanze estranee si depositino sulla finestra di lettura del lettore di codici a barre 2D Ci potrebbe comportare errori di lettura Accertarsi di aver arrestato l emissione laser prima di pulire il lettore Un esposizione al raggio laser potrebbe danneggiare i propri occhi Utilizzare un panno morbido e asciutto per rimuovere qualsiasi sostanza dal lettore di codici a barre Non utilizzare alcol o altre sostanze detergenti Il separatore autoMACS Pro classificato come prodotto laser della classe 1M secondo lo standard IEC 60825 1 1993 A1 1997 A2 2001 ATTENZIONE L utilizzo di comandi o regolazioni o l esecuzione di procedure diverse da quanto qui specificato pu essere causa di esposizione a radiazioni pericolose 1 Important information Figura 1 3 Posizione dei laser Laser invisibili per il rilevamento dei portacampioni portareagenti sono collocati nell area indicata dal rettangolo Il lettore per codice 2D visibile collocato all interno del cerchio A E 1 4 Installazione sicura Questa sezione descrive i requisiti che deve soddisfare il luogo in cui viene collocato il autoMACS Pro Separator onde garantirne un installazione sicura e il funzionamento corretto Leggere le istruzioni fornite nella presente sezione e accertarsi che il luogo prescelto sia adeguatamente predisposto prima di collegare lo strumento alla presa di alimentazione Nel predisporre l ambiente di installazione e nell organiz
183. riale pericoloso WARNING Risk of loss of life if biohazardous material is used Wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Operate device in a safety hood Decontaminate device after spilling of biohazardous material Qualora si utilizzi o sia stato usato del materiale a rischio biologico l operatore deve indossare dispositivi di protezione personale conformi alle avvertenze e alle precauzioni relative alle sostanze impiegate Indossare guanti protettivi indumenti e occhiali di protezione per prevenire il contatto con la pelle e gli occhi L impiego di dispositivi di protezione difettosi o inadeguati rappresenta un rischio per l incolumit dell operatore Lo strumento autoMACS Pro Separator va utilizzato con uno schermo protettivo in caso di trattamento di materiali sconosciuti o pericolosi Qualora sia stato usato o versato del materiale pericoloso il sistema deve essere accuratamente decontaminato Le colonne le piastre le provette e tutti gli altri materiali di consumo entrati in contatto con campioni a rischio biologico vanno sterilizzati in autoclave prima di essere smaltiti rifiuti liquidi vanno autoclavati o decontaminati utilizzando un disinfettante idoneo per il patogeno 1 Important information specifico ed es candeggina al 10 alcol isopropilico o etanolo al 1 70 Figura 1 4 Lo smaltimento dei rifiuti deve avvenire in conformit alle disposizio
184. rnatant completely 8 Protocols 11 Resuspend up to 10 cells in 500 uL of buffer 8 Note For higher cell numbers scale up buffer 12 Proceed to magnetic separation refer to section 5 3 5 volume accordingly 8 3 Direct magnetic labeling of human cells using MACS Whole Blood MicroBeads MACS Whole Blood MicroBeads have been developed for positive selection of many human cell types from anticoagulated whole blood samples or bone marrow samples Perform magnetic labeling with MACS Whole Blood MicroBeads according to the following general instructions Refer to the specific MACS Cell Separation Reagent data sheet for more details 1 Prepare the sample as outlined by the data sheet T 2 Add 50 uL MACS Whole Blood MicroBeads per 1 mL anticoagu lated whole blood When working with larger or smaller blood volumes scale all reagent volumes and total volumes up or down accordingly e g for 500 uL of blood use half the volume of Whole Blood MicroBeads i e 25 uL 3 Mix well and incubate for 15 minutes in the refrigerator 2 8 C 4 Wash cells by adding 2 5 mL of buffer per mL whole blood and centrifuge at 445xg at room temperature for 10 minutes and slow deceleration no brake 5 Aspirate supernatant carefully Do not disturb the cell pellet Leave a small residual volume of supernatant approximately 1 2 mm in height to avoid cell loss 6 Resuspend the cell pellet estimate the volume and dilute with the same volume of b
185. ro er Ersch tter ung ausgesetzt oder fallengelassen wird Sollte wegen Reparatur oder Wartungsleistungen ein R cktransport zum Hersteller notwendig werden dekontaminieren Sie das Ger t vor Versand von jeglichen biologischen Gefahrenstoffen Wenn Sie Fragen zur vorschriftsm igen Dekontaminierung oder zum Versand des Ger tes haben wenden Sie sich bitte direkt an unseren Technical Support 1 5 5 Ger teentsorgung Setzen Sie sich bitte direkt mit unserem Technical Support in Verbindung falls Sie Ihr Ger t entsorgen m chten 1 Important information 49 50 1 Important information Informazioni importanti 1 Leggere prima dell uso Si prega di leggere tutte le informazioni riportate nel presente manuale d uso prima dell utilizzo La mancata lettura e l inosservanza delle istruzioni possono condurre ad un impiego improprio o scorretto dello strumento a manipolazione e manutenzione inadeguate o errate e potrebbero rappresentare un pericolo per l operatore avere conseguenze imprevedibili causare il malfunzionamento o danni allo strumento condurre ad un usura prematura e ad una minore durata di esercizio dello strumento nonch invalidarne la garanzia Conservare il presente manuale in un luogo sicuro accessibile a tutti gi addetti che utilizzano l unit di separazione autoMACS Pro autoMACS Pro Separator Il presente capitolo illustra le istruzioni di sicurezza e i requisiti ambientali per l installazi
186. ro Separator enth lt einen extrem starken Magneten Halten Sie mit magnetischen Datentr gern Kreditkarten Magnetb nder Speichermedien und dergleichen und elektronis chen Ger ten wie H rger te Herzschrittmacher Mess und Steuerg er te PCs Uhren mindestens 20 cm Abstand zur Magnetabdeckung da diese durch das Magnetfeld gest rt und gesch digt werden k nnen 1 Important information Figure 1 1 Warnsymbol f r starken Dauermagnet 1 3 3 Quetsch und Schergefahr CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard WARNUNG Frontabdeckungen nicht ffnen wenn das Ger t l uft Bewegung des Automatikarms und der zugeh rigen Komponenten im Betrieb nicht behindern Finger usw von allen bewegten Teilen des autoMACS Pro Separator und zugeh rigen Komponenten fernhalten es besteht sonst Gefahr von Quetsch und Scherverletzun gen und Sch den am Ger t Fl ssigkeitspumpen nicht ber hren und Leitungen nicht ver ndern wenn das Ger t l uft Ger t immer ausschalten bevor Arbeiten am Fl ssigkeitssystem vorgenommen werden Laufenden Vorgang immer anhalten oder abbrechen bevor Arbeiten an Zusatzger ten wie dem MACS MiniSampler ausgef hrt oder Reagenzgl ser aus dem Reagenzglasgestell im Sampler entnommen oder dort eingesetzt werden Sicherheitsma nahmen und vorrichtungen niemals umgehen oder manipulieren Figure 1 2 Der Kreis zeigt das Warnsymb
187. rtanto un componente del dispositivo di separazione delle cellule Il separatore autoMACS Pro Separator e il campionatore MACS MiniSampler sono concepiti in modo da garantire un funzionamento sicuro dopo l installazione se utilizzati da personale addestrato secondo le norme generali di sicurezza e le istruzioni fornite nel presente manuale d uso Le linee guida riportate nella presente sezione illustrano i potenziali rischi associati al funzionamento dello strumento e forniscono importanti informazioni in materia di sicurezza atte a ridurre al minimo detti rischi Seguendo scrupolosamente le istruzioni possibile proteggere se stessi e le attrezzature dai possibili pericoli e garantire un ambiente di lavoro sicuro Qualora lo strumento venga impiegato in modo non conforme alle istruzioni fornite dal fabbricante la sicurezza potrebbe risultare compromessa IMPORTANTE leggere e seguire tutte le istruzioni operative contenute nel presente manuale e prestare particolare attenzione alle avvertenze indicate sullo strumento Conservare il presente manuale d uso e tutte le altre istruzioni operative e di sicurezza fornite insieme alla strumento in un luogo accessibile a tutti gli operatori per future consultazioni IMPORTANTE autoMACS Pro Separator sono destinati unicamente all impiego interno Non utilizzare lo strumento in ambienti classificati come luoghi pericolosi quali gli ambienti con forte concentrazione di ossigeno Per ulteriori inform
188. rting the cell separation The target cell recovery e g positive cells in the magnetically labeled cell fraction can be calculated as follows No of cells in pos fraction x positive cells in pos fraction Target cell recovery 100 x No of cells in orig sample x positive cells in orig sample The overall cell recovery can be calculated as follows No of cells in pos fraction No of cells in neg fraction Overall cell recovery 100 x No of cells in orig sample 9 2 Purity of isolated cell population For most experiments that follow the cell separation it is necessary to document the purity of the isolated cell subset It is recommended to analyze the cells by flow cytometry Alternatively fluorescence microscopy can be used Cells can also be analyzed by immunocytochemistry 9 Quality control of separations performed with the autoMACS Pro Separator 171 172 Note Efficient staining of the cells with regard to signal intensity and specificity are the pre requisites for an accurate analysis Magnetic cell separation using the MACS Technology is a highly sensitive method and often results in extremely pure cell populations Make sure that the method used for analysis is sensitive enough to accurately analyze the purity Purity of the positively selected cell fraction Purity positive cells in positive magnetically labeled fraction Purity of the depleted cell fraction
189. rwandten Symbole bildlich dargestellt und erkl rt Bezeichnet eine Gefahrensituation die falls sie nicht vermieden wird zu leichteren oder mittelschweren Verletzungen f hren kann Achtung schlagen Sie im Benutzerhandbuch f r weitere Anweisun gen nach und gehen Sie umsichtig vor Warnhinweise deuten auf das Risiko von Besch digungen des Ger tes bzw der Ausr stung die Gefahr von schweren Verletzungen oder Lebensgefahr hin Quetsch und Schergefahr Durch mechanische Gef hrdungen k nnen Br che oder Quetschun gen von K rperteilen verursacht werden Laserstrahlung Gefahr von schweren Augen und Hautverletzungen Starkes Magnetfeld Das Magnetfeld kann auf andere magnetisch aufladbare Gegen st nde und elektronische Ger te st rend einwirken oder magne tische Datentr ger besch digen F r die Tr ger eines Herzschrittmachers oder von eletronischen Implantaten besteht die Gefahr schwerer Verletzungen Gefahr der Kontaminierung bei Verwendung von biologischen Gefahrenstoffen Dieses Symbol weist auf Lebensgefahr das Risiko schwerer Verletzungen f r den Bediener des Ger tes oder Sch den am Ger t selbst hin die durch Arbeit mit m glicherweise gef hrli chen biologischen Substanzen entstehen k nnen Weist auf Lebensgefahr oder das Risiko schwerer Verletzungen f r den Benutzer des Ger tes aufgrund gef hrlicher Stromspannung hin Gleichstrom Mit diesem Symbol ist auf dem Ger t die Art der Energieversorgung
190. s 3 50 mL 50 mL 4x10 tubes MACS Reagent Rack Reagent Rack 4 4 reagent vials Technical specifications of the autoMACS Pro Separator the instrument is UL listed and CE marked Programs 12 pre set Protocol Column capacity Processing volumes Footprint Single and double column enrichment positive selection and untouched isolation of target cells 4x10 cells sample 2x10 magnetically labeled cells sample up to 15 mL of whole blood or bone marrow 0 2 50 mL input 0 5 52 mL output 605 mmx353 mm wxd Footprint with MACS MiniSampler 605 mmx455 mm wxd Height Weight Input voltage Power consumption 392 460 mm adjustable touchscreen 25 kg 100 240 VAC 50 60 Hz 200 VA Depending on the type of power plug chosen the depth increases by 27 mm or 62 mm 2 Introduction Assembly and installation 3 The autoMACS Pro Separator is a benchtop instrument that fits in laminar flow hoods or safety cabinets If the instrument is placed in a laminar flow hood the following accessories might be required autoMACS Pro Laminar Hood Plate 130 093 246 and optionally autoMACS Pro Angle Connector Kit 130 093 245 The Laminar Hood Plate provides a stable and even surface even on potentially bending surfaces The angle connectors reduce the depth of the instrument to 455 mm including MACS MiniSampler for placement in a location with limited space The operating environm
191. s relatives la sant la s curit et a la pr vention des accidents en laboratoire doivent tre observ es en permanence 1 5 1 S curit d utilisation Si l appareil ne fonctionne pas correctement et que les instructions ou les messages sur l cran d affichage conseillent de contacter le service technique la s curit d utilisation de l appareil n est plus garantie D connectez imm diatement le s parateur autoMACS Pro Separator d branchez l appareil et contactez un prestataire de services Miltenyi Biotec agr ou l quipe du service apr s vente de Miltenyi Biotec 1 5 2 Entretien et r paration IMPORTANT N effectuez pas vous m me l entretien du s parateur autoMACS Pro Separator sauf autre consigne sp cifique donn e dans ce mode d emploi ou dans un autre document de Miltenyi Biotec L entretien et la r paration doivent tre effectu s par des r parateurs qualifi s Tout entretien ou toute r paration inappropri e ou incorrecte de votre s parateur autoMACS Pro Separator peut mettre en danger les utilisateurs fournir des r sultats impr visibles entra ner le dysfonctionnement ou la d t rioration de l appareil son usure pr matur e et r duire la dur e de vie de l appareil ainsi qu annuler 1 Important information 35 36 votre garantie Renseignez vous aupr s de votre repr sentant Miltenyi Biotec local sur les conditions d entretien et de support compl men taires propos es par Miltenyi B
192. s and maintenance programs Select appropriate tube rack Prime the autoMACS Pro Separator Define autoMACS Pro Separation program sequence or template 137 140 140 141 143 143 143 143 Chapter 7 145 145 146 146 146 147 149 149 149 153 152 153 154 154 157 160 160 161 161 161 162 162 163 163 164 166 Chapter 8 167 167 168 1 Important information 6 6 1 6 7 6 7 1 6 7 2 6 8 6 8 1 6 8 2 6 8 3 7 1 7 2 7 2 1 7 2 2 7 2 3 7 3 7 3 1 7 3 2 7 3 3 7 3 4 7 3 5 74 7 4 1 7 4 2 7 4 3 7 4 4 7 4 5 7 4 6 7 5 7 6 7 7 7 7 1 7 7 2 7 8 7 9 8 1 8 2 Entering sample separation instructions a workthrough example Monitoring the autoMACS Pro Separator during cell separation Status menu before separation Status menu during cell separation Shut down the autoMACS Pro Separator Sleep as the final wash program Store the program for long term storage Shutdown button Maintenance General considerations Daily maintenance Instrument priming Clean uptake outlet ports Instrument shutdown Periodic maintenance Column exchange Clean the pump syringe Clean the washing station Instrument decontamination Instrument long term storage Planned maintenance Valve exchange Exchange of dilutor valve Exchange of pump syringe Exchange of the peristaltic pump head Exchange of hydrophobic air filter Exchange of autoMACS Columns Exchange of the tubi
193. s et niveaux de danger Avertissements et pr cautions Pr cautions d ordre g n ral Risque d electrocution et de propagation de feu Champ magn tique puissant Content 31 32 33 33 34 34 34 34 34 34 35 35 35 36 36 37 37 37 39 39 39 40 41 41 42 42 43 43 45 45 45 45 45 46 46 46 47 47 47 48 48 49 49 1 3 3 1 3 4 1 4 1 4 1 1 4 2 1 4 3 1 4 4 1 4 5 1 4 6 1 4 7 1 5 1 5 1 1 5 2 1 5 3 1 5 4 1 5 5 1 5 6 1 5 7 1 1 1 2 1 3 1 3 1 1 3 2 1 3 3 1 3 4 1 4 1 4 1 1 4 2 1 4 3 1 4 4 1 4 5 1 4 6 1 4 7 1 5 1 5 1 1 5 2 1 5 3 1 5 4 1 5 5 1 5 5 Risque d crasement et de cisaillement Rayonnement laser S curit de l installation Accessoires de montage Circulation de l air Eau et humidit Produit reli la terre Sources lectriques Accessibilit Appareils p riph riques S curit d utilisation maintenance transport et limination S curit d utilisation Entretien et r paration Nettoyage Mati res dangereuses Transport Elimination de l appareil Compatibilit lectromagn tique Wichtige Information Symbole und Gefahrenstufen Warnhinweis Symbole und Gefahrenebenen Warn und Sicherheitshinweise Allgemeine Sicherheitshinweise Gefahr eines Stromschlags und der Ausbreitung von Feuer Starkes Magnetfeld Quetsch und Schergefahr Laserstrahlung Sichere Ger teinstallation Montagezubeh r Bel ftung Wassereintritt und
194. scription of how to monitor the instrument s status using the A Status menu follows 4 3 1 Status of fluid containers Figure 4 6 Fluid container status symbols Left Fluid containers are ready Right Fluid containers need replaced The status of fluid containers is indicated by color coded graphic symbols and by a text table Container Symbol Symbol color and user action Running Buffer Green no action required Red refill container Gray connect bottle sensor Washing Solution Gray connect bottle sensor Storage solution Gray no liquid detection visually check volume Waste Green no action required Red empty waste Gray connect bottle sensor Table 4 2 Status of fluid containers displayed in the Status menu In addition to color coded graphic symbols of the fluid containers a popup text table also reports fluid container status Touch any fluid bottle symbol to activate the popup textbox To close the popup table select Moreover the symbols are also red when the sensor cables are connected to a wrong bottle closure 4 An overview of the autoMACS Pro touchscreen user interface 97 98 l Figure 4 7 Fluid container status By touching any fluid container symbol a popup box displays a text report of the fluid container status 4 3 2 Columnstatus Figure 4 8 The column status graphic Left The column is ready Right The column must be changed If the column symbol is green
195. separation results Miltenyi Biotec does not honor any warranty or accept any responsibility for instrument failure or damages resulting from the use of inappropriate replacement or spare parts After completing any service or repair work have your authorized Miltenyi Biotec service provider perform all safety checks required by the repair procedure to ensure that the instrument is in proper operational condition Only use options and upgrades recommended by Miltenyi Biotec 1 5 3 Cleaning Unplug the autoMACS Pro Separator from the outlet before cleaning Do not use liquid or aerosol cleaning agents always use adamp cloth 1 5 4 Hazardous material WARNING Risk of loss of life if biohazardous material is used Wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Operate device in a safety hood Decontaminate device after spilling of biohazardous material If biohazardous material have been used the operator shall choose and wear personal safety equipment in accordance with warnings and precautions for the used substances Wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Also protect mouth and nose as aerosols might leak from the system e g Washing Station Defective or inadequate safety equipment might endanger the operator The autoMACS Pro separator shall be operated in a safety hood if hazardous or unknown materials are pro
196. sequent fluorescence analysis The number should not exceed 2x108 cells per sample If necessary split the sample 5 Cells were not labeled with MicroBeads because the MicroBeads were degraded Check the expiration date Check for sterility of the MicroBeads if the vial has been opened before 6 Cells were labeled e g with fluorochrome conjugated antibodies prior to magnetic labeling with direct MicroBeads When antibodies recognizing the same epitope are used for fluorescent and magnetic labeling fluorochrome conjugated antibodies and MicroBeads might compete for the binding sites This can result in insufficient magnetic labeling It is recommended to perform magnetic labeling prior to staining with fluorochrome conjugated antibodies Alternatively use indirect MicroBeads 7 Anon optimal separation program was chosen e g Possel instead of PosselS for dimly labeled cells Especially when using autolabeling 1 Check whether the uptake needle is positioned in the middle of the sample tube during the mixing procedure If not inadequate mixing will be performed 2 Check whether the reagent volume contents of reagent vial is sufficient for labeling 3 Verify whether the sample volume was programmed correctly A reduced input volume will result in insufficient labeling of cells Low purity of magnetically labeled cell fraction 1 Check for cell aggregates Negative cells may be retained when forming clusters with positive cells
197. setting of this option is described in section 4 7 O_init Optional priming of the instrument at startup 5 Ensure that the MACS MiniSampler is installed correctly For more details concerning correct installation of the MACS MiniSampler refer to section 3 Figure 6 20 MACS MiniSampler status graphic Left The MiniSampler was successfully installed Right No MiniSampler was detected 6 The autoMACS Pro Separator is now ready for priming Select Separation and Wash Now from the lower navigation bar Figure 6 21 Wash Now command displays two options Qrinse quick rinse and Rinse A full rinse is required for instrument priming 7 Select Rinse and Run 6 6 Define autoMACS Pro Separation program sequence or template After completing the priming process or if a wash program has been completed the autoMACS Pro Separator is ready for separation The status of the instrument is displayed in the menu status The bottles are illuminated green Select the Separation menu 1 Select the desired position s in the sample separation template field by touching it 2 Select an autolabeling program and the sample volume a separation program and a washing program for each sample position The selected programs will be displayed in the programming field 6 Cell separation using the autoMACS Pro Separator 3 Optional Templates can be saved by selecting Save template 6 from the lower navigation bar Follow
198. skin and eyes Operate device in a safety hood Decontaminate device after spilling of biohazardous material Si une mati re nocive pour l organisme est ou a t utilis e l op rateur doit choisir et porter un quipement de protection individuelle conforme aux avertissements et aux pr cautions pour les substances utilis es Portez des gants de protection des v te ments de protection et des lunettes de s curit afin d viter tout contact avec la peau et les yeux Un quipement de s curit d fectueux ou inad quat peut mettre l op rateur en danger Le s parateur autoMACS Pro Separator doit tre manipul dans un couvercle de protection si des mati res dangereuses ou inconnues sont trait es Si une mati re dangereuse a t utilis e ou s est d vers e prenez les pr cautions appropri es pour d contaminer soigneusement le syst me 1 Important information Les colonnes les plaques les tubes et tous les autres consommables qui ont t en contact avec des chantillons nocifs pour l organisme doivent tre trait s l autoclave avant l limination Les d chets liquides doivent tre trait s l autoclave ou d contamin s l aide d un d sinfectant adapt l agent pathog ne sp cifique par ex 10 eau de Javel alcool isopropylique ou 70 d thanol 4 mate BACH Pra Separator Figure 1 4 L limination des d chets doit tre effectu e conform ment aux r glementations locales 1 5
199. sschutzbestimmungen Laborrichtlinien Sicherheitsnormen und Unfallverh tungsvorschriften m ssen auf jeden Fall beachtet werden 1 5 1 Sicherer Betrieb des Ger tes Falls Ihr Ger t nicht einwandfrei arbeitet und Anzeigen auf dem Display Sie dazu auffordern den technischen Kundendienst zu kontaktieren ist die Betriebssicherheit nicht mehr l nger gew hrleistet Schalten Sie das Ger t sofort aus unterbrechen die Stromzufuhr und kontaktieren einen autorisierten Miltenyi Biotec Service Provider oder das Miltenyi Biotec Customer Support Team 1 5 2 Wartung WICHTIG Versuchen Sie nicht den autoMACS Pro Separator selbst zu warten oder zu reparieren es sei denn es ist in diesem Benutzer handbuch oder anderen technischen Unterlagen der Miltenyi Biotec GmbH ausdr cklich vermerkt Wartung und Reparaturen m ssen durch geschulte Fachkr fte ausgef hrt werden Falsche oder unsachgem e Wartung oder Reparatur an Ihrem Ger t kann zur Gef hrdung des Anwenders unvorhersehbaren Resultaten Fehlfunk tionen Ger tesch den vorzeitigem Verschlei und verringerter Lebensdauer f hren und kann den Verlust Ihrer Garantieanspr che zur Folge haben Fragen Sie Ihren rtlichen Miltenyi Biotec Vertriebsmitarbeiter nach unseren weit reichenden Vereinbarungen zum Ger teser vice und Kundendienst oder besuchen unsere Website www miltenyibiotec com support WICHTIG Wenn Ersatzteile ben tigt werden stellen Sie sicher dass Ihr Service Pro
200. surface markers can be sorted easily and quickly Even rare cells can be enriched efficiently Multiparameter sorting with MACS MultiSort Kits allows sequential positive selections of cells The target cells are first labeled with MACS MultiSort MicroBeads and positively selected for the first parameter Subsequently the cells are incubated with the MultiSort Release Reagent which enzymatically removes the MicroBeads from the antibodies In the next step these cells are magnetically labeled with MicroBeads conjugated with an antibody directed against a second marker After labeling the cells are again magnetically separated Magnetic labeling strategies There are two basic approaches to magnetic labeling direct labeling with MACS MicroBeads and indirect labeling with MACS MicroBeads against primary antibodies other specific ligands or their conjugates Both approaches can also be used for a MACS MultiSort strategy Direct magnetic labeling Direct magnetic labeling is the fastest way of magnetic labeling It requires only one labeling step as the specific antibody is directly coupled to the magnetic particle Direct labeling minimizes the number of washing steps and therefore avoids cell loss Highly specific monoclonal antibodies have been selected by Miltenyi Biotec to produce a large variety of antibody conjugated MicroBeads targeting many human mouse rat and non human primate cell surface markers The high specificity of the antib
201. t N utilisez pas l appareil dans un endroit humide ou mouill vitez l humidit et la condensation et prot gez la machine des clabous sures 1 4 4 Produit reli la terre L appareil est quip d une fiche lectrique trifilaire avec mise la terre dont la troisi me borne est pr vue pour relier l appareil la terre Cette fiche fonctionne uniquement dans une prise de courant avec contact de mise la terre Il s agit d un dispositif de s curit N essayez pas d ins rer la fiche dans une prise de courant sans contact de mise la terre Si vous ne pouvez pas ins rer la fiche dans la prise de courant veuillez vous adresser votre lectricien local qui remplacera la prise de courant 1 4 5 Sources lectriques Ne faites fonctionner l appareil qu partir d une source lectrique indiqu e sur la plaque signal tique du produit Si vous avez des questions sur le type de courant lectrique que vous pouvez utiliser contactez votre prestataire de services Miltenyi Biotec agr ou votre fournisseur d lectricit local N utilisez pas de rallonges ni de barrettes de connexion Ne surchargez pas la prise lectrique La charge totale du syst me ne doit pas d passer 80 de la valeur nominale du circuit de d rivation 1 4 6 Accessibilit Assurez vous que l interrupteur principal et le connecteur du cable lectrique soient facilement accessibles et plac s aussi pr s que possible de l op rateur de l appareil
202. t Rack set 130 093 532 autoMACS Pro 130 093 292 MACS Fluid Container Protection Cover 1 5 L 130 093 303 autoMACS ProSensor 130 094 574 MACS Reagent Rack 4 Cable 130 093 669 Cover MiniSampler Table 7 2 List of available accessories 166 7 Maintenance Protocols 8 The following chapter describes protocols that may be used with the autoMACS Pro Separator using manual labeling Please visit www miltenyibiotec com for an up to date list of manual labeling and autolabeling protocols using the autoMACS Pro Separator Note Detailed protocols and product documentation are provided on the data sheet for the particular MACS Cell Separation Reagents Work fast keep cells cold and use pre chilled solutions This will prevent capping of antibodies on the cell 8 1 surface and non specific bi cell labeling Volumes for magnetic labeling procedures described below are for up to 10 total cells When working with higher cell numbers scale up all reagent volumes and total volumes accordingly e g for 2x10 total cells use twice the volume of all indicated reagents and double the total volume When working with fewer than 10 cells do NOT scale down the volumes Keeping the samples on ice may result in poor cell o labeling Temperatures 6 Optional Add staining antibodies e g 10 uL of MACS Fluoro higher than 8 C and or chrome conjugated Antibodies and incubate according to the data sheet
203. t information 1 2 Advertencias y precauciones 1 El separador autoMACS Pro emplea tecnologia puntera Se trata de un dispositivo controlado por ordenador desarrollado para la separaci n de c lulas magn ticamente rotuladas con la tecnologia MACS El MACS MiniSampler conecta el separador autoMACS Pro y forma de esta manera parte del dispositivo de separaci n de c lula El separador autoMACS Pro y el MiniSampler estan dise ados para funcionar con seguridad una vez instalados y siempre que sean manejados por personal cualificado de conformidad con las normas generales de seguridad y las instrucciones contenidas en este manual del usuario Las directrices del presente capitulo explican los potenciales riesgos asociados al manejo del instrumento y suminis tran informaci n importante a fin de reducir dichos riesgos al minimo Si sigue cuidadosamente las instrucciones se proteger a si mismo y al equipo de posibles peligros y crear un ambiente de trabajo seguro Si este instrumento es manejado de un modo no previsto por su fabricante la seguridad se ver mermada IMPORTANTE Por favor lea y siga todas las instrucciones de uso recogidas en el presente manual de usuario y preste atenci n a todas las advertencias que aparecen en el instrumento Guarde para futuras consultas este manual y todas las dem s instrucciones de seguridad o de funcionamiento recibidas junto con el instrumento en un lugar accesible para todos sus usuarios IMPORTA
204. tato di quattro laser a emissione superficiale a cavit verticale VSCL per il rilevamento automatizzato classe 1M La radiazione non visibile Non osservare direttamente il laser con strumenti ottici p es lenti lenti di ingrandimento e microscopi Osservando la porta VSCL a una distanza inferiore a 100 mm con uno strumento ottico si potrebbero danneggiare i propri occhi L apparecchio dotato anche di un lettore di codici 2D dotato di un laser a semiconduttore visibile come puntatore per regolare la posizione di lettura e di potenti diodi luminosi LED per illuminare la zona di lettura Non guardare direttamente le radiazioni di diodi laser o LED o le radiazioni di diodi laser o LED riflesse da una superficie a specchio In caso contrario si potrebbero danneggiare i propri occhi Non rivolgere intenzionalmente il raggio laser verso altre persone Non smontare modificare o rimuovere le sorgenti di radiazioni laser o LED installate o le loro staffe di montaggio Le sorgenti di radiazioni laser o LED non cessano automaticamente di emettere radiazioni una volta smontate Le radiazioni di unit smontate possono essere causa di lesioni agli occhi Fare attenzione al percorso del raggio laser o a riflessi da una superficie a specchio Durante l installazione del separatore autoMACS Pro accertarsi che il percorso del raggio laser non si trovi alla stessa altezza degli occhi di chi esegue il lavoro Evitare che acqua grasso polve
205. te prematuro y reducir el tiempo de vida del instrumento pudiendo anular su garantia Mantenga este manual de usuario en un lugar seguro al alcance de todo aquel que este usando la Unidad de Separaci n autoMACS Pro separador autoMACS Pro El presente capitulo describe las instrucciones de seguridad y los requisitos de emplazamiento de su separador autoMACS Pro Las siguientes advertencias y precauciones tienen como objetivo evitar da os personales o materiales 1 1 S mbolos y niveles de peligro Disposici n de advertencias de seguridad Ejemplo CAUTION Moving parts stay clear Hazard of puncturing or crushing fingers Operate only with attached needle guard Las advertencias de seguridad informan al usuario sobre riesgos potenciales en caso de que no se sigan las advertencias y precauciones resumidas a continuaci n El s mbolo que aparece a la izquierda especifica el riesgo de que se trate El nivel de peligro que encabeza la advertencia clasifica el peligro de que se trate tal y como se mencion m s arriba El nivel tipo y fuente del peligro as como las posibles consecuencias prohibiciones y remedios se indican tal y como aparece a continuaci n 1 Important information 63 64 CAUTION WARNING o 0 6 gt gt OG S mbolos y niveles de peligro El siguiente recuadro recoge un glosario ilustrado que describe los s mbolos usados en el presente manual de usuario y en el separador autoMACS Pro
206. ted Miltenyi Biotec products and services are for research use only and not for therapeutic or diagnostic use autoMACS Pro Separator User manual Manufacturer Miltenyi Biotec GmbH Friedrich Ebert Stra e 68 51429 Bergisch Gladbach Germany Technical Support Phone 49 2204 8306 0 Fax 49 2204 85197 macstec miltenyibiotec de www miltenyibiotec com Version 02 Content Chapter Content 1 15 15 15 16 17 17 17 18 19 19 20 21 21 21 21 21 22 22 22 22 22 23 23 24 24 24 27 27 27 28 29 29 30 30 1 2 1 3 1 3 1 1 3 2 1 3 3 1 3 4 1 4 1 4 1 1 4 2 1 4 3 1 4 4 1 4 5 1 4 6 1 4 7 1 5 1 5 1 1 5 2 1 5 3 1 5 4 1 5 5 1 5 6 1 5 7 1 2 1 3 1 3 1 1 3 2 Content Important information Symbols and hazard levels Setup of safety notices Symbols and hazard levels Warnings and precautions General precautions Hazard of electric shock and spread of fire Strong magnetic field Hazard of crushing and shearing Laser radiation Secure installation Mounting accessories Air circulation Water and moisture Grounded earthed product Power sources Accessibility Peripheral devices Secure operation maintenance transport and disposal Secure operation Servicing Cleaning Hazardous material Transport Instrument disposal Electromagnetic compatibility Informations importantes Symboles et niveaux de danger Mise en place des consignes de s curit Symbole
207. tes using the navigation 6 arrows M W 3 Select the template for deletion 4 Select Delete Template Cell separation at a glance 1 Assign MicroBead reagent vials to positions on the MACS Reagent Rack It is recommended to use the 2D code reader Alternatively the product order number may be manually programmed N os gt Figure 6 15 CD4 MicroBeads human vial was placed onto the MACS Reagent Rack position R1 2 Onthe Separation tab the recommended separation strategy and wash program are pre selected for the chosen autolabeling reagents 3 Program the cell separation protocol using the Separation tab menu For full details refer to section 4 4 6 Cell separation using the autoMACS Pro Separator 129 130 Note Refer to the respective product data sheet for more detailing on sample magnetic labeling volumes BR gt Figure 6 16 Performing MicroBead CD4 cell separation with autolabeling left and without autolabeling right The cell separation Possel and wash conditions Qrinse for both processes are identical Disabling autolabeling 7 influences the initial sample volume For manual labeling it is recommended to dilute cells to a volume of 500 uL 108 total cells refer to corresponding data sheet for further information 4 Select Run Manual labeling at a glance Cells are labeled with MACS MicroBeads according to the MACS Cell Separation Reagent data sheet For general immunomagne
208. test calibration Installation of the autoMACS Columns Priming the autoMACS Pro Separator Checking the fluid levels Test of barcode scanner An overview of the autoMACS Pro touchscreen user interface Working with the lower menu bar Chapter Chapter Content 5 6 95 96 97 98 99 99 99 106 106 108 109 109 113 113 113 114 114 114 115 117 118 118 118 119 120 120 122 130 132 133 135 136 4 2 4 3 4 3 1 4 3 2 4 3 3 4 3 4 4 4 4 5 4 6 4 7 4 7 1 4 7 2 5 1 5 1 1 5 1 2 5 2 5 2 1 5 2 2 6 1 6 2 6 2 1 6 2 2 6 3 6 3 1 6 3 2 6 3 3 6 3 4 6 4 6 5 6 6 The main menu screen Status menu Status of fluid containers Column status Tube rack status MACS MiniSampler status Separation menu Reagent menu Log list menu Option menu Special User settings Materials required Materials required for operation Solutions Hardware Materials required for maintenance Solutions Hardware Cell separation using the autoMACS Pro Separator Select a cell labeling strategy Prepare cell samples Prepare single cell suspensions Reagent volumes and labeling volumes must be adjusted to the total cell number Select a cell separation strategy and a suitable magnetic labeling strategy Choosing a cell separation strategy Choose a magnetic labeling strategy autoMACS Pro Cell Separation programs autoMACS Pro Separator wash program
209. than MACS Antibodies are used carefully titer staining reagents For tips and hints on titration refer to the FAQs under Customer Support at www miltenyibiotec com the column To avoid this the primary antibody should be 1 O titrated carefully For tips and hints on titration refer to the FAQs under Customer Support at www miltenyibiotec com 3 The isotype of the primary antibody is not recognized by the Anti Immunoglobulin MicroBeads Make sure to use appropriate MicroBeads 4 Check the expression of the antigen epitope with flow cytometry 5 Antibodies may have degraded Check the antibody for its function Storage of diluted reagents at 4 C or 20 C may lead to degradation 6 Cells were not labeled with MicroBeads because the MicroBeads were degraded Check the expiration date Check for sterility of the MicroBeads if the vial has been opened before Antibodies may have degraded Check the antibody for its function Storage of diluted reagents at 4 C or 20 C may lead to degradation Positive fraction shows poor purity 1 The concentration of the primary antibody was too high leading to non specific binding of unwanted cells The primary antibody should be titrated carefully For tips and hints on titration refer to the FAQs under Customer Support www miltenyibiotec com 2 The primary antibody shows non specific binding Add blocking reagents such as BSA or immunoglobulin before labeling the cells 3 The incuba
210. the affected tubing by loosening the tube connectors using the black wrenches 3 Replace the tubing with the correct part 4 Pull back the connector from the tubing so that the tubing can be easily inserted into the appropriate port 5 Insert the connector cautiously and precisely and fasten it by hand Make sure not to overwind the screw 7 Maintenance 161 162 Note Fuse specifications are given in section 11 2 and on the marking plate on the rear of the instrument close to the fuse holder Do not use other fuses than specified 6 Plug in and switch ON the instrument Run the program Rinse and check for leakages 7 Run the program Calibr_2 to calibrate the volume control of the instrument For details refer to section 12 2 8 If there are any leakages refer to to section 10 1 2 for advice on how to improve the tubing connection 7 6 Exchange of fuses If the instrument fails to start upon switching it on or if operation suddenly stops and the screen is dark an exchange of the fuses might be required CAUTION Replace fuses only with those of the same type and rating 1 Switch OFF the instrument 2 Unplug the main power cord from the power outlet as well as from the instrument The fuse holder is located below the main power connector on the rear panel of the instrument 3 Pull out the fuse holder from the housing and exchange fuses LK ce Figure 7 25 Removing the fuse holder 4 Push the f
211. the user is encouraged to try to correct the interference by one or more of the following measures Reorient or relocate the receiving antenna e Increase the separation between the equipment and receiver Connect the equipment into an outlet on a circuit different from that to which the receiver is con nected Consult the dealer or an experienced radio TV technician for help 1 Important information 25 26 1 Important information Informations importantes 1 lire avant utilisation Veuillez imp rativement lire toutes les informations fournies dans ce mode d emploi avant d utiliser l appareil Le non respect de ces consignes peut donner lieu une utilisation une manipulation et une maintenance inappropri e ou incorrecte de votre appareil ce qui pourrait mettre en danger les utilisateurs fournir des r sultats non attendus entra ner le dysfonction nement ou la d t rioration de l appareil son usure pr matur e et r duire la dur e de vie de l appareil ainsi qu annuler votre garantie Conservez ce mode d emploi dans un endroit s r et accessible aux utilisateurs de l unit de s paration autoMACS Pro s parateur autoMACS Pro Ce chapitre d crit les consignes de s curit et les exigences d installation s appliquant votre s parateur autoMACS Pro Les avertissements et les mesures de pr caution suivantes ont pour objectif de vous aider viter toute blessure corporelle ainsi que
212. tialization failed Valve may be blocked or worn and cannot be turned correctly or valve drive is damaged 27 28 Motor rotation detection failed Valve may be blocked or worn and cannot be turned correctly Otherwise valve or magnet drive malfunction Valve rotation hindered Valve may be blocked or worn and cannot be turned correctly 29 257 Needle arm movement has been hindered 263 Unable to read rack bar code correctly 10 Troubleshooting Possible remedies 1 0 Restart instrument Please call Technical Support if error is thrown again Switch off the instrument wait for 5 seconds and switch the instrument on again If error persists touch the button DETAILS Exchange displayed valve If error is displayed again call Technical Support Touch button DETAILS Exchange displayed valve if applicable If error is displayed again or magnet is displayed call Technical Support Switch off the instrument for 5 seconds and turn it on again If error persists switch off instrument loosen air filters on buffer waste bottles by a half turn exchange columns for dummy columns verify that Washing Station is free of clogs and salt deposits eventually clean Washing Station verify that uptake needle is not clogged Switch on instrument if error persists touch DETAILS Exchange displayed valve if applicable Make sure to prepare samples as specified If error is displayed again call Technical Support
213. tic cell separation protocols please refer to section 8 1 The specific MACS Cell Separation Reagent data sheets can be found at www miltenyibiotec com 6 3 3 autoMACS Pro Cell Separation programs The autoMACS Pro Cell Separation programs are based on different cell separation strategies There are ten pre set separation programs to choose from depending on the cell isolation strategy the frequency of target cells and the level of antigen expression Positive selection programs POSSEL Positive selection in standard mode isolation of cells with normal antigen expression and frequencies higher than 5 select Possel if purity is the highest priority POSSEL_S Positive selection in sensitive mode isolation of cells with low antigen expression and frequencies higher than 5 select Possel_s if yield is the highest priority POSSELD Positive selection in standard mode I double column program for isolation of rare cells in low elution volume POSSELDS Positive selection in sensitive mode double column program for isolation of rare cells with low antigen expression POSSELD2 Positive selection in standard mode Il double column program for isolation of rare cells if purity is the highest priority POSSELWB Special positive selection in special mode double column program for isolation of cell subsets from whole blood Cell samples are automatically diluted with Running Buffer 6 Cell separation using the autoMACS Pro
214. tilisez pas l appareil dans des zones class es dangereuses telles que des environnements teneur lev e en oxyg ne Pour de plus amples d informations concernant l installation de l quipement veuillez vous adresser votre organisme local respons able de l approvisionnement en lectricit des travaux de b timent de la maintenance et de la s curit Si vous avez un doute s rieux quant la s curit d utilisation de votre appareil veuillez contacter votre prestataire de services Miltenyi Biotec agr ou appelez le service apr s vente de Miltenyi Biotec 1 3 Pr cautions d ordre g n ral Afin de r duire les risques potentiels li s l utilisation du s parateur autoMACS Pro Separator veuillez observer les pr cautions g n rales 1 Important information 1 29 suivantes La non observation de ces pr cautions peut provoquer un incendie des dommages corporels et ou d t riorer l appareil 1 3 1 Risque d lectrocution et de propagation de feu WARNING Hazardous voltages Risk of loss of live severe personal injury Unplug before cleaning Do not use if device is opened or damaged if liquids have been spilled into device if objects device through ventilation slots AVERTISSEMENT Les appareils lectriques pr sentent un risque d lectrocution Afin de r duire le risque d lectrocution n ouvrez aucun cache du s parateur autoMACS Pro Separator ni d autres quipements accessoir
215. ting Possible remedies 1 0 Please close protective cover and touch CONTINUE Touch CANCEL to abort Check electrical connection of MACS MiniSampler If detected the MACS MiniSampler symbol would be displayed in the status screen Check bar code on rack or try different rack Make sure ambient or direct sun light does not hit sensor Touch RETRY to try again touch SELECT to set chill rack type without automatic bare code reading Always use SLEEP to shut down the instrument Do not use sample volumes exceeding the maximum whole blood sample volume specified for the used chill rack type Split sample into several tubes reprogram separation param eters and restart separation If volumes are in specified range but error is displayed anyway please call Technical Support Check buffer supply Check for leakage of system unintended air intake Start a rinse program WASH ONLY and then retry calibration Otherwise call Technical Support Set time and date to actual values Make sure foam ontop of sample is not higher than 5 mm above liquid level Check for leakage at transition of needle to tubing and tubing to bubble sensor Touch CONT to process currently uptaken sample volume Touch CANCEL to abort Make sure foam on top of sample is not higher than 5 mm above liquid level Check for leakage at transition of needle to tubing and tubing to bubble sensor 191 192 Error code 7036 7037 7038 7039
216. tion strategies are classified as e positive selection or depletion e direct or indirect magnetic labeling cell sorting based on one or more marker s 4 Determine number of samples Select an appropriate tube rack for the number of samples and also take the sample volume into account 5 Prime the autoMACS Pro Separator Before performing a cell separation the autoMACS Columns must be filled with Running Buffer by running the Rinse program 6 Define autoMACS Pro Separation Programs on the sample rack template Select a separation and a washing program for every cell sample or labeling separation wash program Separation programs Positive selection or depletion Standard mode or sensitive mode Single or double column selection 6 Cell separation using the autoMACS Pro Separator 117 118 Note Use Pre Separation Filters 130 041 407 when cell clumping is suspected especially when working with previously frozen material cord blood dissociated tissue or whole blood Cell pre filtration removes aggregates dead cells and debris Special programs for efficient depletion of unwanted cells or cells that are only dimly labeled Special programs for cell separation directly from whole blood Special programs for the sequential depletion of up to 14x108 cells Washing programs e Quick Rinse Qrinse or Rinse are washing steps between separations Sleep to follow the last program before the instru
217. tion 3 2 7 for more information Check_up Performing a system check up The Check_up program allows the user to perform a system check up It is recommended to use the program if hardware errors occur The program starts after highlighting check_up and pressing Run 4 An overview of the autoMACS Pro touchscreen user interface 109 110 Note In case of malfunction during cell separation this feature must be disabled in order to perform a cell rescue procedure O_bcr 2D code reader setup configuration and initialization The O_ber program allows the user to setup configure and initialize a recently installed autoMACS Pro Separator 2D code reader O_init Optional priming of the instrument at startup By default the autoMACS Pro Separator does not perform a wash program after initialization The option O_init allows the user to add an initial rinse program that will be performed automatically after each initialization to prime the instrument O_led Activating deactivating bottle illumination The autoMACS Pro Separator has a bottle illumination designed to facilitate monitoring the instrument s status The bottle illumination can be switched ON or OFF O_progs Enabling disabling special separation protocols The O_progs are used to enable or disable special separation programs To enable or disable these protocols perform the following steps 1 Select Option User settings and O_progs 2 Select Run A dialog box wi
218. tion time during labeling was too long Do not exceed the incubation times specified in the respective data sheet 4 Check the primary antibody for its specificity When antiserum is used it is recommended to pre adsorb the antiserum e g on cells which do not express the antigen or to purify it by affinity chromatography ammonium sulfate precipitation ion chromatography etc in order to avoid non specific cross reaction 10 4 Fluorescent staining 10 4 1 Cells are poorly stained 1 Fluorochrome conjugated antibodies may have deteriorated Store MACS Fluorochrome conjugated Antibodies at 2 8 C 2 Fluorochrome conjugated antibodies and MACS MicroBeads might compete for epitopes Check the efficiency of the fluorescent staining on cell samples that are not magnetically 10 Troubleshooting 185 Note To prevent contamination runa Sleep program before turning off the instrument 186 labeled Especially for weakly expressed antigens it might be necessary to use a fluorochrome conjugated antibody that is directed against an epitope different from that recognized by the MicroBeads 3 For indirect staining of biotinylated antibodies with streptavidin fluorochromes please note that buffers supplemented with fetal bovine serum FBS or bovine serum albumin BSA may contain free biotin which can inhibit streptavidin binding to the biotinylated antibody Alternatively use MACS Anti Biotin VioBlue FITC PE or APC MACS
219. to deplete certain cell types from the cell sample in order to isolate the target cells The depletion strategy allows for example to isolate a target cell for which no specific antibody is available or to isolate untouched cells With a depletion strategy all unwanted cells are magnetically labeled During magnetic separation the labeled cells are retained on the column while the target cells pass through and form the negative non labeled fraction 6 Cell separation using the autoMACS Pro Separator A depletion strategy is recommended 6 forthe removal of unwanted non target cells e if no specific antibody is available for the target cells if binding of minimal amounts of antibody to surface molecules can interfere with downstream applications e if isolated cells are to be sorted subsequently utilizing a second marker Multiparameter sorting depletion followed by positive selection Cells can also be isolated by first depleting the non target cells followed by magnetic labeling and positive selection of the target cells This strategy is useful if unwanted cells in the cell suspension express the same antigen that was chosen as the marker for positive selection Ss It can also be used to deplete non target cells from the initial cell suspension before isolating extremely rare cell types by positive selection MACS MultiSort strategy With MACS MultiSort Kits high numbers of cells characterized by multiple cell
220. to section 5 3 5 8 Protocols 167 168 Note Primary antibodies should be titrated to determine the optimal staining dilution After incubation with primary antibodies wash the cells carefully If unbound primary antibodies have not been completely removed they may inhibit labeling of cells with indirect MACS MicroBeads Note When using Anti FITC MicroBeads or Streptavidin MicroBeads resuspend cell pellet in 90 uL of buffer per 10 total cells and add 10 uL of MicroBeads per 10 total cells Note Keeping the samples on ice may result in poor cell labeling Temperatures higher than 8 C and or longer incubation times may lead to non specific cell labeling 8 2 Indirect magnetic labeling Indirect magnetic labeling is the method of choice if direct MACS Cell Separation products are not available Almost any monoclonal or polyclonal primary antibody can be used A variety of Anti Fluoro chrome MicroBeads is available for magnetic labeling of cells stained with primary antibodies conjugated to VioBlue fluorescein isothiocyanate FITC R phycoerythrin PE allophycocyanin APC peridinin chlorophyll protein PerCP or tandem molecules like PE Cy5 or APC Cy7 Anti Biotin MicroBeads and Streptavidin Micro Beads are recommended when using biotinylated primary antibod ies Streptavidin MicroBeads must be used with biotin free buffer for labeling This is not necessary when using Anti Biotin MicroBeads
221. toMACS Pro Separator as described in section 7 2 1 13 Run the program Calibr_2 to calibrate the fluidic volume control of the instrument Refer to section 12 2 for details 7 3 3 Clean the washing station The washing station is designed for the automated rinsing of the outlet and uptake ports as well as surface cleaning of the uptake port needle The washing station should be cleaned as necessary to remove spills and salt crusts 1 Switch OFF and unplug the instrument ss 2 Make sure that the needle arm is in the uppermost position 3 Swivel the front cover to the right side swivel the cover of the washing station to the left side The cover can be removed by lifting it Figure 7 9 Removing the washing station 4 Press the tubing clamp on the right hand side of the washing station and pull to remove the tubing Note Potentially F contaminated liquid may spill out of the orifice of the washing station and the tubing Therefore wear protective gloves protective clothing and safety glasses to avoid contact with skin and eyes Figure 7 10 Remove the peristaltic pump tube from the washing station 7 Maintenance 151 Note Do not autoclave the washing station or wash using a dish washer Note Run the Store program and exchange the autoMACS Columns with substitute columns if you intend to store the instrument for a period longer than two weeks refer to section 6 8 2 Note If sample tubes and fr
222. tor Qrinse is the standard short wash program that only uses Running 6 Buffer It is recommended to use this program between separations of cells with normal frequency Rinse is an extensive rinsing program that uses Washing Solution Note The autoMACS Pro and Running Buffer It is recommended to use this program between Separator will not start separations of rare cells e g stem cells the separation of cells from a separation program different species and is mandatory between whole blood before a wash program separations has been completed Sleep it is mandatory to use Sleep as the last wash program before overnight storage Upon completion of the Sleep program the fluidic system contains 70 ethanol Maintenance programs The program Safe is designed for decontamination of the fluidic system For details refer to section 7 Upon completion of the Safe program the fluidic system contains Running Buffer The program includes an exchange of autoMACS columns The program Store should be applied to prepare the instrument for long term storage Upon completion of the Store program the fluidic system contains 70 ethanol For details refer to section 7 The program Col_ex is used for column exchange Upon completion of the Col_ex program the fluidic system contains Running Buffer 6 4 Select appropriate tube rack Three different tube racks are available for processing sample volumes between 0 2 mL and 50 mL Unless otherwis
223. torizzato o la societ elettrica locale Non utilizzare prolunghe o ciabatte multi presa Non sovraccaricare la presa di corrente Il carico complessivo del sistema non deve superare l 80 della potenza del circuito 1 4 6 Accessibilit Accertarsi che l interruttore principale e l attacco del cavo dell alimentazione siano facilmente accessibili e posti quanto pi vicino possibile all operatore dello strumento Qualora si renda necessario interrompere l alimentazione staccare il cavo dalla presa di corrente 1 4 7 Periferiche Soltanto le periferiche conformi a UL 60950 possono essere collegate al connettore RS232 contrassegnato con COM Il connettore 1 Important information contrassegnato come RS232 AUX non in uso E inoltre possibile 1 collegare soltanto apparecchi autoMACS Pro originali ai connettori contrassegnati con External CAN CAN1 e CAN2 I livelli di tensione su questi connettori non devono superare i livelli di tensione ammessi pari a 30 V rms e 42 4 V di picco o 60 V cc Al connettore del sensore i flacone pu essere esclusivamente collegato il cavo del sensore del flacone autoMACS Pro Al connettore RS232 BCR pu essere collegato soltanto un lettore per codice 2D raccomandato da Miltenyi Biotec dispositivi laser esterni collegati al connettore contrassegnato come RS232 BCR devono essere conformi alla norma IEC 60825 1 Utilizzare unicamente cavi di connessione infer
224. uffer and proceed to section 5 3 5 7 Fluorescent labeling should be performed after the separation process 8 Protocols 169 170 8 Protocols Quality control of separations 9 performed with the autoMACS Pro Separator Note To count the cells of the original fraction collect an aliquot of the cell sample after magnetic labeling directly before the magnetic separation to analyze whether cell losses are due to centrifugation steps OR to magnetic separation Also take counting statistics into consideration The standard deviation when counting cells is N N1 2 Therefore cell counting might be associated with large statistical errors To evaluate any MACS Cell Separation the separated cells can be analyzed with regard to purity recovery and viability Using MACS MicroBeads the magnetically labeled cells can be simultaneously stained with fluorochrome conjugated antibodies Antibodies of the same specificity can be used in most cases MACS Fluorochrome conjugated Antibodies are standardized to evaluate MACS Cell Separations The stained cells can subsequently be analyzed by flow cytometry fluorescence microscopy or other techniques 9 1 Recovery of cells In most cases the number of isolated cells will be compared to the number of cells theoretically expected from the heterogeneous starting population To calculate the target cell recovery take an aliquot from the magnetically labeled fraction just before sta
225. ure that a Reagent Rack 4 is placed on the MACS MiniSampler Click Done to continue test calibration 4 Test calibration Control position of needle in the washing station The instrument will automatically position the needle arm in the washing station If the needle is not correctly positioned click Move Up or Move Down to adjust the needle position accordingly Click Done to continue the test calibration sequence Figure 3 16 Test calibration of the needle arm position in the washing station 5 Test calibration Control position of needle in tube The instrument will insert the needle arm into tubes located in rack positions A1 B1 C1 Click Move up or Move down to adjust the position of the needle in the tube The needle arm should almost touch the bottom of the middle point of a standard 5 mL plastic sample tube If the needle is correctly positioned click Done 3 Assembly and installation Note After test completion the instrument will proceed to the next test calibration step control position of needle in vial Verify sangis tube positon O 107 Figure 3 17 Click Done if the needle is correctly positioned click Move up and Move down to adjust the position 6 Test calibration Control position of needle in vial The instrument will insert the needle arm into the MACS Reagent Rack vial positions R1 R2 R3 and R4 Click Move up or Move down to adjust the position of the needle in the vial The
226. use holder back into the housing and reconnect the main power cord 7 7 Rescue procedure Should the separation be interrupted before target cells are eluted it is possible to perform a cell rescue procedure to recover the sample If the instrument can be restarted follow procedure A if the instrument cannot be restarted follow procedure B 7 7 1 Rescue procedure A 1 Restart the instrument by switching it OFF and ON again 7 Maintenance 2 Undo the tubing connector at the negative port and place into a 50 mL tube 3 Take out the uptake port needle from the needle holder and place it into a 50 mL tube 4 Undo the tubing connector of the waste tube at the waste bottle and place it into a 50 mL tube place a second 50mL tube beside this one 5 Run the program Qrinse This will rinse the complete fluidic system with autoMACS Pro Running Buffer eluting the cells into the 50 mL tubes Depending on which step of the separation program that the interruption occurred the cells will be found in any one of the vials 6 Combine all fractions and centrifuge at 350xg for 10 minutes Ss Note Depending on the nature of your sample it is recommended to wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Dispose tissues swabs and vials appropriately 7 Discard the supernatant and apply cells to a reseparation as soon as possible Keep cells on ice until the separation 8 Reconn
227. ve selection of abundant or rare cells to the isolation of untouched cells by depletion of non target cells 12 preset separation programs simplify and standardize the application IMPORTANT The autoMACS Pro Separator is intended for research applications only and not for diagnostic or therapeutic use The autoMACS Pro Separator features automated sample labeling autolabeling sample loading elution of the negative non labeled as well as the positive labeled cell fractions Up to six samples can be processed in one programming step Furthermore automated procedures for maintenance of the system are included Different wash programs are available to rinse the columns before a new separation is performed One pair of columns can be used for up to 2 Introduction 100 cell separations or for two weeks whichever comes first The thin film transistor TFT color touchscreen with intuitive screen menus makes operation and monitoring of the instrument intuitive and easy Finally standard MACS Fluid Containers which are directly attached to the instrument and ready to use sterile MACS Buffers are available for maximum convenience The autoMACS Pro Separator is supplied with the MACS MiniSampler and therefore offers an additional sampling option This new feature allows the sequential processing of multiple samples without further manual handling The sampler is supplied with three different tube racks and an additional reagent rack The sam
228. vider nur Originalteile der Miltenyi Biotec GmbH oder Teile von Drittanbietern verwendet die von der Miltenyi Biotec GmbH spezifiziert und empfohlen werden Die Verwendung unautorisierter Ersatzteile kann Fehlfunktionen des Ger tes verursachen und die Ergebnisse von Zellseparationen beieintrachtigen Die Miltenyi Biotec 1 Important information 47 48 GmbH akzeptiert keinerlei Garantieanspr che und haftet nicht f r Fehlfunktionen oder Sch den am Ger t die auf Verwendung ungeeigneter Verschlei oder Ersatzteile zur ckzuf hren sind Nach jedweder erfolgter Wartungs oder Reparaturleistung lassen Sie Ihren autorisierten Miltenyi Biotec Service Provider alle notwendigen Sicherheitspr fungen durchf hren um sicherzustellen dass das Ger t sich in vorschriftsm igem Zustand befindet Nutzen Sie nur von Miltenyi Biotec empfohlenes Zusatzger t und von uns empfohlene Upgrades zu Ihrem Ger t 1 5 3 Reinigung Ziehen Sie den Netzstecker vor Reinigung des Ger tes Verwenden Sie keine Fl ssig oder Spr hreinigungsmittel sondern nur ein feuchtes Tuch 1 5 4 Gef hrliches Material WARNING Risk of loss of life if biohazardous material is used Wear protective gloves protective clothing and safety glasses to prevent contact with skin and eyes Operate device in a safety hood Decontaminate device after spilling of biohazardous material Wird oder wurde mit biologischen Gefahrenstoffen gearbeitet muss der Bediener des Ge
229. with the appropriate spare part Insert the tubing into the appropriate port Then cautiously insert and fasten the tube connector 5 Run the Rinse program and check if the leakage persists If so unscrew the tubing and check if the screw thread is damaged If this is the case order and install new tubing 6 Ifthe problem persists contact Technical Support 10 1 3 Pump syringe leakage Verify that the Running Buffer has equilibrated to room temperature before performing a washing or separation program Cold buffer will make the plunger seal constrict more than usual and may lead to leakage Salt crystals may also damage the pump syringe seal To clean the pump syringe refer to section 7 3 2 and retry If the problem persists follow the guidelines below 1 Runthe Sleep program Press the shutdown button on the upper right hand corner of the screen D Click Yes to shutdown the instrument 2 Switch OFF the instrument 3 Wash the pump syringe as described in section 7 3 2 4 Switch ON the instrument and run a Qrinse program to ensure that the problem is solved From the Separation menu select Wash Now lower menu bar Highlight Qrinse and click Run 5 If the leakage persists order and install either a new pump syringe 130 090 339 or anew pump seal 130 022 101 For details on the installation refer to section 7 3 2 10 Troubleshooting Note Depending on the level of use the pump syringe should be
230. x 1 530 888 8925 Phone 800 4020120 Freephone 800 FOR MACS Fax 0800 4020100 Freephone 1 800 367 6227 macs miltenyibiotec com China Miltenyi Biotec Shanghai Office Australia Fareast International Plaza A Rm 2301 Miltenyi Biotec Australia Pty Ltd No 319 Xianxia Rd Unit 16A 2 Eden Park Drive Shanghai 200051 P R China North Ryde NSW 2113 Phone 86 21 62351005 Australia Fax 86 21 62350953 Phone 61 02 8877 7400 macs miltenyibiotec com cn Fax 61 02 9889 5044 macs miltenyibiotec com au 13 Technical service 213 214 Singapore Miltenyi Biotec Asia Pacific Pte Ltd 7 Temasek Boulevard 16 06 Suntec Tower One Singapore 038987 Phone 65 6238 8183 Fax 65 6238 0302 macs miltenyibiotec com sg Japan Miltenyi Biotec K K Nittsu Etai Building 5F 16 10 Fuyuki Koto ku Tokyo 135 0041 Japan Phone 81 3 56 46 8910 Fax 81 3 5646 8911 macs miltenyibiotec jp France Miltenyi Biotec 10 rue Mercoeur 75011 Paris France Phone 33 1 56 98 16 16 Fax 33 1 56 98 16 17 macs miltenyibiotec fr 13 Technical service Italy Miltenyi Biotec S r l Via Persicetana 2 D 40012 Calderara di Reno Bologna Phone 39 051 64 60 411 Fax 39 051 64 60 499 macs miltenyibiotec it Spain Miltenyi Biotec S L C Luis Bu uel 2 Ciudad de la Imagen 28223 Pozuelo de Alarc n Madrid Phone 34 91 512 1290 Fax 34 91 512 1291 macs miltenyibiotec es United Kingdom Miltenyi Biotec Ltd Almac House Church Lane Bis
231. y There are two basic strategies for separating specific cell populations positive selection and depletion Using the positive selection strategy the target cells are magnetically labeled and collected as the positive fraction Using the depletion strategy the unwanted cells are labeled separated from the target cells and eluted as a positive cell fraction i e cells labeled with MicroBeads The target cells will be collected in the negative fraction Both strategies can be combined for multiparameter sorting Positive selection Positive selection allows up to 10 000 fold enrichment of the magnetically labeled target cells Positive selection takes advantage of the high specificity of monoclonal antibodies to isolate highly pure cell populations that specifically express the corresponding antigen The positively selected cells are virtually unaffected by the separation procedure and can be used immediately for culturing or other applications There is no need to remove the MACS MicroBeads from the cells The small size and the composition of the MACS MicroBeads iron oxide and polysaccharide make them biodegradable Typically MACS MicroBeads do not activate cells or influence function or viability Positive selection is recommended for highly specific labeling e for excellent purity and recovery e for the isolation of rare cells e ifno subsequent separation step is needed Depletion For some experiments it may be desirable
232. y adjust the central positioning of the uptake needle by using the needle navigation buttons Move left Move right Height The uptake needle must be positioned above the center of the tube in row A of the tube rack si 4 ve Figure 12 10 The uptake needle should be positioned as shown above Calibration 4 5 z as P 14 10 a 5 lt s Figure 12 11 Adjust MiniSampler position 25 Select Save to store new settings Calibration 1 Test current calibration settings step 5 5 26 Select Test to test new configurations The autoMACS Pro Separator will perform a complete test of Calibration 1 settings Calibration SS Test Tant camteston sata Figure 12 12 It is important to test the new calibration settings To navigate back through the calibration steps select Previous 12 Hardware and software installation 27 Place a MACS Cooling Tube Rack onto the MACS MiniSampler 1 2 28 Select Use to start the calibration at the current position recommended Select Reset to reset the position to factory settings 29 Place aMACS Reagent Rack 4 onto the MACS MiniSampler 30 Select Done 31 Check all approached positions by using the needle navigation buttons Move up and Move down then select Done 32 If any errors or misalignments were noted repeat the entire process Calibration 2 1 Select menu Option 2 Highlight User settings and select Calibr_2 3 Select Run a gt Figure 12 13 Performing Cali
233. y Miltenyi Biotec for immunomagnetic labeling and subsequent separation of cells or biomolecules in a high gradient magnetic field MACS MicroBeads Super paramagnetic particles conjugated to antibodies for magnetic labeling of cells or biomolecules Magnet cover Black cover surrounding the magnets The magnet cover is located in the center of the fluidic system and has slots for the autoMACS Columns Memory card Removable compact flash SD RAM card containing the autoMACS Pro Programs Memory card slot Slot located on the right hand side of the autoMACS Pro Touchscreen giving access to the memory card The memory card should be removed by trained personnel only Negative fraction Sample fraction containing the non labeled cells that pass through the autoMACS Column while the column is placed in the magnetic field PE Phycoerythrin PerP Peridinin chlorophill protein i Ports The automated arm carries two ports the port located proximal to the instrument is designed for computer controlled fluid detection and distribution such as magnetic labeling sample mixing sample uptake and release of the magnetically labeled fraction while the port in the front releases the non labeled fraction Positive fraction Sample fraction containing the cells labeled with MACS MicroBeads These cells are retained on the column while the column is placed in the magnetic field The cells are eluted from the column after the column has been remove
234. yringe must be cleaned every 1 3 months 1 Runa Sleep Program refer to section 7 2 3 switch OFF the power and unplug the autoMACS Pro Separator from the electric supply Open the front access cover The syringe pump plunger should be at the top right position 2 Loosen the plunger lock screw by turning it counter clockwise IL j Hy u dive Figure 7 5 Unscrewing the plunger lock screw 3 Lower the plunger holder by firmly pressing the plunger lock screw downward Figure 7 6 The plunger was gently lowered 4 Unscrew the syringe from the dilutor valve housing 7 Maintenance 149 L h a e Figure 7 7 To remove the pump syringe the top screw must be turned anti clockwise 5 Carefully remove the plunger from the syringe 6 Remove salt crusts with distilled or deionized water Figure 7 8 Washing the pump syringe with distilled water Note If symptoms of wear 7 Gently push the plunger back into the syringe and dry the such as leakage persist plunger lock screw before proceeding with installation of the contact Technical syringe Support 8 To reinstall the syringe carefully insert the syringe into the plunger holder 9 Push the plunger holder up until the syringe reaches the dilutor valve 150 7 Maintenance 10 Fasten the syringe at the dilutor valve by turning it until a resistance can be felt 11 Push the plunger holder up once again and tighten the plunger lock screw 12 Prime the au
235. zare la disposizione delle attrezzature tenere in considerazione le precauzioni descritte nella presente sezione in modo da evitare guasti allo strumento e ridurre la possibilit di spegnimenti causati dall ambiente IMPORTANTE vanno sempre osservate le istruzioni di sicurezza sul luogo di lavoro le procedure di laboratorio e le norme relative alla salute e alla sicurezza del laboratorio nonch alla prevenzione degli infortuni 1 4 1 Accessori di montaggio Non collocare il autoMACS Pro Separator su un piano un carrello un supporto un treppiede o una staffa poco stabile che potrebbe provocare la caduta dello strumento con la possibile conseguenza di lesioni personali gravi o di gravi danni allo strumento Utilizzare unicamente piani carrelli supporti treppiedi o staffe raccomandati da Miltenyi Biotec o venduti insieme allo strumento Non collocare il autoMACS Pro Separator su sistemi incassati o in spazi angusti come uno scaffale a meno che non siano stati concepiti specificamente 1 Important information 57 58 per accogliere lo strumento e a condizione che vi sia una ventilazione adeguata e che siano state seguite le istruzioni di montaggio dello strumento 1 4 2 Circolazione dell aria Lo strumento non deve essere collocato in prossimit di radiatori stufe o altri dispositivi inclusi gli amplificatori che producono calore Accertarsi che via sia un adeguata circolazione d aria attorno al autoMACS Pro Separator ma

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