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Fluoroskan Ascent FL - User Manual

1. Solvent Polysulfone Teflon KelF Polypropylene Acetaldehyde 0 0 0 Acetates 0 0 Acetic Acid 0 0 0 0 Acetic Anhydride 0 Acetone 3 0 0 0 Acetyl Bromide 0 Ammonia 0 0 0 Ammonium Acetate 0 Ammonium Hydroxide 0 0 0 Ammonium Phosphate 0 0 Ammonium Sulfate 0 0 Amy Acetate 0 3 Aniline 0 0 0 Benzene 3 0 3 Benzyl Alcohol 0 0 0 Boric Acid 0 0 0 Bromide 0 0 Butyl Alcohol 2 0 0 1 Butyl Acetate 3 0 E Carbon Sulfide 0 Carbon Tetrachloride 0 0 1 3 Chloroacetic Acid 0 0 Chlorine 0 1 3 Chlorobenzene 3 3 Chloroform 3 0 3 Chromic Acid 3 0 0 Cresol 0 dd Cyclohexane 0 0 3 Dimethyl Sulfoxide DMSO 0 0 0 0 Ethers 0 ER Ethyl Acetate 3 0 0 Ethyl Alcohol 0 0 0 Ethyl Chromide 0 1 3 Continued 48 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Cont Solvent Polysulfone Teflon Kel F Polypropylene o o o Formaldehyde 0 Formic Acid Freon Gasoline Glycerine Hydrochloric Acid Hydrochloric Acid conc NIOJo o D N 0 0 0 wm 0O o Hydrofluoric Acid Hydrogen Peroxide Hydrogen Peroxide conc Hydrogen Sulfide OoO lo 0o 0o 0O 0Olo o 0O NIo Kerosene Methyl Ethyl Ketone MEK Methyl Alcohol Methylene Chloride Naphtha I ojojwjojojojojojo
2. 0 1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 Diameter mm Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 41 4 5 2 Incubator The incubator contains two heating element plates in the measurement chamber one heating element plate under the microplate and another above it Both heating element plates are temperature controlled The upper plate is slightly warmer than the lower plate to avoid condensation on the plate lid The measurement chamber is large to adopt different plate formats and therefore extensive evaporation may cause some variations in the temperatures between the wells Consequently when using incubations with extended periods of time in the instrument a plate lid is recommended If the incubation period is long without any other important functions the Fluoroskan Ascent FL Fluoroskan Ascent automatically changes the place of the plate within the measurement chamber to minimize temperature differences between the wells 4 5 3 Dispensers The optional dispensers 1 to 3 dispensers numbered from left to right are located inside the instrument housing under the dispenser cover Fig 2 1 2 The dispensing heads have three alternative dispensing positions one of these dispenses into the well in the measurement posi
3. Recommended W Possible 1 Not allowed 1080 1020 gt 960 900 gt 840 gt 780 gt 720 4 660 600 gt 540 gt 480 420 360 gt 300 gt 240 180 4 120 60 5 0 1 3 5 7 9 11 131517 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 Diameter mm Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Table 45 10 Shaking speeds for 6 well plates 1200 1140 Recommended 1080 1020 E Possible Not al lowed 960 900 gt 840 gt 780 gt 720 4 660 gt 600 gt 540 gt 480 420 360 gt 300 gt 240 gt 180 4 120 4 60 gt Speed rpm 0 1 Table 4 5 1e 3 5 7 9 111315 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 Diameter mm Shaking speeds for 95 mm Petri dish 1200 1140 y 1080 1020 960 900 gt 840 gt 780 gt 720 4 660 gt 600 gt 540 gt 480 4 420 4 360 gt 300 gt 240 gt 180 120 4 60 4 Speed rpm Recommended E Possible Not al lowed
4. Th ermo Fluoroskan Ascent Fluoroskan Ascent FL amp Fluoroskan Ascent AA AAA A AA User Manual User Manual Rev 2 4 Nov 2008 Cat no 1506450 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Table of Contents Symbols and Markings n nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnvnner 7 1 Introduction m nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnunnnnnn 9 2 Instrument Layout nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 13 3 Installation ennnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnuennnn 19 4 Operation iniciada iii diia 31 5 Maintenance nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnunnnnnnnnnen 61 6 Troubleshooting Guide nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnr 71 7 Frequently asked questions FAQ about the Fluoroskan Ascent FL and Fluoroskan Ascent seseeeeeeeeeeeeeeeeeeeees 75 8 Instrument Service r rnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnen 81 9 Disposal of the Instrument and Materials annnnnnnnnnnnnnn 87 10 Ordering Information anannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnr 89 11 Glossary and Abbreviations nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 101 12 Literature nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnne 105 13 Warranty Certificate nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
5. mmmsvvrerenavvvrrrnavrrrrrrnnrresrsnsvvrsssnsrsessennnnee 87 9 2 Disposal of materials mrsssvvvvravavvvnnannvvvrnrnsvrenrsnsvrvenrersvrnnssssvveneen 87 10 Ordering Information a axannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnvnnnnr 89 10 1 Product code numbers ccoococcccccccnnnncannnononncnnnnnncn naar nn nc nn nn nn 89 10 2 List of recommended spare parts ocoococccononoccnnnnononnnnnonananoncnnnnn nn 96 10 3 Ordering filter ccoo ara 96 10 4 Fluorochromes and corresponding filter pairs seer 99 11 Glossary and Abbreviations nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 101 12 A AAA O A A 105 13 Warranty Certificate annnnnnnnnnnnnnnnnnnnnnnnnvnnnnnnnnnnnnnnnnnnnnn 107 13 1 Warranty limitations 00ooococcnnnnnnnoconccnnnnnnnnnnann ere 108 14 Specifications coiccocinocanicnadonncc ni 109 14 1 General specifications essere reen rn 109 14 2 Safety specificatiONS o ooccccnnnoccccnanonncnnanannnnnnnn nn nn ramon nn nnnnrnnnnnnos 111 14 3 In conformity with the requirements ooccconncccnccanonnncnaninnnnnnannnns 112 14 4 Performance specificatiONS conoocccccnnonnncnancnnnnnannnncrnnnnnncnnnnn on 113 15 Ind x A ea ee 115 Appendix A Brief User s Guide nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 119 Appendix B Filter Order Form nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 121 Appendix C Addresses x nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 123 6 Fluoroskan Ascent FL amp Fluoroskan
6. 6 Fit the optical unit light cover and the instrument cover back into their places 4 8 Installing the filters 1 Switch off the instrument and disconnect the mains power supply cable Fig 3 3 2 Remove the instrument covers as described in Section 3 2 Releasing the transportation lock points 1 4 3 If the optical unit is above the measurement chamber undo the four finger nuts Fig 4 6a 2 and remove the light cover Fig 4 6a 1 You can find more detailed instructions in Section 4 6 Changing the measurement direction 4 If the optical unit is below the measurement chamber undo the six screws Fig 4 6b 1 fixing the measurement chamber to the chassis Lift the measurement chamber into the upper position There are hinges in the rear and a gas spring Fig 4 6c 3 holds the chamber in the upper position 56 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual leave your fingers between the measurement chamber and Caution The measurement chamber is very heavy DO NOT the bottom case 5 Unplug the flat cable Fig 4 6b 3 and the lamp cable located beside the flat cable from the optical unit Undo the three long finger nuts Fig 4 6b 4 and remove the optical unit and place it on a table 6 The excitation filter wheel Fig 4 8a can be removed by undoing the fitting screw Fig 4 8a 1 DO NOT touch the filter surfaces 7 The emission filter wheel Fig 4 8a can be removed by undoing the fittin
7. Trademarks and Ascent Software Fluoroskan Fluoroskan Ascent Immulon Luminoskan Luminoskan Ascent Microfluor and Microtiter are registered trademarks of Thermo Fisher Scientific BioBind and Microlite are trademarks of Thermo Fisher Scientific alamarBlue is a trademark of AccuMed International Inc AlexaFluor PicoGreen and Texas Red are registered trademarks of Molecular Probes OliGreen and RiboGreen are trademarks of Molecular Probes AttoPhos is a trademark of JBL Scientific Cy is a trademark of Amersham Biosciences StarBright Green is a trademark of Chromagen Virkon is a registered trademark of Antec International Ltd All other trademarks registered trademarks and copyrights are the property of their respective holders Disclaimer Thermo Fisher Scientific reserves the right to change its products and services at any time to incorporate technological developments This manual is subject to change without prior notice Although this manual has been prepared with every precaution to ensure accuracy Thermo Fisher Scientific assumes no liability for any errors or omissions nor for any damages resulting from the application or use of this information This manual supersedes all previous editions No liability for consequential damages Thermo Fisher Scientific shall not be liable for any damages whatsoever arising out of the use or inability to use this product Contact information Thermo Fisher Scientific Oy
8. You can find the filter pair scaling entry in Ascent Software The menu selection is Setup Filters Select the corresponding filter pair and key in the scaling factor The following filter pairs if any of these are factory installed in the instrument are scaled as follows Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 35 Filter pair Scaling reference Scaled reading approx Ex 355 nm Em 460 nm 1000 pmol 4 Methylumbelliferone 1000 free acid 4 MeU Sigma M 1381 Ex 485 nm Em 518 nm 10 pmol Fluorescein Sodium salt 10 527 and 538 nm Sigma F 6377 Ex 544 nm Em 590 nm 625 pmol Tetramethylrhodamine 625 isothiocyanate TRITC Sigma T 5646 Ex 584 nm Em 612 nm 500 pmol Texas Red Sigma S 3388 500 Ex excitation filter Em emission filter Note The scaling factors used can be seen from the instrument status report 4 3 3 Dynamic series test solutions Dynamic series for the Fluoroskan Ascent FL Fluoroskan Ascent filter pair excitation filter 485 nm emission filter 538 nm Fluorescein Sigma F 6377 M 376 3 g mol store at amb temp Reagents 0 01 M PBS pH 9 0 dilution buffer K2HP04 3 H20 M 228 23 g mol 2 289 NaCl M 58 44 g mol 9 0g Distilled water ad 1000 ml Adjust the pH if necessary with a few drops of 0 1 M NaOH to pH 9 0 25 mmol l Fluorescein stock solution Fluorescein Sodium salt Sigma F 6377 M 376 3 g mol 94 mg Pure ethanol 94 3 ml 0 01 M PBS pH 9 0 ad 10 ml
9. 36 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Dilution series No pmol 100 yl Make the dilutions in 10 ml tubes as follows 13 5000 100 ul Fluorescein stock sol ad 50 ml 0 01 M PBS pH 9 0 12 1000 2 ml dilution 13 ad 10ml 8ml 11 100 1 ml dilution 12 ad 10ml 9 ml 10 50 3 mi dilution 11 ad 6ml 3 ml 9 10 1 midilution 11 ad 10ml 9 ml 8 5 3 mi dilution 9 ad 6ml 3 ml 7 1 1midilution9 ad 10mil 9 ml 6 0 5 3 ml dilution 7 ad 6ml 3 ml 5 0 1 1 mi dilution 7 ad 10ml 9 ml 4 0 05 3 ml dilution 5 ad 6ml 3 ml 3 0 025 2 ml dilution 5 ad 8ml 6 mi 2 0 01 1 ml dilution 5 ad 10ml 9 ml 1 0 005 3 ml dilution 2 ad 6ml 3 ml BLANK 0 01 MPBS pH 9 0 Pipetting Use a Black Microtiter Strip plate Cat no 95029450 Blank 100 ul well in eight replicates into column 1 Dilutions 100 ul well dilutions 1 9 in eight replicates into columns 2 10 100 ul well dilutions 10 13 in four replicates into columns 11 12 4 4 Luminometric measurement Only the Fluoroskan Ascent FL can measure luminometric labels Luminometric reading is optimized for 96 and 384 well plates and top reading To obtain the best sensitivity and the lowest crosstalk a light shield Fig 4 7a between the optics and the plate must be used The light shield does not interfere in fluorometric measurements In luminometric measurements the dispenser cover must be closed F
10. To change the measurement direction the optical unit and the cover plate are interchangeable Installing or removing the light shield The light shield is only present in the Fluoroskan Ascent FL 54 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Light shield Positioning guide Fig 4 7a Installing the light shield 1 Switch off the instrument and disconnect the mains power supply cable Fig 3 3 2 2 Remove the instrument cover as described in Section 3 2 Releasing the transportation lock 3 Undo the four finger nuts Fig 4 6a 2 and remove the light cover Fig 4 6a 1 4 Undo the three long finger nuts Fig 4 6b 4 and remove the optical unit or if the optical unit is below the measurement chamber remove the cover plate Fig 4 6c 1 5 When removing the light shield use your little finger to lift the light shield up When installing the light shield ensure that the positioning guide is placed towards the front of the instrument against the corresponding guide in the holder Fig 4 7a and Fig 4 7b If the light Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 55 shield is not positioned correctly the optical unit does not slot down into its correct place but stays swinging Caution Mispositioning of the light shield will cause discrepancies in the measurement results or may cause the instrument not to work Positioning guide Fig 4 7b Top view of the light shield
11. User Manual 4 3 Fluorometric measurement A measurement function has several phases 1 The instrument switches on the halogen lamp and waits for 5 seconds to balance the intensity if the lamp was off The lamp is switched off if the instrument is idle and has not received any commands from the computer in 3 minutes The anode voltage of the PMT is selected according to the excitation and emission wavelengths to compensate for the spectrum of the halogen lamp and the spectral response of the photomultiplier tube Selected filters are driven to the measurement positions by rotating the excitation and emission filter wheels The plate carrier is driven in and the reference chip corresponding to the selected filter pair is driven to the measurement position In the autocalibration procedure the instrument reads the fluorescence from the reference chip compares it to the value in memory and sets a factor to correct the reading The original chip value is in memory and due to the drift of the PMT and the drift of the lamp the measured chip value may differ slightly from the original one This difference is used to correct the reading in order to obtain the original signal levels Also see Section 4 3 1 Validation In long measurement procedures autocalibration is performed in a suitable phase without disturbing the measurement timing The time interval between autocalibrations is 5 minutes After autocalibration the wells
12. 61 81 83 84 85 87 BIE eo E E AE A A E SE si 11 16 110 DIMENSIONS incida a a re hadd 46 77 109 110 DITECUNOS iii a aa aa Aa 112 Disinfection haran a aae druer 59 61 81 82 83 87 101 POC Uri i 61 81 82 83 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 115 Dispenser Head tube naninuenadtunrnuan G te vridde dad caida idiota ci 46 68 SVINE viii laa knee caver dada 70 91 96 TUDING a EE aoe E E 16 43 65 119 Dispensing head seerne 15 17 25 42 43 44 65 66 68 69 73 A O 15 25 42 Dispensing tip ananannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 47 59 67 68 69 73 91 96 Dispensing tubing sanannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 67 68 91 96 Dritt eee 33 38 74 Drop platica dai 26 27 72 91 DUMMY HEr cnica aida EE Dd haa 58 DA O 15 42 E End weightia id a E A 26 73 91 Environmental requirement cccccccccscecscececscececececceceeceeeeeeeeeeeeeness 21 119 Error messadg esse aser ae ia sete a 31 74 101 F Feedback Form nit a vde kadett sevedevedber Stusas 81 Filter O ERE 62 Ordek cts ius Seri india bid ia dis 96 order O Mita tii idad 96 121 122 A E 11 16 33 38 57 58 74 77 110 JU TO 07 11 q EPA OOOO C NDOR 36 37 99 100 102 113 Fluorochrome rmuerrananvvnnnnenvvnnanensvnnenern 71 72 99 100 101 102 121 122 Fluorom eteruskn sadk ndrudehudnevdideg a a aa aaa 9 10 61 102 G ETE A E AAAA EAE E E EAT aa 31 33
13. In this case switch the instrument OFF and ON again If this failure is repeated contact authorized technical service The instrument is ready for operation The instrument also performs different kinds of additional runtime hardware tests before each measurement run Autocalibration in fluorometry and automatic blanking in luminometry are performed Because of the relative nature of fluorometry and luminometry we recommend you use known samples or controls to verify instrument operation 30 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 4 Operation The Fluoroskan Ascent FL Fluoroskan Ascent is fully computer controlled Ascent Software controls the reader functions and provides complete data handling and report formatting For further details on how to use the software refer to the Ascent Software User s Guide Warning DO NOT use any other software than Thermo Scientific software designed for this instrument Use only valid combinations of the PC software and the instrument s internal EEPROM Warning The Fluoroskan Ascent FL Fluoroskan Ascent instruments do not verify the logic flow of the received commands The instrument is equipped with a power switch ON OFF and a three color LED indicator When the instrument is switched ON the color indicates the state of the instrument Green The instrument is ready and waiting for a command Orange The instrument is busy executing a command Red The ins
14. Microplate Instrumentation Ratastie 2 P O Box 100 Fl 01621 Vantaa Finland Tel 358 9 329 100 Fax 358 9 3291 0415 www thermo com 8 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 1 Introduction The Thermo Scientific Fluoroskan Ascent designed and made by Thermo Fisher Scientific is a microplate fluorometer which offers versatility and flexibility for even the most demanding fluorometric applications The extended Thermo Scientific Fluoroskan Ascent FL is equipped with both fluorometric and luminometric detection technologies As a combination instrument it covers the full range of fluorometric as well as glow and flash luminometric applications Fig 1 Fluoroskan Ascent FL Fluoroskan Ascent Thermo Scientific Ascent Software controls all the reader functions and provides the features for data handling and report formatting The software for the Fluoroskan Ascent is a dedicated software for fluorometric applications while the software for the Fluoroskan Ascent FL enables both fluorometric and luminometric detection methods to be used even during a single session The advanced optical system based on direct illumination optics produces a sharply focused light beam for fluorometric measurements prevents crosstalk and ensures accurate readings The choice of two beam diameter settings allows optimal reading of 1 to 96 and 384 wells High sensitivity is one of the main benefits of fiberless optics and a critical fea
15. defines the level of transient overvoltage which the instrument is designed to withstand safely It depends on the nature of the electricity supply and its overvoltage protection means For example in CAT Il which is the category used for instruments in installations supplied from a supply comparable to public mains such as hospital and research laboratories and most industrial laboratories the expected transient overvoltage is 2500 V for a 230 V supply and 1500 V for a 120 V supply 2 The pollution degree describes the amount of conductive pollution present in the operating environment Pollution degree 2 assumes that normally only nonconductive pollution such as dust occurs with the exception of occasional conductivity caused by condensation Both of these affect the size of the electrical dimensioning within the instrument Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 111 14 3 In conformity with the requirements Fluoroskan Ascent FL Fluoroskan Ascent bears the following markings Type 374 100 240 Vac 50 60 Hz 200 VA CE mark CSA monogram with US designator Fluoroskan Ascent FL Fluoroskan Ascent conforms to the following requirements 2006 95 EC Low Voltage Directive 2004 108 EC Electromagnetic Compatibility Directive EMC FCC Part 15 Subpart B Class B July 2004 2002 96 EC Waste of Electrical and Electronic Equipment Safety performance EN 61010 1 2001 EMC performance EN 61000 6 3
16. tubing syringe seals and valves If they become worn out the symptoms are Poor precision and accuracy A variable or moving air gap Leakage Drops and spills 66 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual The frequency of replacement will depend on the duty cycle fluids used and instrument maintenance If any of these symptoms occur and it is not obvious which component is causing the problem the easiest and most economical way is to replace one component at a time in the following order 1 dispensing or aspirate tubings and or dispensing tip and 2 syringe 5 6 1 Replacing the dispenser tubings 1 To remove either the dispensing tube or the aspirate tube assembly from the valve gently loosen the fittings either manually or using a 7 9 mm 5 16 in wrench Unscrew the fittings and remove the tubing 2 We recommend you replace the complete assemblies always when replacement is necessary Alternatively only some parts of the assemblies can also be replaced If only the dispensing tubing is changed first remove the complete dispensing tube assembly from the dispenser unit 2 3 1 Dispensing head tube Dispensing tip Silicone tubing Internal dispenser head tube Brass tube holder o a fF O N Brass locking ring 7 Dispensing tubing Fig 5 6 1 Structure of the complete dispensing tube assembly Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 67 10 11 68 Remove the disp
17. 34 113 H Half bandwidth eevvnverrnnnnnnnnnnvernnvnnnnanenenvnvrnnnnnevener 75 89 90 97 102 110 I INGUbat ruasnshulmianegrhimibelmsdtibiesilrnrdin 10 17 42 110 114 INitialiZatiON aria ii tina adan bad 30 31 32 102 103 Instrument layou re ta 13 Integration MS mamita ae ease cote deepen 33 Intended USE mana aa i a aa ieai E aaa i 10 116 Fluoroskan Ascent FL Fluoroskan Ascent User Manual LiMP ia Sieg Ohi tenes 11 16 33 34 54 57 64 91 96 109 Leakage tray caina ponin ed must 15 91 LED sandaler karen Ende 30 31 32 102 LOCKING PICO iii nina aa dad edda 22 23 Lockingring dd eek 68 M Maintenance orerar paiard akairo r aeeie hiai 61 65 66 67 81 85 N E SEE 21 33 73 74 O Operational check srrrrnanavvvnnnanavvvnnansvvvnssnsnnnnssnsnnnnssnsnnnnsssennvnnssssnnrnessennnne 30 Ordering infOrmatiOn cccccccccsssssssscseceeecsceesesseaeceeeeessssceseaeeeseesessssseaeess 89 P Packin iia at ai 19 20 81 85 87 ASTUCIA ii a a a ea al 19 85 A OT 19 20 Material iia a rial 19 87 PER ein titi adn 28 78 79 87 91 Photomultiplier tube 11 12 16 33 34 38 75 90 102 110 Plate adaptei asra annaa araea ane Era aaea Ea APATA ERRAT 27 28 29 73 74 79 PUNGE E aa a dada 25 26 70 73 91 96 POSITIONING lever a iaraa AR ad la 18 32 Prime TIP a A E E A ERE RER ES AN LEES 44 R Reference Chip s scccccsecsssssssseeeeeseeesees 18 33 34 63 74 76 102 103 Replace siii it
18. and amplifies the current Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Reference chip RFU or rfu RH RLU or rlu rpm Scaling Self tests Stokes shift A filter pair is validated against a reference chip in Ascent Software If a filter pair does not have a suitable reference chip the asterisk will appear after the filter wavelengths given Relative Fluorescence Fluorometric Units The arbitrary units in which fluorescence intensity is reported Relative humidity Relative Luminescence Luminometric Light Units The arbitrary units in which luminescence intensity is reported Revolutions per minute The measured values are expressed as RFU or RLU Scaling is a way to convert readings to show desired values In fluorometry the normal beam and the small beam have separate scaling factors A user defined scaling factor can be set to the indicated filter pair in Ascent Software see Procedure Desktop Setup Filters The measured values are then multiplied by this factor Initialization tests and adjustments that the instrument performs prior to operation as well as autocalibration The difference between the wavelengths of the excitation and emission peaks Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 103 104 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 12 Literature Practical Fluorescence 1990 Guilbault G G ed Second edition Marcel Dekke
19. and the accessories for any possible transport damage If the carton has been damaged in transit it is particularly important that you retain it for inspection by the carrier in case there has also been damage to the instrument 20 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Visually check all interconnections in the basic instrument Check that there are no loose parts inside the instrument If any parts are damaged contact your local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy 3 1 4 Environmental requirements and noise When you set up your Fluoroskan Ascent FL or Fluoroskan Ascent avoid sites of operation with excess dust vibrations strong magnetic fields direct sunlight draft excessive moisture or large temperature fluctuations e Make sure the working area is flat dry clean and vibration proof and leave additional room for cables connections controlling computer printer etc e Make sure the ambient air is clean and free of corrosive vapors smoke and dust e Make sure the ambient temperature range is between 10 C 50 F and 40 C 104 F e Make sure relative humidity is between 10 and 90 non condensing Leave sufficient space at least 10 cm at both sides of the instrument and at the back of the unit to allow adequate air circulation Make space for the controlling computer on one side of the Fluoroskan Ascent FL or Fluoroskan Ascent The Fluoroskan As
20. before relocating the instrument from one laboratory to another Example of disinfectants Formaldehyde solution 10 Pure ethanol 70 in distilled water Virkon solution 1 3 Glutaraldehyde solution 4 Chloramine T Caution Always use disposable gloves and protective clothing and operate in a well ventilated area 1 Prepare the disinfectant 200 ml 10 formaldehyde solution or 200 ml 4 glutaraldehyde solution or another agent recommended by your safety officer 2 Empty the plate carrier 3 Switch off the power and disconnect the mains power supply cable and the computer cable 4 Disinfect the outside of the instrument using a cloth dampened with 70 pure ethanol in distilled water 5 Place the instrument in a large plastic bag Ensure that all the lids are open 6 Place a cloth soaked in the prepared solution into the bag Ensure that the cloth does not make contact with the instrument 7 Close the bag firmly and leave the instrument in the bag for at least 24 hours 8 Remove the instrument from the bag 9 Clean the instrument using a mild detergent 10 Remove any stains using 70 pure ethanol in distilled water 82 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 11 After performing this decontamination procedure label the instrument with a signed and dated Certificate of Decontamination 8 3 Certificate of decontamination The decontamination procedure is required prior to shippin
21. box 100 box 10 box 1 box 95 10 2 List of recommended spare parts Cat no Item 1 2 unit s year 10 units year 1410130 Halogen lamp 12V 30W 1 10 24071580 Complete dispensing tube 1 5 assembly 24071500 Dispensing tubing 2 8 1047661 Dispensing tip 4 20 2805690 Aspirate tube assembly 1 5 24071490 Dispenser syringe 1 0 1 5 ml and plunger 1006470 Light shield 1 3 1002820 Red adjustment collar 2 10 10 3 Ordering filters Filters play an essential role in fluorometric measurements When ordering special filters the better information you provide the better the measurement results will be A special filter order should include the following details Name of the application Excitation and emission spectra Peak or center wavelengths of the excitation and emission spectra Reference article RANE The peak or center wavelengths of both filters are not enough for optimizing the filters If you only want to order an excitation or emission filter you also have to tell which filter is used as the pair The excitation and emission spectra are the best and easiest way to design the best possible filter pair for the user If you have a reference article in addition to the name of your application the order will be perfect Enclosed you can find the filter ordering form see Appendix B 96 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Fluorescent materials absorb light energy of some characteristic waveleng
22. corner of the measurement chamber O Turn the locking piece upside down Fig 3 2b 2 22 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 7 Fit the locking piece back with the fitting screws Fig 3 2b 3 8 Refit the cover by first fixing the front corners 1 Connected locking piece with two screws 2 Disconnected locking piece turned upside down 3 Reconnected locking piece Fig 3 2b Removing the transportation lock Caution If the locking piece is not fitted into its place light may enter the measurement chamber and affect the results 3 3 Power and computer connections 1 Ensure that the power switch Fig 3 3 1 is in the OFF position 2 Connect the mains supply cable to the mains power supply socket Fig 3 3 2 The instrument box contains two different mains supply cables with North American and European types of plugs Select the correct type used in your laboratory If any other type of mains supply cable is needed use only cables certified by the local authorities 3 Connect the instrument to a correctly installed line power outlet that has a protective conductor also called ground or earth 4 Connect the serial cable to the serial connector Fig 3 3 3 and secure it with the locking screws Connect the other end similarly to the controlling computer Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 23 1 Power switch ON OFF 2 Mains power supply socket 3 Serial
23. dispenser tubings if equipped with dispensers p 25 e Plug in the instrument p 23 e Switch the instrument ON and ensure that the green light is lit p 31 e Install Ascent Software p 31 e Create an Ascent Software session see Ascent Software User s Guide Procedure Desktop Session New e We recommend you perform a dummy run first with an empty plate when you have created a new session and then a run with known samples or controls e Load the microplate with prepared samples blanks and controls Place the microplate onto the plate carrier of the instrument so that the A1 well is located in the upper left corner of the plate carrier p 32 e Start the session by pressing the START button on the Procedure Desktop toolbar of Ascent Software e Follow the instructions provided by the software e Switch the instrument OFF after routine operation p 59 e Maintain the Fluoroskan Ascent FL or Fluoroskan Ascent instrument on a regular basis p 61 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 119 120 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Appendix B Filter Order Form Thermo SCIENTIFIC Filter Order Form PO 4 Filter pairs Excitation filter Emission filter Fluorochrome fluorescent dye nm nm Specify the wavelength to the nearest 5 nm Provide the excitation and emission spectra of the Fluorochrome if possible Only excitation filter For single f
24. the sample vessel s on the corresponding adapter select the correct plate template marked with adapter from Ascent Software and run the instrument normally Note Always remember to remove the plate adapter before using the instrument with any other plate types 3 7 Adjusting the plate carrier The plate carrier of the Fluoroskan Ascent FL and Fluoroskan Ascent has been designed for plates with different footprints and to be robot compatible The plate carrier has two adjusting knobs with two different orientations These knobs have been set for standard plate types at the factory The setting is also valid for the robotic 384 adapter but readjustment of the knobs will be needed with the plate types requiring special adapters see Section 3 6 Fig 3 7 shows how the knobs should be adjusted for special plate adapters Both knobs should be rotated 90 clockwise after which all special adapters fit into the plate carrier Fig 3 7 Two plate carrier knobs rotated 90 clockwise for special plate adapters Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 29 3 8 Operational check Switch the instrument ON The instrument automatically performs a complete set of initialization tests or adjustments When the initialization tests and adjustments have been completed successfully the indicator LED turns from yellow to green If anything fails in the initialization tests or adjustments the indicator LED will turn red
25. the correct instrument model Loose serial cable Check the cable connections connections Software in simulation Switch the connection to mode the instrument Too high background No liquid in the well Always use some liquid in the blank wells Unclean plate Use a disposable plate only once Contaminated reagents Replace the reagents used Microplate material Check if the plate manufacturer or material has changed Unsuitable filter pair Ensure that the filters used are suitable for the fluorochrome Contaminated tubing Replace clean the tubing Phosphorescence from Use only plastics the plastic designed for fluorometry luminometry without phosphorescence Continued Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 71 Problem Cause Action Cont Too low high signal Changed scaling factor Check the scaling factor used Microplate material Check if the plate manufacturer or material has changed Unsuitable filter pair Ensure that the correct filter pair is used Unsuitable filter pair Ensure that the filters used are suitable for the fluorochrome Even low signal level for the whole plate Bottom reading of non transparent plate Use top reading Bottom reading when drop plate in its place Remove the drop plate Too large deviation between replicates Unclean plate Use a disposable plate only
26. the left hand side of the dispensers First push the plunger manually upwards into the upper position before tightening the plunger lock screw Fig 3 4 5 Ensure that the plunger lock screw is sufficiently tightened Note that the plunger can be extremely stiff Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 25 Aspirate tube assembly incl tubing and end weight Complete dispensing tube assembly 3 port valve Dispensing syringe 1 0 ml and plunger a fF WOW N Plunger lock screw Fig 3 4 Automatic dispenser unit 3 5 Installation of the drop plate The instrument is supplied with a special drop plate Fig 3 5a The drop plate is used to protect the instrument from damage caused by accidental dispensing without any microplate If the user forgets to place a microplate onto the plate carrier but has the drop plate in place the reagent will be dispensed into the drop plate not inside the instrument The drop plate is placed like an adapter into the plate carrier Fig 3 5b and the microplate is placed onto the drop plate The holding capacity of the drop plate is 19 ml of liquid Note The drop plate cannot be used for bottom reading 26 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Fig 3 5a Drop plate Fig 3 5b Drop plate placed into the plate carrier 3 6 Plate adapters Reading certain plate types with the Fluoroskan Ascent FL and Fluoroskan Ascent instruments requir
27. used with the plate DO NOT use plate lids with the dispensers Damaged dispensing tip Replace the tip Incorrect installation of the dispensing head Check that the dispensing head is inserted deep enough Incorrectly adjusted dispenser head assembly Contact authorized technical service Noise from the Light shield not Remove the light shield instrument removed when using FL only high plates Continued Fluoroskan Ascent FL Fluoroskan Ascent User Manual 73 Problem Cause Action Cont Noise from the instrument Lid left on the plate Remove the light shield or the lid Dispenser head installed to the incorrect height Check the dispenser plate height Filter wheel too tight Loosen the filter wheel screw Transportation lock not removed Remove the transportation lock Plate adapter for low 384 well plates left under a 96 well plate Remove the adapter when using 96 well plates Plate carrier moves incorrectly Foreign object e g a tip priming strip obstructs the plate carrier movement Ensure that there are no loose objects inside the instrument Too high background drift message Dirty measurement window or reference chip protection glass Clean the measurement window and the reference chip protection glass Accidental dispensing into the instrument Contact authorized tech
28. 0 Black Strip 1x12 assembled EB Flat 50 box 95029800 Black Breakable Strip 1x8 Assembled UB Flat 50 box Cat no Description Qty Microtiter 384 Well Plates 95040000 384 Round Well Plate 50 box 95040010 White 384 Round Well Plate 50 box 95040020 Black 384 Round Well Plate 50 box 95040130 384 Round Well Plate Sterile with lid 40 box 95040230 White 384 Round Well Plate Sterile with lid 40 box 95040330 Black 384 Round Well Plate Sterile with lid 40 box 94 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Thermo Scientific Microtiter Plates Cont Cat no Description Microtiter Streptavidin Coated Plates 95029263 95029293 95029273 95029303 95029283 Cat no BioBind Strip 1x8 Assembled BioBind Breakable Strip 1x8 assembled White BioBind Strip 1x8 Assembled White BioBind Br Strip 1x8 Assembled Black BioBind Strip 1x8 Assembled Description Microtiter Plate Accessories 5500 5550 9503210 2 9503220 6305 6604 6000 Universal Polystyrene Lid Sterile Styrene Individually Wrapped Lid Sterile Microplate Lid 96 well Microplate Lid Vinyl Lid for 1x12 strip assembly Holder for 1x12 strip assembly Workstation 1x12 strip assembly Accessories for ex Dynex products 2 Accessories for ex Thermo Scientific products Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Bottom Flat Flat Flat Flat Flat Qty 5 box 5 box 5 box 5 box 5 box Qty 100 box 50 box 15 box 15
29. 2 The Fluoroskan Ascent FL is a combined high quality microplate fluorometer and luminometer intended for laboratory research use by professional personnel The Fluoroskan Ascent FL is used to measure fluorescence or luminescence from suitable 1 to 384 well plates mentioned in this manual It also has incubation shaking and reagent dispensing capabilities 3 For verification of the entire system it is recommended that Good Laboratory Practices GLP be followed to guarantee reliable analyses 4 Use for self testing is excluded 10 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 1 2 Method descriptions 1 2 1 Fluorometric measurement principle Measurement with the Fluoroskan Ascent FL and Fluoroskan Ascent occurs in a narrow angle The source of the excitation light and the detector of the emission light are both located on the same side of the microplate Filters are held in 8 position filter wheels The Fluoroskan Ascent FL Fluoroskan Ascent can carry out both top and bottom reading The position of the whole optic unit is easily changed 1 2 2 Luminometric measurement principle Fluoroskan Ascent FL A luminometric measurement uses the same optics as the fluorometric but the lamp is switched off The emission filter slots 7 and 8 are reserved only for luminometric measurements Filter slot 7 is empty for measurement and filter slot 8 is blocked to enable measurement of the PMT photomultiplier tube dark current Thi
30. 2 1425382 or 527 or 1425182 or 1425272 Fluo 3 485 538 1424852 1425382 or 527 or 1425272 Fluo 4 485 518 1424852 1425182 Fluorescein 485 538 or 518 1424852 1425382 or or 527 1425182 or 1425272 Ex Excitation filter Em Emission filter Continued Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 99 Cont Fluorochrome Ex Em Filters Cat no Notes Fluorescein diacetate 485 538 1424852 1425382 Fura 2 340 and 1423402 and Only slow 380 510 1423802 1425102 Ca flux Hoechst dyes 33258 and 355 460 1423552 1424602 33348 HPPA 320 405 1423202 1424052 4 Methylumbelliferone 355 460 1423552 1424602 OliGreen 485 538 1424852 1425382 Phenylalanine 390 485 1423902 1424852 Ninhydrin reaction R Phycoerythrin 544 590 1425442 1425902 PicoGreen 485 518 14248852 1425182 or or 538 1425382 Resorufin 544 590 1425442 1425902 Rhodamine 110 485 518 1424852 1425182 RiboGreen 485 538 1424852 1425382 Rox 578 604 142578206 1426042 StarBright Green 444 510 1424442 1425102 Sulforhodamine 101 584 612 1425842 1426122 Tamra 544 590 1425442 1425902 Texas Red 584 612 1425842 1426122 TRITC 544 590 1425442 1425902 wtGFP 390 510 1423902 1425102 Ex Excitation filter Em Emission filter 100 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 11 Glossary and Abbreviations ATP Chassis Chemiluminescence Crossover point Crosstalk Cr
31. 20 54 62 64 67 69 71 73 81 IAI kver kue kane as SE SE EET Een 64 RFE uk SS ed 35 103 113 S SA O 35 36 38 72 76 103 Tactics 35 36 72 76 103 Selassie oro 10 102 103 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 117 Shaker e ii A ts 10 39 72 110 113 SHUTH OWN ist cdi aner 51 55 56 59 62 64 70 82 Sia a E AIA li 72 Spacer colla ui taa 58 Specificatii NSi errendi eiee eaa aia iia 85 109 110 111 114 MV iaa a ada A ai 111 Spring WNES Z oe aaen oi 58 Koo AETA DT EPEAN ne E AS AS T A aboutus bear A E T 33 97 103 SOPP uria a a aka en 46 SWit hing ON cuisine a AT Tied 24 31 59 T TIP pMn graa EG kl aida 18 44 45 59 72 74 TOPE Dacia bs 37 50 72 STRAWS uastsdusrneGleeGaatkeeeind 20 Transportation lOCK cccccccccccncncnonononnncnnnononononononononononos 20 23 74 85 119 Troubleshooting esinin en raian aaa ia 81 fo U Mo RE E A AA ee a ER SEEREN 71 U Unpacking sis sisal ida tee els ise dalen 19 119 V Validation 33 34 58 102 W Warnings ici ee 7 21 24 29 31 74 W rranty usariaddidte adjunkt iaaa aa aeiaai 19 62 108 118 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Appendix A Brief User s Guide e Unpack the instrument p 19 e Check delivery for completeness p 20 e Check for damage during transport p 20 e Place the instrument according to environmental requirements p 21 e Release the transportation lock p 22 e Install the
32. 2001 EN 61000 6 1 2001 EN 61326 1 1997 A1 1998 A2 2001 A3 2003 Test standards EN 55022 1998 EN 61000 3 2 2000 EN 61000 3 3 1995 A1 2001 ANSI C63 4 2003 EN 61000 4 2 1995 A1 1998 A2 2001 EN 61000 4 3 2002 A1 2002 EN 61000 4 4 2004 EN 61000 4 5 1995 A1 2001 EN 61000 4 6 1996 A1 2001 EN 61000 4 8 1993 A1 2001 EN 61000 4 11 1994 A1 2001 112 Generic emission standard Residential commercial and light industry Generic immunity standard Residential commercial and light industry Product family standard Performance limits Class B 150 kHz 1 GHz Class A Class B 450 kHz 1 GHz 30 MHz 1000 MHz 4 kV CD 8 kV AD Criteria B 3 V m 80 MHz 2 GHz Criteria A 1 kV Criteria B 2 kV line to ground 1 kV line to line Criteria B 3 Vims 150 kHz 80 MHz Criteria A 3 A m Criteria A 30 10 ms Criteria B 60 100 ms Criteria C gt 95 5 s Criteria C 100 20 ms Criteria B Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 14 4 Performance specifications Warm up time lt 15 min to rated accuracy Measuring speed Depends on the plate type and the measurement type The minimum kinetic interval time is 15 s for a 96 well plate from well A1 back to the same well A1 Excitation wavelengths 320 700 nm Fluorometric measurement range Up to approx 5000 Relative Fluorescence Units RFU Emission wavele
33. 52 Emission filter 485 nm 14 nm 1425102 Emission filter 510 nm 10 nm 1425182 Emission filter 518 nm 12 nm 1425202 Emission filter 520 nm 10 nm 1425272 Emission filter 527 nm 15nm 1425382 Emission filter 538 nm 25 nm 1425552 Emission filter 555 nm 25 nm 1425902 Emission filter 590 nm 14 nm 1426042 Emission filter 604 nm 10nm 1426122 Emission filter 612 nm 6 nm 1426202 Emission filter 620 nm 10 nm 142678210 Emission filter 678 nm 10nm 1426802 Emission filter 680 nm 20 nm Specified to be used as either excitation or emission filter Other wavelengths available on request see Section 10 3 Ordering filters Upgrade Kits for Fluoroskan Ascent Item Cat no 1st Dispenser Kit 2805620 Additional Dispenser Kit 2805630 Upgrade Kits for Fluoroskan Ascent FL Item Cat no 1st Dispenser Kit 2805621 Additional Dispenser Kit 2805630 Wide Wavelength Range Kit PMT 2805760 90 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Accessories Item Adapter for Costar Petri dish 95 mm Adapter for Falcon Greiner Petri dish 93 mm Adapter for Petri dish 2 x 59 mm Adapter for low 384 well plates Robotic 384 adapter Adapter for Petri dish 6 x 40 mm Adapter for Terasaki plate Adapter for PCR tubes and 96 well PCR plates Drop plate Halogen lamp Light shield Dispensing tip Complete dispensing tube assembly Fig 3 4 2 and Fig 5 7 1 Dispensing tubing Fig 5 7 1 7 Dispensing tip Fig 5 7 1 2 10 pie
34. Ascent User Manual Symbols and Markings SYMBOLS USED IN THE FLUOROSKAN ASCENT FL AND FLUOROSKAN ASCENT Power ON C Power OFF Connection to the protective grounding system WARNING MARKINGS USED ON THE INSTRUMENT Caution risk of electric shock Caution risk of personal injury to the operator or a safety hazard to the surrounding area See the accompanying documentation CE compliance mark WEEE symbol This product is required to comply with the European Union s Waste Electrical Electronic Equipment WEEE Directive 2002 96 EC ap WARNING MARKINGS USED IN THE DOCUMENTATION Caution risk of electric shock Caution risk of personal injury to the operator or a safety hazard to the surrounding area Caution risk of serious damage to the instrument other equipment or loss of performance or function in a specific application Caution biohazard risk Pb Fluoroskan Ascent FL Fluoroskan Ascent User Manual 7 Fluoroskan Ascent Cat no 5210470 and 5210480 with dispenser Fluoroskan Ascent FL Cat no 5210450 and 5210460 with dispenser User Manual Rev 2 4 Nov 2008 Cat no 1506450 Copyright 2008 Thermo Fisher Scientific Corporation All rights reserved First edition published in 1995 Reproduction of the accompanying user documentation in whole or in part is prohibited Patents The Fluoroskan Ascent FL and Fluoroskan Ascent have national and international patents and patents pending
35. F Herring P J Robison B H Haddock S H D Kricka L J and Stanley P E eds World Scientific Publishing Co Pte Ltd Singapore 517 pages Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 105 106 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 13 Warranty Certificate Thermo Fisher Scientific Microplate Instrumentation Business products are fully guaranteed against defective parts and materials including defects caused by poor workmanship for a period of one year from the date of delivery Thermo Fisher Scientific will repair or replace defective parts or materials during the term of warranty at no extra charge for materials and labor provided that the products were used and maintained in accordance with Thermo Fisher Scientific s instructions The warranty is invalid if products have been misused or abused For the warranty to be effective the product must have been purchased either directly from Thermo Fisher Scientific or from an authorized Thermo Fisher Scientific distributor The guarantee is not transferable to a third party without prior written approval from Thermo Fisher Scientific This guarantee is subject to the following exclusions e Any defects caused by normal wear and tear e Defects caused by fire lightning flood earthquake explosion sabotage war riot or any other occurrence of the type listed above e Refurbished products that are subject to different warranty co
36. How is the scaling factor in Ascent Software used A The scaling factor is a constant that multiplies all the readings from the instrument It is defined separately for each filter pair and beam size fluorometry or filter position luminometry The scaling factor can be used to convert the instrument reading to a desired level or to adjust several instruments to give similar signal values Q What plate colors can be used for fluorescence A Best sensitivity is generally obtained with black plates which also have the lowest background fluorescence However with some fluorochromes white plates can also be used for obtaining a slightly better sensitivity Transparent plates can be used but the sensitivity is slightly lower and the difference is dependent on the filter pair used When you carry out bottom reading only transparent or transparent bottomed plates can be used Q When can I use white plates in fluorometry In assays such as DNA quantitation with PicoGreen white plates can be used as the obtained fluorescent signal level is not very high The sensitivity is also slightly better when using white plates White plates can also be used in for example GFP quantitation however white plates should not be used when the signal level is very high 76 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual FAQs about the Fluoroskan Ascent FL amp Fluoroskan Ascent Cont Q What plate colors can be used for luminesc
37. L amp Fluoroskan Ascent User Manual 123 NOTES 124 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual NOTES Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 125 NOTES 126 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual
38. Nitric Acid o o o Nitric Acid conc Nitrobenzene o Phenol Pyridine 3 Silver Nitrate Soap Solutions I D 0o Stearic Acid Sulfuric Acid o 0 Sulfuric Acid conc 3 Sulfurous Acid OQIOIO IuM JO Ju N o o 0o 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0D o0 o ojojojo ojo Tannic Acid Tannin Extracts Tartaric Acid Toluene Trichloroethylene Turpentine Es OlNIuM I VM ojojojojo ojojoj Water Xylene 3 0 0 ld Polypropylene satisfactory to 22 C 72 F Polypropylene satisfactory to 49 C 120 F Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 49 4 5 6 Excitation light beam selector Excitation beam selection means a possibility to select the normal beam with a diameter of 3 mm in the measurement well or the small beam with a diameter of 1 5 mm The small beam is required to measure for example 384 well plates If the normal beam is used with 384 well plates the light beam also excitates the walls of the well and the result is not optimal Measurement of larger wells with the small beam is possible and depends on the application EXCITATION BEAM Fig 4 5 6a Beam selection when carrying out top reading The selection lever of the beam is on the front panel of the optical unit When the optical unit is above the plate the selection knob can be
39. a the not recommended Possible and the white area the prevented speed and diameter combinations Not allowed Table 4 5 1a Shaking speeds for 48 or more well plates 1200 1140 4 Recommended Not allowed 1080 1020 960 900 840 780 E 7204 amp 660 4 5 6004 g 54014 G amp 4804 420 360 300 240 4 180 120 60 0 1 3 5 7 9 11 1315 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 Diameter mm Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 39 40 Speed rpm Speed rpm Table 4 5 1b Shaking speeds for 24 well plates 1200 1140 gt 1080 Recommended W Possible O Not allowed 7 1020 gt 960 900 840 780 gt I 720 420 360 300 gt 240 180 120 4 60 4 0 660 600 540 gt 480 1 Table 4 5 1c 1200 3 5 7 9 11131517 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 Diameter mm Shaking speeds for 12 well plates 1140 gt
40. anual FAQs about the Fluoroskan Ascent FL amp Fluoroskan Ascent Cont Q What kind of plate adapters are available for the Fluoroskan Ascent FL and Fluoroskan Ascent A The adapters listed below are available Other adapters are available on request 5210310 5210300 5210380 5210330 5210320 5210390 5210340 5210630 Adapter for Petri dish 2 x 59 mm Adapter for Costar Petri dish 95 mm Adapter for Falcon Greiner Petri dish 93 mm Adapter for Petri dish 6 x 40 mm Adapter for low 384 well plates Robotic 384 adapter Adapter for Terasaki plates Adapter for PCR tubes and 96 well PCR plates Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 79 80 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 8 Instrument Service 8 1 Service request protocol If the Fluoroskan Ascent FL or Fluoroskan Ascent requires service contact your local Thermo Fisher Scientific representative or the Thermo Fisher Scientific technical service department DO NOT under any circumstances send the instrument for service without any prior contact It is imperative to indicate the fault and nature of the required service This will ensure a faster return of the instrument to the customer The Thermo Fisher Scientific representative or distributor takes care of sending the Thermo Fisher Scientific technical service department a Feedback Form i e the Complaint Order which contains a more detailed description of the
41. are measured according to the selected measurement parameters The A D converter has several gain steps but the instrument automatically selects the best gain in each measurement The measurement is performed with a selected integration time The integration time can be from 20 to 1000 ms which defines the length of time used to obtain one measurement result The minimum time is 20 ms default during which time the instrument sums 150 readings from the A D converter If the integration time is longer the result is the mean value of the number of 20 ms readings during the total integration time Longer than 20 ms integration times can be used to reduce background noise if the signal level of the sample is very low With small Stokes shift values optical crosstalk of excitation and emission filters may dominate the Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 33 instrument background noise In these cases an extended integration time does not help The plate carrier acceleration and maximum speed are adjustable see the Ascent Software User s Guide General step Settings samples high speed and acceleration may cause splashing Caution When measuring large wells with low viscosity and extra variation in the results Use the default acceleration 2 for 48 well plates and the default acceleration 1 for microplates with 24 or less wells the plate carrier door when measurement is started DO NOT Note Ens
42. ate and place Signature Name block capitals 84 Fluoroskan Ascent FL Fluoroskan Ascent User Manual 8 4 Shipping the instrument or items When you ship the instrument for service remember to 1 L NBR 10 11 Empty the dispenser s and remove the tube assembly Remove any loose items from the plate carrier for example adapters plates and priming vessels Remove the power cable as well as the serial cable Decontaminate the instrument beforehand Fit the transportation lock Pack the instrument according to the enclosed packing instructions Use the original packaging to ensure that no damage will occur to the instrument during shipping Any damage will incur additional labor charges Inform about the use of hazardous materials Enclose a dated and signed Certificate of Decontamination see Section 8 3 Certificate of Decontamination both inside and attached to the outside of the box in which you will return your instrument or other items Indicate the fault after you have been in touch with your local Thermo Fisher Scientific representative or the Thermo Fisher Scientific technical service department Enclose the return authorization number RGA given by the Thermo Fisher Scientific representative See Section 14 1 General specifications for details on storage and transportation temperatures 8 5 Service contracts We strongly recommend that you maintain and service the instrument eve
43. ation Fluorescein Fluorescence Fluorochrome Fluorophore Fluorometer Fluorometry HBW Initialization LED Luminescence Luminometer Luminophore Photomultiplier tube PMT 102 Procedure in which a new filter pair is introduced to the autocalibration system The instrument reads the fluorescence of all the reference chips and selects the chip for which the value is most suitable The selected value is then saved in memory for the autocalibration procedure An example of a fluorescent dye emitting green fluorescence The emission of light from a fluorochrome the wavelength of the light generally being of longer wavelength than that of the absorbed light A molecule or chemical group which emits fluorescence Instrument used for measuring the intensity of fluorescent radiation Also known as fluorimeter Fluorometry is the measurement of fluorescence Half bandwidth of filters Initialization tests are so called self tests which are carried out prior to operation to ascertain that the necessary instrument adjustments have been carried out Light emitting diode Emission of light other than from thermal energy causes such as bioluminescence An instrument used for measuring the intensity of luminescent radiation A substance which emits light at room temperature A group of atoms that can make a compound luminescent A photoelectric cell that converts light into electric current
44. bled C 100 box 6506 Immulon 2 HB 2x8 Strip assembled Flat 100 box 6522 Immulon 2 HB 2x8 Strip assembled C 100 box 6405 Immulon 4 HBX 1x12 Strip assembled Flat 100 box 6404 Immulon 4 HBX 1x12 Strip Flat 320 box 6484 Immulon 4 HBX 1x8 Strip assembled C 100 box 6508 Immulon 4 HBX 2x8 Strip assembled Flat 100 box 6524 Immulon 4 HBX 2x8 Strip assembled C 100 box 95029350 Strip 1x8 assembled UB Flat 50 box 95029100 Strip 1x8 assembled EB Flat 50 box 95029440 Strip 1x8 assembled UB Round 50 box 95029370 Strip 1x12 assembled UB Flat 50 box 95029140 Strip 1x12 assembled EB Flat 50 box 95029390 Breakable Strip 1x8 Assembled UB Flat 50 box 95029180 Breakable Strip 1x8 Assembled EB Flat 50 box 95029430 Breakable Strip 1x8 Assembled EB Round 50 box 92 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Thermo Scientific Microtiter Plates Cont Cat no Description Microtiter Plates for Luminescence 7416 7417 7418 7571 7572 7521 7522 9502887 95029580 95029770 Cat no Microlite 1 Plate Microlite 2 Plate Microlite TCT Plate sterile Microlite 1 Plate Microlite 2 Plate Microlite 1 Plate Microlite 2 Plate White 96 Well Plate UB White 96 Well Plate EB White 96 Well Plate Sterile Description Microtiter Plates for Luminescence Strips and Assemblies 7421 7403 7400 7410 7561 7566 7562 7567 95029510 95029530 95029660 Microlite 1 1x12 Strip assembled Microlite 1 1x12 Strip Microlite 2 1x12 Strip assemb
45. calibration procedure Ascent Software shows the validated filter pairs and those pairs which uyn have not found a suitable chip are marked with Fluoroskan Ascent FL Fluoroskan Ascent User Manual 4 3 2 Scaling The scaling factor is a value that multiplies all the readings from the instrument It is defined separately for each filter pair fluorometry or filter position luminometry The scaling factor can be used to convert the instrument reading to a desired level or to adjust several instruments to give similar signal values 1 The measured results are expressed as Relative Fluorescence Units RFU Scaling is a way to convert readings to show desired values The normal beam and the small beam have separate scaling factors 2 Prepare a scaling reference solution and a blank solution and pipette several wells of blanks and the known concentration 3 Measure the wells using the correct filter pair and beam 4 Calculate the average values of the measured reference values and blank values 5 Calculate the scaling factor using calculated average values Factor Known reference Measured reference Measured blank Example The known concentration is 500 pmol well The calculated average of the measured reference wells is 1825 RFU The calculated average of the measured blank wells is 0 2 RFU Factor 500 1825 0 2 0 274 After scaling the measured result is 0 274 x 1825 RFU 500 pmol well
46. cent FL or Fluoroskan Ascent does not produce operating noise at a level which could be harmful No sound level measurements are needed after installation Warning DO NOT operate the instrument in an environment where potentially damaging liquids or gases are present 3 1 5 Things to avoid DO NOT smoke eat or drink while using the Fluoroskan Ascent FL or Fluoroskan Ascent Wash your hands thoroughly after handling test fluids Observe normal laboratory procedures for handling potentially dangerous samples Use proper protective clothing Use disposable gloves Be sure the working area is well ventilated Never spill fluids in or on the equipment Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 21 3 1 6 Technical prerequisites Place the instrument on a normal laboratory bench close to the mains power supply socket The net weight of the unit is approx 21 kg 46 Ibs The instrument operates at voltages of 100 240 Vac The frequency range is 50 60 Hz 3 2 Releasing the transportation lock Cover retaining screws 4 pieces Dispenser and optics cover Rear of the cover Point to where cover lifted Fig 3 2a Removing the instrument cover 1 Remove the four cover retaining screws Fig 3 2a 1 2 Open the dispenser and optics cover Fig 3 2a 2 3 Lift the rear of the cover at first about 3 cm Fig 3 2a 3 4 Lift the cover aside Fig 3 2a 4 5 Undo the two screws Fig 3 2b 1 at the right rear
47. ces min order Aspirate tube assembly Fig 3 4 1 incl tubing and end weight Dispenser syringe 1 0 ml and plunger Fig 3 4 4 Syringe seal 3 port valve Leakage tray Fig 2 3 4 Red adjustment collar Fig 4 5 4b 2 RS 232C interface cable D9 Female D25 Female Dust cover Fluoroskan Ascent and FL upgrade Cat no 5210300 5210380 5210310 5210320 5210390 5210330 5210340 5210630 2805880 1410130 1006470 1047661 24071580 24071500 1047661 2805690 24071490 2805600 2805610 10478030 1002820 2305290 1610460 2805660CD 91 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Thermo Scientific Microtiter Plates Cat no Description Bottom Qty Microtiter Plates for Immunoassay 3355 Immulon 1B Plate Flat 50 box 3555 Immulon 1B Plate U 50 box 3455 Immulon 2 HB Plate Flat 50 box 3655 Immulon 2 HB Plate U 50 box 3855 Immulon 4 HBX Plate Flat 50 box 9502227 96 Well Plate UB Flat 50 box 95029330 96 Well Plate EB Flat 50 box 95029780 96 Well Plate Sterile Flat 40 box Cat no Description Bottom Qty Microtiter Plates for Immunoassay Strips and Assemblies 6310 Immulon 1 B 1x12 Strip assembled Flat 100 box 6301 Immulon 1 B 1x12 Strip Flat 320 box 6381 Immulon 1 B 1x8 Strip assembled C 100 box 6505 Immulon 1 B 2x8 Strip assembled Flat 100 box 6521 Immulon 1 B 2x8 Strip assembled C 100 box 6309 Immulon 2 HB 1x12 Strip assembled Flat 100 box 6302 Immulon 2 HB 1x12 Strip Flat 320 box 6382 Immulon 2 HB 1x8 Strip assem
48. connector Fig 3 3 Power and computer connections Warning Always make sure the power switch on the instrument is in the OFF position and remove the mains power supply cable from the back of the instrument prior to any installation or relocation of the instrument Warning Never operate your instrument from a power outlet that has no ground connection gt gt 24 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 3 4 Installation of dispensers The optional dispensers 1 to 3 are installed in number order from left to right The complete dispensing assemblies are packed with the accessories The aspirate tubing Fig 3 4 1 is factory installed into the left hole of the valve Ensure that the aspirate tubing is finger tight If necessary turn the fitting another quarter to half turn using a 7 9 mm 5 16 in wrench The aspirate tubing is used to fill the syringe with reagent When using the dispensers make sure that the aspiration tube end is completely submerged in the reservoir and there is a sufficient volume of the reagent in the reservoir for all primings and actual dispensing Fit the complete dispensing tube assembly Fig 3 4 2 into the right hole of the valve and tighten it finger tight Then turn the fitting another quarter to half turn using a 7 9 mm 5 16 in wrench The dispensing tube is used to dispense reagent from the syringe into a microplate Place the dispensing heads in the dispensing head holder on
49. d contaminating neighboring wells The correct tip height is also important to prevent damaging the tip or the plate Note Ensure that the dispenser tips are always correctly inserted sufficiently deep into their slots Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 45 The plate height is one of the template parameters in Ascent Software and it is defined as the height of the uncovered well from the bottom of the plate not the inside height of the well The selected dispenser head height the used plate and the template selected in Ascent Software must match to avoid problems To see the heights of plates in Ascent Software select Setup and edit Plate Templates Some 384 well plates are lower than the standard 96 well plates If the plate height is less than 13 5 mm an adapter must be used to raise the plate Measure the height of the plate and the adapter together and enter this value to the plate template Caution Plate manufacturers may change the dimensions of plates without any prior notice or change in order numbers Check the dimensions when you start using plates from a new box The dispenser head height is adjusted with red adjustment collars by moving them from either side of the fixed stopper collar Fig 4 5 4b BEE ET B TL TE E 1 Dispenser head tube 2 Red adjustment collars Fig 4 5 4b Changing the position of the red adjustment collars 1 Remove the dispenser head tube Fig 4 5 4b 1 from
50. de the plate carrier door when measurement is started DO NOT Note Ensure that there are no items preventing the closure of open the plate carrier door during measurement 4 4 1 Luminometric scaling The measured results are expressed as Relative Light Units RLU Scaling is performed similarly as in fluorometric scaling see Section 4 3 2 Scaling 38 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 4 5 Other functions 4 5 1 Orbital shaking The track movement system can perform the shaking action The speed is adjustable from 60 to 1200 rpm revolutions per minute and the diameter of the orbital movement is adjustable from 1 to 50 mm Some combinations of speed and diameter would cause too high g forces inside the well area resulting in spills inside the measurement chamber Therefore only certain combinations are available When you use plates with small wells such as plates with 48 wells or more you can select any of these available speed and diameter combinations to serve your application best Table 4 5 1a The following tables Table 4 5 1a Table 4 5 1e show the recommended and the not recommended but available and unavailable speed and diameter combinations with different plate types These tables are based on the liquid used being of low viscosity like water and the volumes being appropriate the wells are not full The gray area in the tables show the recommended Recommended the black are
51. de wavelength range option for the Fluoroskan Ascent FL A The Fluoroskan Ascent FL photomultiplier tube PMT provides good sensitivity in both fluorometry and luminometry With this PMT the emission wavelength range for the Fluoroskan Ascent FL ranges from 360 to 670 nm If the Wide Wavelength Range Option is selected the PMT of the Fluoroskan Ascent is installed into the FL The emission wavelength range which can then be measured ranges from 360 to 800 nm However the luminometric sensitivity is decreased about one decade The fluorometric sensitivity is the same with both types of PMTs Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 75 FAQs about the Fluoroskan Ascent FL amp Fluoroskan Ascent Cont Q How do the fluorescence reference chips work which are installed into the Fluoroskan Ascent FL and Fluoroskan Ascent plate carrier A The fluorescent chips are installed into the unit to keep the instrument signal level stable When a filter pair is validated the instrument measures all the four chips in the plate carrier and then automatically selects the most suitable chip The chip measurement value is stored in the instrument memory When the measurement is started autocalibration is performed The instrument measures this chip during the autocalibration process and compares it with the value in memory The correction factor is calculated and all further measurement values are multiplied by this factor Q
52. ded cleaning maintenance instructions are followed Both fluorometry and luminometry are very sensitive detection technologies Therefore take special care to avoid any contamination of any parts of the dispenser tubings and follow all GLP Good Laboratory Practice recommendations 5 5 1 Basic maintenance 1 The basic maintenance procedure should be performed regularly to ensure proper dispenser operation 2 Rinse the dispenser tubings out thoroughly with distilled water after each use 3 DO NOT allow the dispensers to run dry for more than a few cycles 4 Inspect the dispensers for leaks and rectify any problems immediately 5 Wipe up all spills on and around the dispensers immediately 5 5 2 Extended maintenance Clean the fluid path thoroughly using one of the procedures outlined below There are three ways that the dispensers may be cleaned e Weak detergent 10 bleach e Weak acid and base 5 5 2 1 Weak detergent or 10 bleach Remove the dispensing heads from the dispensing positions and DO NOT let any cleaning fluids enter the measurement chamber Use external containers 1 Prime the dispensers with a weak detergent or 10 bleach solution and leave it in the dispensers with the syringes full for 30 minutes 2 After the 30 minute period remove the aspirate tubing from the detergent or bleach solution and remove all the fluid from the syringes and tubing into a waste container Fluoroskan Ascent FL amp Fluor
53. e recommended volume is 2700 pl The Fluoroskan Ascent FL also has control switches for priming the dispenser tubing With these switches priming can be carried out alternatively by using an external priming vessel or by using an empty microplate on the plate carrier as a priming vessel When priming is performed with the dispensing heads installed into the M X or Y positions place the empty priming vessel into the plate carrier Initiate priming by pressing the corresponding switch when the plate carrier is located outside the unit Priming is carried out as long as the corresponding switch is pressed Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 43 the plate carrier is located outside the instrument DO NOT use priming vessels higher than the actual plate intended to be used in the assay Notice the dispensing height adjustment Note The priming control switches are functional only when Priming with Ascent Software must always be performed with the dispensing heads removed from the M X or Y dispensing positions clotting or contain any mechanical particles with the automatic Note Never use liquids that can cause any precipitation dispensers The instrument has a Prime Tip feature If this function is selected in the Dispense or Dispense And Measure steps in Ascent Software the dispenser dispenses about 5 ul reagent into the tip priming vessels every time the instrument fills the syringe This makes the v
54. e special plate adapters Fig 3 6 These plate adapters are required for the following reasons Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 27 1 Raising the 384 well plate with a height of less than 15 mm to the proper height for luminometric measurement 2 If the 384 well plate height is less than 13 5 mm an adapter must be used to raise the plate for dispensing 3 Positioning of other sample vessel types than microplates for example Petri dishes or Terasaki plates onto the plate carrier for measurement and dispensing 4 Supporting flexible PCR plates and tubes on the plate carrier for measurement and dispensing When using any adapter measure the height of the plate and the adapter together and ensure that this value is correctly entered into the Ascent Software plate template parameter file Refer to the Thermo Scientific Ascent Software User s Guide for further information LABSYSTEMS OY 5210630 Adapter for Petri dish 6 x 40 mm Adapter for 96 well PCR plates Adapter for Petri dish 93 mm b Q N Adapter for low 384 well plates Fig 3 6 Example of plate adapters 28 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Note If the orientation of the plate adapter is important the correct orientation is printed on the adapter or the adapter has been designed in such a way that only one orientation is possible When the adapter is needed place it into the plate carrier Then place
55. ea is measured beam The black area between the This is the recommended way measurement points is not measured at all Fig 4 5 7 Area measurements 4 6 Changing the measurement direction The measurement direction can be changed by moving the whole optical unit from above the measurement chamber to below the measurement chamber or vice versa 1 Switch off the instrument and disconnect the mains power supply cable Fig 3 3 2 There are anode voltages inside the mains power supply box and the optical unit if the power is on 2 Remove the instrument cover as described in Section 3 2 Releasing the transportation lock Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 51 3 Undo the four finger nuts Fig 4 6a 2 to release the light cover Fig 4 6a 1 4 Remove the light cover 1 Light cover 2 Finger nuts 4 pieces Fig 4 6a Removing the light cover of the optical unit 5 Undo the six screws three screws on each side of the instrument Fig 4 6b 1 fixing the measurement chamber to the chassis 6 Lift the measurement chamber into the upper position from the handle Fig 4 6b 2 There are hinges in the rear and a gas spring Fig 4 6c 3 holds the chamber in the upper position leave your fingers between the measurement chamber and Caution The measurement chamber is very heavy DO NOT DA the bottom case 7 Remove the three short finger nuts holding the cover Fig 4 6c 2 Then remove t
56. ence A White plates are generally used for luminometric measurements to enhance the signal and to avoid possible crosstalk from high signals in neighboring wells Q Can dispensing be performed into 864 well plates A No dispensing into 864 well plates is not possible Q Can the light shield be removed from the Fluoroskan Ascent FL while carrying out fluorometric measurements A Yes as the light shield is only needed with luminometric measurements Q When is the light shield needed with the Fluoroskan Ascent FL A The light shield is needed in luminometric measurements and does not interfere with fluorometric measurements The light shield should be used with 96 and 384 well plates If the plates are higher than 15 mm the light shield must be removed Q How many filters can be installed into the Fluoroskan Ascent FL and Fluoroskan Ascent A Both excitation and emission filter wheels in the Fluoroskan Ascent can hold a maximum of eight filters The Fluoroskan Ascent FL excitation filter wheel can hold a maximum of eight filters and the emission wheel a maximum of six filters Q What plate types can be measured with the Fluoroskan Ascent A The Fluoroskan Ascent FL and Fluoroskan Ascent can measure plate formats from 1 to 864 Customized plates with maximum dimensions of 90 mm x 143 mm x 25 mm can be used Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 77 FAQs about the Fluoroskan Ascent FL amp Flu
57. ensing head tube Fig 5 6 1 1 from the brass tube holder Fig 5 6 1 5 Loosen the brass locking ring Fig 5 6 1 6 Remove the dispensing tip Fig 5 6 1 2 and the connecting piece of silicone tubing Fig 5 6 1 3 Remove the internal dispenser head tube Fig 5 6 1 4 the brass tube holder and the brass locking ring Insert the new dispensing tubing into the brass tube holder locking ring and the internal dispenser head tube Connect the dispensing tip with the connecting piece of silicone tubing into the new dispensing tubing Fit the dispensing tip and tubing into the dispenser head tube When you push the dispensing tubing gently towards the tip the internal dispenser head tube should be visible about 1 mm This can be adjusted through the length of the connecting piece of silicone tubing First screw the brass tube holder onto the dispenser head tube Gently tighten the tubing and ensure that the tube end stays in the silicone tubing Then tighten the brass locking ring To fit a new tubing insert the fitting into the valve and tighten it finger tight Using a 7 9 mm 5 16 in wrench turn the fitting another quarter to half turn Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 5 6 2 Replacing the dispensing tip 1 Dispensing head tube 2 Dispensing tip 3 Brass tube holder Fig 5 6 2 Replacing the dispensing tip 1 Remove the dispensing head tube Fig 5 6 2 1 from the brass tube holder Fi
58. ent User Manual 4 9 Shutdown e Remove any microplates left on the plate carrier or breakable strips from the tip priming vessel Dispose of all microplates and strips as biohazardous waste e Flush the pump s out thoroughly with distilled water Empty all the tubings e Place the dispensing tips into the tip holder Fig 2 3 5 e Switch off the Fluoroskan Ascent FL Fluoroskan Ascent by pressing the power switch on the left hand side of the instrument into the OFF position e Wipe the plate carrier surface and the external surfaces of the instrument with a soft cloth or tissue paper moistened with distilled water or a mild detergent solution e Push the plate carrier manually in e If you have spilt infectious agents on the plate carrier disinfect with 70 pure ethanol in distilled water or some other disinfectant See Section 8 2 Decontamination procedure e If you are not using the Fluoroskan Ascent FL Fluoroskan Ascent for an extended period of time always clean the external surfaces of the instrument e Finally put the dust cover on Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 59 60 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 5 Maintenance to biohazardous infectious radiologic or toxic materials when A Note Follow normal laboratory safety procedures with regard maintaining the instrument 5 1 Routine cleaning of the instrument For reliable operation keep the instrum
59. ent free of dust and spills of liquids We recommend you clean the case of the instrument periodically A soft cloth dampened in mild detergent is sufficient We recommend you service the instrument at least yearly If you believe liquid has entered the fluorometer switch the instrument off and contact your local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy for technical service see Sections 8 1 Service request protocol and 8 4 Shipping the instrument or items If any surfaces have been contaminated with biohazardous material a sterilizing solution must be used The prescribed decontamination procedure see Section 8 2 Decontamination procedure or similar routine must be performed before returning the instrument to the supplier for service or repair All instruments must be accompanied by a completed and signed Certificate of Decontamination securely attached to the exterior of the packaging see Section 8 3 Certificate of Decontamination Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 61 5 2 Cleaning the optical system Fig 5 2 Cleaning the optical unit 1 Switch off the instrument and disconnect the mains power supply cable Locate the optical unit and remove it according to the instructions in Sections 3 2 Releasing the transportation lock and 4 6 Changing the measurement direction 2 Clean the main lens Fig 5 2 the filters and the light shield Fig 4 7a with a cloth dampened w
60. est Ab Cold IEC 60068 2 2 test Bb Dry heat Storage conditions 25 C 50 C packed in transportation packaging Tested according to IEC 60068 2 1 test Ab Cold IEC 60068 2 2 test Bb Dry heat Mains power supply 100 240 Vac 50 60 Hz nominal Power consumption 200 VA max 32 VA standby Heat dissipation 683 BTU max Computer interface Serial RS 232C port Baud rate 9600 Character format 1 start bit 8 data bits 1 stop bit no parity Flow control XON XOFF Light source Quartz halogen lamp Power consumption 30 W Continued Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 109 General specifications Cont Detector Photomultiplier tube Fluorescence reference Lisa KL 3 9402 Polymethylmethacrylate Filters for fluorometric measurements Both the excitation and emission filter wheels can comprise max eight selectable filters Fluoroskan Ascent However in the Fluoroskan Ascent FL eight excitation and six emission filters can be installed into the filter wheels since the emission filter slots 7 and 8 are always reserved for luminometric measurements Transmittance 30 100 half bandwidth 6 50 nm depending on the wavelength Filters included in the instrument Excitation filters 355 nm and 485 nm Emission filters 460 nm and 538 nm Filters for luminometric Filters can be used L
61. fault symptom or condition Give all the necessary information to the distributor who will fill out and forward the Feedback Form to the technical service department Check Section 8 4 Shipping the instrument or items You will find instructions on how to proceed before shipping the instrument for service Check that any necessary decontamination procedure has been carried out before packing See Sections 8 2 Decontamination procedure and 8 3 Certificate on Decontamination Ensure that the Certificate of Decontamination is sent with the instrument The Thermo Fisher Scientific technical service department will keep you up to date with the progress of service and provide you with any further details you might need for example on maintenance serviceability troubleshooting and replacement 8 2 Decontamination procedure Decontamination should be performed in accordance with normal laboratory procedures Any decontamination instructions provided with the reagents used should be followed A decontamination procedure is only recommended when infectious substances have been in direct contact with any part s of the instrument Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 81 If there is any risk of contamination with biohazardous material the procedure recommended below or some other corresponding decontamination procedure must be performed We strongly recommend that the complete decontamination procedure is performed
62. g 5 6 2 3 2 Replace the dispensing tip Fig 5 6 2 2 connected with a small piece of silicone tubing in the dispensing tube 3 Replace the dispensing head tube Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 69 5 6 3 Replacing the dispenser syringe Aspirate and complete dispensing tube assemblies Plunger lock screw Plunger holder arm Dispenser syringe 1 0 ml and plunger Fig 5 6 3 Replacing the dispenser syringe 1 2 3 4 1 Remove the liquid from the dispenser syringe Fig 5 6 3 4 and from the tubings 2 Switch off the power from the instrument Push the plunger manually into the upper position 4 Loosen the plunger lock screw Fig 5 6 3 2 approximately three full turns 5 Pull the plunger holder arm Fig 5 6 3 3 firmly down 6 Unscrew the syringe from the valve 7 Tofit the new dispenser syringe screw the syringe into the valve pull the syringe plunger down to the plunger holder arm and screw it into place Make sure the plunger lock screw is securely tightened 70 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 6 Troubleshooting Guide Note DO NOT use the instrument if it appears that it does not function properly 6 1 Troubleshooting Problem Cause Action Instrument does not No power connection Check the power cable turn on correctly No connection between Incorrect software type Check that the software is instrument and PC installed for
63. g screw Fig 4 8a 2 DO NOT touch the filter surfaces 8 Select the filter wheel into which you want to install a filter Normally the filters are factory installed in ascending wavelength order starting from position 1 Select the first free filter slot Emission filter wheel Excitation filter wheel 1 Fitting screw holding the excitation filter wheel in place 2 Fitting screw holding the emission filter wheel in place Fig 4 8a Filter wheels in the optical unit 9 Undo the fitting screws Fig 4 8b 1 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 57 10 Remove the spring wheel Fig 4 8b 2 11 Remove the spacer collar Fig 4 8b 3 12 Remove the dummy filter or previously installed filter used Fig 4 8b 4 Install a new filter so that the small arrow on the filter rim points away from the filter wheel The arrow shows the direction of the light flow 13 Fit the spacer collar spring wheel and the fitting screws Fit the filter wheel back into place Fitting screw s Spring wheel Spacer collar RR U N Dummy filter or previous filter used Fig 4 8b Filters in a filter wheel 14 Replace the optical unit connect the flat cable and lamp cable Lower the measurement chamber and replace the instrument cover 15 Validate the changed filter pairs refer to the instructions in Section 4 3 1 Validation and the Ascent Software User s Guide 58 Fluoroskan Ascent FL amp Fluoroskan Asc
64. g the instrument to Thermo Fisher Scientific Oy for example for repair If for any reason the instrument is shipped back to Thermo Fisher Scientific Oy it must be accompanied by a dated and signed Certificate of Decontamination which must be attached to the outside of the package containing the instrument See Section 8 2 Decontamination procedure Failure to confirm decontamination will incur additional labor charges or at worst the items will be returned for proper cleaning Before returning any instrument s or items ensure that they are fully decontaminated Confirm A or B status see below Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 83 Certificate of Decontamination Name Address Tel Fax Instrument Serial no A confirm that the returned items have not been contaminated by body fluids toxic carcinogenic or radioactive materials or any other hazardous materials B confirm that the returned items have been decontaminated and can be handled without exposing the personnel to health hazards Materials used in the unit Chemicals Biological e Radioactive Specific information about contaminants Decontamination procedure Date and place Signature Name block capitals The signature of a Radiation Safety Officer is also required when the unit has been used with radioactive materials This unit is certified by the undersigned to be free of radioactive contamination D
65. he User Manual Caution DO NOT touch or loosen any screws or parts other than those specifically designated in the instructions Doing so might cause misalignment and will void the instrument warranty Caution The Fluoroskan Ascent FL or Fluoroskan Ascent weighs approximately 21 kg 46 Ibs without dispensers and should be lifted with care It is recommended that two persons lift the instrument together taking the proper precautions to avoid injury Retain the original packing materials and shipping carton for future transportation Also retain all the documentation provided with the instrument If you relocate your instrument or ship it for service remember to 1 Empty the dispenser s and remove the tube assembly Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 19 2 Remove any loose items from the plate carrier for example adapters plates and priming vessels 3 Remove the power cable as well as the serial cable 4 Replace the transportation lock See Section 8 4 Shipping the instrument or items for further information Intended lifting area Fig 3 1 Fluoroskan Ascent FL Fluoroskan Ascent 3 1 2 Checking delivery for completeness Check the enclosed packing list against order If any parts are missing contact your local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy 3 1 3 Checking for damage during transport Visually inspect the transport package the instrument
66. he cover plate Fig 4 6c 1 from the opposite position of the optical unit under the measurement chamber 52 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 1 Six screws three screws on each side of the instrument 2 Handle 3 Flat cable 4 Long finger nuts 3 pieces Fig 4 6b Removing the optical unit 1 Cover plate 2 Short finger nuts 3 pieces 3 Gas spring Fig 4 6c Cover plate Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 53 10 11 12 13 14 15 16 4 7 Lower the measurement chamber back into the down position from the handle Unplug the flat cable Fig 4 6b 3 and the lamp cable located beside the flat cable from the optical unit DO NOT remove any other parts from the optical unit Undo the three long finger nuts Fig 4 6b 4 and remove the optical unit Lift the measurement chamber into the upper position from the handle Place the optical unit into the position where you removed the cover plate Fig 4 6c 1 Fix it with the long finger nuts and plug in the cable connectors the flat cable and the lamp cable No adjustments are needed Lower the measurement chamber back into the down position Place the cover plate instead of the removed optical unit and fix it with the short finger nuts Fit the six screws back to hold the measurement chamber and replace the instrument cover The unfitted cover may increase stray light and harm the measurement
67. ht Fig 2 4 Principle of the optical system 2 5 Control switches The control switch box FL only Fig 2 3 7 contains three rocker switches for priming and emptying dispenser tubings one rocker switch for driving the plate carrier in or out and a sensor Fig 2 3 8 to monitor that the dispenser cover is in the closed position in luminometric measurements The priming control switches are only functional when the plate carrier is located outside the instrument 16 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 2 6 Incubator The incubator contains two heating element plates in the measurement chamber one heating element plate under the microplate and another above it The incubator only heats but does not cool 2 7 Excitation beam selector The excitation beam selector makes it possible to select the optimal beam to measure different sizes of wells The normal beam diameter 3 mm is recommended for 96 well plates and larger wells The small beam diameter 1 5 mm is recommended for example for 384 well plates The beam selector is situated in the optics module Fig 2 3 9 A manually operated knob changes the diameter of the light beam and the instrument automatically detects the position of the knob 2 8 Dispensers The optional dispensers 1 to 3 dispensers from left to right are located inside the instrument housing under the dispenser cover as seen in Fig 2 3 The dispensers consist of modular digital p
68. ilter order also specify the emission filter to be used Excitation filter Emission filter Fluorochrome fluorescent dye nm nm Specify the wavelength to the nearest 5 nm Provide the excitation and emission spectra of the Fluorochrome if possible Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 121 Filter Order Form Cont Only emission filter For single filter order also specify the emission filter to be used Excitation filter Emission filter Fluorochrome fluorescent dye nm nm HE Specify the wavelength to the nearest 5 nm Provide the excitation and emission spectra of the Fluorochrome if possible List of standard fluorometric filters Excitation 320 nm 355 nm 390 nm 430 nm 440 nm 444 nm 485 nm 530 nm 544 nm 578 nm 584 nm 646 nm Emission 405 nm 440 nm 460 nm 485 nm 510 nm 518 nm 520 nm 527 nm 538 nm 555 nm 590 nm 604 nm 612 nm 620 nm 678 nm 680 nm Thermo Fisher Scientific contact information e mail to your sales representative or fax to 358 9 3291 0580 122 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Appendix C Addresses For the latest information on products and services visit our websites at http www thermo com Manufactured for Distributed by Thermo Fisher Scientific Oy Ratastie 2 P O Box 100 Fl 01621 Vantaa Finland Tel 358 9 329 100 Fax 358 9 3291 0415 www thermo com Fluoroskan Ascent F
69. ith 96 pure ethanol and afterwards with a lint free cloth or a lens tissue DO NOT use any other liquids to clean the optical unit A Avoid any harsh treatment 3 Replace all the removed parts and reconnect the instrument to the mains power supply 5 2 1 Visual filter check The useful life of a filter depends on environmental factors such as dust humidity and temperature Filters have a one year warranty Carry out the visual check in the following way Visually check the filter s by holding it them against an even light source If the color of the filter is even then the filter is suitable for use On the other hand if the filter appears to be mottled or discolored discard the filter since it is either damaged or defective The best alternative is to measure the filters with a spectrophotometer 62 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 5 3 Cleaning the plate carrier 1 Plate carrier 2 Reference chip protection glass 3 A1 corner of the microplate Fig 5 3 Plate carrier 1 Clean the plate carrier Fig 5 3 with a cloth dampened with distilled water In case of any spills of infectious agents clean the plate carrier with disposable towels dampened with a disinfectant solution containing 2 glutaraldehyde Always use disposable gloves Place the disposable towels and gloves in a biohazardous waste container 2 Clean the reference chip protection glass with a cloth dampened with 96 p
70. led Microlite 2 1x12 Strip Microlite 1 1x12 Strip assembled Microlite 1 1x12 Strip Microlite 2 1x12 Strip assembled Microlite 2 1x12 Strip White Strip 1x8 assembled UB White Strip 1x12 assembled UB White Breakable Strip 1x8 Assembled UB Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Bottom Flat Flat Flat Flat Flat U U RE RE RE Bottom Flat Flat Flat Flat Flat Flat Flat Flat Flat Flat Flat Qty 50 box 50 box 50 box 50 box 50 box 50 box 50 box 50 box 50 box 40 box Qty 100 box 320 box 100 box 320 box 100 box 320 box 100 box 320 box 50 box 50 box 50 box 93 Thermo Scientific Microtiter Plates Cont Cat no Description Bottom Qty Microtiter Plates for Fluorescence 7605 Microfluor 1 Black Plate Flat 50 box 7005 Microfluor 1 Black Plate U 50 box 7805 Microfluor 2 Black Plate Flat 50 box 7205 Microfluor 2 Black Plate U 50 box 7705 Microfluor 1 White Plate Flat 50 box 6905 Microfluor 1 White Plate U 50 box 7905 Microfluor 2 White Plate Flat 50 box 7105 Microfluor 2 White Plate U 50 box 9502867 Black 96 Well Plate UB Flat 50 box 95029120 Black 96 Well Plate EB Flat 50 box 95029840 Black 96 Well Plate Sterile Flat 40 box Cat no Description Bottom Qty Microtiter Plates for Fluorescence Strips and Assemblies 95029450 Black Strip 1x8 assembled UB Flat 50 box 95029490 Black Strip 1x8 assembled EB Flat 50 box 95029470 Black Strip 1x12 assembled UB Flat 50 box 9502950
71. luoroskan Ascent FL amp Fluoroskan Ascent User Manual 37 Reading 1 to 48 well plates as well as bottom reading can cause increase in background signal and effect sensitivity in luminometric measurements because the light shield cannot be used and most of these plates are transparent which enables light leakage from one well to another A measurement function has several phases 1 The anode voltage of the PMT is set according to the selected value of the software The excitation filter wheel is driven in between the two filter positions and the excitation light is blocked even when the light is off The emission filter slot 8 block is held in place until the plate door is closed The empty position selected filter is then driven to the measurement positions The emission filter slot 8 is also used for the blanking procedure to compensate for the possible PMT drift In the blanking procedure the instrument reads the PMT dark signal If the bottom value is drifting the results are compensated To obtain the most accurate results the measurement time of the dark must be as long as the measurement time of the sample The measurement time of the dark is divided into two parts The first half is measured before the sample measurement and the second half after the sample measurement The time used for the instrument blanking procedure can be selected from Ascent Software refer to the Overview part of the Ascent Software User s Gui
72. nditions THIS WARRANTY IS IN LIEU OF ALL OTHER EXPRESSED OR IMPLIED WARRANTIES INCLUDING BUT NOT LIMITED TO ANY IMPLIED WARRANTIES OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE The seller is not liable for any loss or damage arising out of or in connection with the use of the product or other indirect damages These warranty terms and conditions can be obtained from your local Thermo Fisher Scientific dealer This card acts as a warranty certificate Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 107 13 1 Warranty limitations The following items are not included in the warranty e Consumables e Software programs e Lamps 108 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 14 Specifications Thermo Fisher Scientific reserves the right to change any specifications without prior notice as part of our continuous product development program 14 1 General specifications Weight Basic unit 21 kg 46 Ibs 3 optional dispensers add 3 5 kg to the weight Overall dimensions 420 mm 16 5 in W x 420 mm 16 5 in D x 340 mm 13 4 in H options included Operating conditions indoor use 10 C 40 C RH 90 max Tested according to IEC 60068 2 1 test Ab Cold IEC 60068 2 2 test Bb Dry heat IEC 60068 2 3 test Ca Damp heat Transportation conditions 40 C 70 C packed in transportation packaging Tested according to IEC 60068 2 1 t
73. ngths Fluoroskan Ascent FL 360 670 nm optional 360 800 nm Fluoroskan Ascent 360 800 nm Fluorometric sensitivity 2 fmol Fluorescein in a black 96 well plate normal beam Fluorometric dynamic range gt 6 decades Fluorescein in a black Thermo Scientific 96 well strip plate Luminometric spectral range 270 670 nm Luminometric measurement range Up to approx 5000 Relative Light Units RLU Luminometric sensitivity in FL only 40 amol ATP well using flash reaction white 384 well plate Luminometric dynamic range in FL only gt 9 decades over whole gain setting area Shaker Orbital method speed 60 1200 rpm 1 50 mm Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Continued 113 Performance specifications Cont Incubator Heaters Warm up time from RT 25 C to 37 C 15 min Liquid in the well Ambient temperature 23 C covered 96 well plate 200 ul well Temperature accuracy Mean temperature of the wells 19C Uniformity lt 19C Warm up speed 1 h from 23 C to 37 C on an average 90 of the set value and the ambient temperature Dispensers Dispensing volume 1 1000 pl in 1 ul increments Accuracy 3 ul avg for volumes 5 ul and above Precision 5 19 ul lt 5 20 1000 ul lt 2 Minimum dispensing speed 25 s 96 well plate 5 ul well The dispenser specifications are given for dis
74. nical service 6 2 Error messages Error messages issued by the instrument are listed in the Reference part of the Ascent Software User s Guide in the chapter on warning and error messages 74 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 7 Frequently asked questions FAQ about the Fluoroskan Ascent FL and Fluoroskan Ascent Q Can Fluoroskan Ascent be upgraded for luminometric applications A It is not possible to add the luminometric option to the Fluoroskan Ascent Only the Fluoroskan Ascent FL can measure both fluorescence and luminescence Q Is the wavelength marked on each Fluoroskan Ascent FL and Fluoroskan Ascent filter A Wavelength as well as date information is printed on each Fluoroskan filter frame On some filters the half bandwidth HBW is also printed Q Are Petri dish adapters standard accessories that come with the Fluoroskan Ascent FL and Fluoroskan Ascent A Petri dish adapters are not standard accessories that come with the reader and thus they should be ordered separately Q When is the adapter for low 384 well plates needed A The adapter for low 384 well plates is needed when you dispense into low 384 well plates Fluoroskan Ascent FL and Fluoroskan Ascent If the plate height is less than 13 5 mm an adapter must be used to raise the plate In luminometry it is also needed for the measurement of low 384 well plates Fluoroskan Ascent FL Q What is the wi
75. nn nana rn 38 45 COTO PTUNCHONS ii ida 39 4 5 1 Orbital SHAKING vincia s 39 4 5 2 Iheubator sarte hr 42 4 5 3 DISPENSEls cidad 42 4 5 4 Dispenser head height adjustment rnrnrrnnnnnnnnnnnnrnrrnnnnnnnnnnnnn 45 4 5 5 Chemical resistance of the dispenser rruuarvvvrvrrvrrnvrvrvrnnrnnn 47 4 5 6 Excitation light beam selector sssersere renere erne ner 50 4 5 7 Area measurements ccccconononnoncnnnnnnnnnnnnnnnnnnnnnn nan nnnnnn nn nr rn nnnnnnnn nes 51 4 6 Changing the measurement direction oooocccnncccnncnancnnnnnaninnnnnnnnnnn 51 4 7 Installing or removing the light shield rerrnnnnnnnnnrnvnnnnnnnnnnnnnnnnrenn 54 4 8 Installing the filterS o occcnnnnncnnnnicnncnanonnnnnanannnncnnnn cnn rrnna nn nr rana rnnrrnnnnn 56 4 9 SHUTHOWN inicia dd at 59 5 Maintenance iia tease 61 5 1 Routine cleaning of the instruMeNt oooccccnnncccccnanoncnnnanoncnnnnnnnanoninnos 61 5 2 Cleaning the optical system esserne renerne renerne 62 5 2 1 Visual filter Check oil eee aner 62 5 3 Cleaning the plate carrier m rrrruunssrvrravvrrrrrrsrrrerrrsnnvrrsrennvnssvennnnne 63 5 4 Replacing the halogen laMP cocccccnnnnnnnoncnnninnnonanannncnnnnnn nan nnnnnn nn 64 5 5 Routine maintenance of optional dispensers and main lens 65 5 5 1 Basic Maintenance asrnane a aaea an 65 5 5 2 Extended maintenance mrnverrvnnvnannnnnnnvarnnnnnnarnnnnnrnrnnnnnnnennnenn 65 5 5 2 1 Weak detergent or 10 bleach rarnrrnrnrnvvnn
76. nnnnnnnn 107 14 Specifications a nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnannnnr 109 EOS 115 Appendix A Brief User s Guide nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnen 119 Appendix B Filter Order ForM ooooconcccoccconnconaconananenanenanenanenane 121 Appendix C Addresses annnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 123 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 3 Contents Symbols and Markings a a a nnnnnnvnnnnnnnnnnvnnnnvnnnnnnnnvnnnnnnvnvnnnnnnvnvnnen 7 1 Introduction ccoo cciccin nic aaa aa or ene 9 1 1 intended Use ian as 10 1 2 Method descriptions 2222 gar rari A pa aaa nn 11 1 2 1 Fluorometric measurement principle ooccoccnnonnconccnnnnnnannnnano 11 1 2 2 Luminometric measurement principle Fluoroskan Ascent FL 11 2 Instrument Layout nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnner 13 2 1 ErontVIEN ua SE 13 2 2 REAL VIEW an pb ee LESA 14 23 internal View seder dei eh ete aes eee eee ee 15 2 4 Optical System cin eee 16 25 Control switthes uu unatdmust gun dmAnsdte 16 2 6 INCGUD ator sess aches Ae aos aceite in Ea 17 2 7 Excitation beam selector rurarannnnrannnnnnnnnnnnrnrrannnnnennnennnnrannnnnennnnnn 17 2 8 Dispensers aiia hice acid 17 29 Plate carrier o O 18 3 Installati nM 00 0 ici iia 19 3 1 Upon delivery toria is 19 3 1 1 Unpacking urere ts rta dd Dee ste da de da 19 3 1 2 Checking delivery for COMpletenesSS no
77. nvnvrennnnnvvnnersvennnrsrenverner 65 5 5 2 2 Weak acid and base in SequenNCe ooooncccccccnnnnnonnncnnncnnnnnnnnnnnnnnnnnnnnns 66 5 5 2 3 Cleaning the main lens m rveurrvvrrvrvrvvrrrvvennrsvennsrnvrnnsrrvvnnssrsrenssnnen 66 5 6 Periodic maintenance sisirin atr i ieir biaa 66 5 6 1 Replacing the dispenser tUDINYQS cccooconococcccnininonananannnnccnnnnnnns 67 5 6 2 Replacing the dispensing tip ooccccccnnnccononononnnanonananananannncnnnnnnns 69 5 6 3 Replacing the dispenser SYriNGe cccceeeeeeeeeeeteeteeeeeeeees 70 6 Troubleshooting Guide a anannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnner 71 6 1 Troubleshooting 2 2 ios e 71 62 Error messages ideea eh Ratio teh Pde tegen bd 74 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 5 7 Frequently asked questions FAQ about the Fluoroskan Ascent FL and Fluoroskan Ascent nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 75 8 Instrument ServiCe a annnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnunnnnnnnnnnnnnnn 81 8 1 Service request Protocol mmmmmnsvrrerrensrerrrnsrrerrsnarreessnsrreessnsrreessnnnn 81 8 2 Decontamination procedure occccnncnnoconcnnnnnnnnnnnononccnnnnnnnnnnnn nn ncnnnnnnns 81 8 3 Certificate of decontamination cesser rr 83 8 4 Shipping the instrument or items rrrnnnannvrvnnannvvvnnrnnrvennrsnsvvvnrnnn 85 8 5 Service contracts eshare aa aAA EA AA a a 85 9 Disposal of the Instrument and Materials 87 9 1 Disposal of the instrument
78. o Fisher Scientific representative 9 2 Disposal of materials Refer to local regulations for the disposal of infectious material The samples can be potentially infectious Dispose of all used F microplates PCR tubes disposable gloves syringes disposable tips etc as biohazardous waste Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 87 88 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 10 Ordering Information 10 1 Product code numbers Microplate Fluorometers Cat no Instrument 5210470 Fluoroskan Ascent 5210480 Fluoroskan Ascent with dispenser 5210450 Fluoroskan Ascent FL 5210460 Fluoroskan Ascent FL with dispenser Fluorometric Filters Cat no Item HBW 1423202 Excitation filter 320 nm 40 nm 1423552 Excitation filter 355 nm 38 nm 1423902 Excitation filter 390 nm 20 nm 142430210 Excitation filter 430 nm 10 nm 142440210 Excitation filter 440 nm 10 nm 1424442 Excitation filter 444 nm 12 nm 1424852 Excitation filter 485 nm 14 nm 1425302 Excitation filter 530 nm 10 nm 1425442 Excitation filter 544 nm 15 nm 142578206 Excitation filter 578 nm 7 nm 1425842 Excitation filter 584 nm 16 nm 1426462 Excitation filter 646 nm 10 nm Specified to be used as either excitation or emission filter Fluoroskan Ascent FL Fluoroskan Ascent User Manual Fluorometric Filters Cont Cat no Item HBW 1424052 Emission filter 405 nm 50 nm 1424602 Emission filter 460 nm 24 nm 14248
79. occccnnnocccnnononnnnnanann nono 20 3 1 3 Checking for damage during tranSpoOlt cccconoccccnononnnnnanannnnnnna 20 3 1 4 Environmental requirements and noise 21 3 1 5 THINGS Ol coca tio ska 21 3 1 6 Technical prerequisites ooooccccconononoccnnnonnnnnnnannncnnn nerne nerne 22 3 2 Releasing the transportation lock rrnnnnnnnnnnnnrvnnnnnvnnnnnnnnnrennnnrennnnnn 22 3 3 Power and computer CONNECTIONS cccccccccconononnnccnnnnnnnnnnancnnccnnnnnnns 23 3 4 Installation of dispensers rrrnnannvvvvnnannvvvnnannvrvnnsnnnvvenssnsvrnnsensvveneen 25 3 5 Installation of the drop plate uwssrrveravavvrvrravavrvrnrnnrvrnrsnsrvnnssnsrveneen 26 3 6 Plate ad pt ers ii eee se 27 3 7 Adjusting the plate carrier rrrrsnrornnnnnnnnnnnrnrrnnnnnrentnennnnrrnnnnnennnnnn 29 3 8 Operational check rrrrnnnnvnnnnnnnnnannnnnnvnnnnnnrnernnnrnnnnennesnnnnnsrnennnennnnnnn 30 4 Operation LL LEEDS aae AE Dra ANEREN ES 31 ALT Switching OMe tiori id 31 4 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 4 2 Loading the microplate oooonnnnnocccncnnninnnnnnnonnnccnnnnnnnnnnn no nncnnnnnnnnnnnn nes 32 4 3 Fluorometric measurement rnrennnnnnnnnnnnnnrnnnnnnrnrnrnnnnrnnnnnnentnnnnnnnnnn 33 4 3 1 Validation inician aiii 34 4 3 2 SC ii E A tds 35 4 3 3 Dynamic series test Solutions ssseersee nerne ennen 36 4 4 Luminometric measurement scenerne nerne 37 4 4 1 Luminometric SCALING ooooonmnnconccnnonnnancnnnccnnnncnnn
80. olume of the first well equal to that of the others We recommend you use the tip priming feature when the dispensing volumes are small for example 5 20 ul 1 A1 corner of the microplate Fig 4 5 3b Tip priming vessel 1 is a piece ofa breakable strip 44 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual The recommended tip priming vessel consists of four wells of a breakable 96 well plate strip Fig 4 5 3b There is a holder for the tip priming vessel in the right rear corner of the plate carrier The four well piece of a strip should be exchanged after about 300 tip primings You may need to adjust the dispenser speed The default setting is for a water based liquid You can find the adjustments and selections in Ascent Software When dispensing is started the liquid volume in the well should be less than half of the total well volume for example the volume should be less than 200 ul in a typical 96 well plate 4 5 4 Dispenser head height adjustment Plate height 13 5 15 0 mm Plate height 15 1 18 0 mm Plate height 18 1 21 0 mm e g 96 well plate e g 48 well plate e g 24 to 6 well plates No adjustment collars One adjustment collar Two adjustment collars Fig 4 5 4a Dispenser head height The correct dispenser head height Fig 4 5 4a is very important to avoi
81. once Dust or dirt in the wells Keep the plates protected before use Foaming in the sample Use a lower dispensing or shaking speed Use correct pipetting techniques reverse lllogical results Plate inserted with incorrect orientation Rotate the plate with the A1 well facing towards the upper left corner Too low or variable volume from dispenser Inadequate priming Reprime the dispenser Use tip priming during the session Incorrect dispensing height Check that the dispenser heads are positioned at the correct height Continued 72 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual Problem Cause Action Cont Too low or variable volume from dispenser Incorrect dispensing height Check that the dispenser head positions in the instrument and in the software settings are identical Use plate adapters with low 384 well plates Loose syringe plunger or tubings Check all the connections in the dispenser Aspiration tube not in the liquid Ensure that there is a sufficient volume of liquid in the vial Ensure that the aspiration tube end weight is at the bottom of the vial and attach it firmly if necessary Liquid droplets outside the wells after dispensing Incorrect plate template used Check the plate type used in the software Too high total volume used Use smaller volumes Lid
82. oroskan Ascent Cont Q Does the Fluoroskan Ascent have different beams for different plate types A The Fluoroskan Ascent FL and Fluoroskan Ascent have two beam diameter settings The normal beam with a diameter of 3 mm is generally recommended for 96 well plates and larger wells The small beam with a diameter of 1 5 mm is recommended for 384 well plates Q Is it possible to measure 96 well PCR plates with the Fluoroskan Ascent FL and Fluoroskan Ascent A Yes and you can use the Fluoroskan Ascent FL and Fluoroskan Ascent for measuring both PCR plates and 0 2 ml PCR tubes but you need a special adapter This adapter is suitable for for example Hybaid Omniplates Robbins Cycloplates and Axygen PCR microplates or tubes from Greiner or Sarstedt Q Do I have to make any modifications to the instrument if I want to measure fluorescence and luminescence from the same well A No you must only select separate measurement steps in the software which means that you can use both measurement technologies in one session for measuring the same wells Q What kind of plates are best to use with the Fluoroskan Ascent FL luminometric measurements when you have to reduce the crosstalk as much as possible and minimize the background A There are clearly two options You should use either white Microlite 1 plates Cat no 7571 or white universal binding plates Cat no 9502887 78 Fluoroskan Ascent FL amp Fluoroskan Ascent User M
83. oskan Ascent User Manual 65 3 Flush the dispenser a minimum of 10 cycles with distilled water 5 5 2 2 Weak acid and base in sequence Remove the dispensing heads from the dispensing positions and DO NOT let any cleaning fluids enter the measurement chamber Use external containers 1 Prime the dispensers with 0 1 M NaOH and leave the solution in the dispensers for 10 minutes with the syringes full Note DO NOT spill any 0 1 M NaOH onto any instrument surfaces to avoid damage of the instrument If needed use suitable protection covering 2 Flush the dispensers with distilled water 3 Prime the dispensers with 0 1 M HCI and leave the solution in the dispensers for 10 minutes with the syringes full 4 After the 10 minute period remove the aspirate tubing from the 0 1M HCI solution and remove all the fluid from the syringes and tubing into a waste container 5 Flush the dispensers a minimum of 10 cycles with distilled water 5 5 2 3 Cleaning the main lens It is recommended that you check the main lens of the optical unit weekly 1 Remove the optical unit if it is above the measurement chamber or the cover plate if the optical unit is below the measurement chamber as described in Section 4 6 Changing the measurement direction 2 If needed clean the main lens according to the instructions in Sections 5 2 Cleaning the optical system 5 6 Periodic maintenance There are three items which require periodic maintenance
84. osstalk area Decade Decontamination Disinfection Dynamic range Emission Error message Excitation Adenosine triphosphate a biological molecule that is commonly used as a reference chemical for luminometric sensitivity The framework of the instrument A compound that emits light following a chemical reaction is said to be chemiluminescent The point where the excitation and emission spectra meet Interfering signal from neighboring wells The common area for excitation and emission filters used Order of magnitude A logarithmic value that is used for presentation of dynamic range Removal or neutralization of radiologic bacteriological chemical or other contamination The destruction of pathogenic bacteria usually with an antiseptic chemical or disinfectant Dynamic range refers to the range of signals an instrument can read from the minimum to the maximum detectable For example dynamic range of seven decades means that the difference between the lowest and highest signals that can be measured is 10 The release of light from a fluorochrome when an electron falls from an excited state to a lower energy state of the molecule Indication that an error has been detected The absorption of light energy by a fluorochrome during which electrons in the fluorochrome molecule are boosted to a higher energy level Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 101 Filter valid
85. r Molecular Probes Handbook of Fluorescent Probes and Research Products 2001 Haugland R P ed Eighth edition Molecular Probes CD ROM Principles of Fluorescence Spectroscopy 1999 Lakowicz J R ed Second edition Plenum Pub Corp Fluorescent and Luminescent Probes for Biological Activity A Practical Guide to Technology for Quantitative Real Time Analysis 1999 Mason W T ed Second edition Biological Techniques Series Academic Press Bioluminescence and chemiluminescence Part B Methods in Enzymology Vol 133 1986 DeLuca M A and McElroy W D eds Academic Press Orlando 649 pages Chemiluminescence Principles and Applications in Biology and Medicine 1988 Campbell A K ed Horwood Chichester U K 608 pages Nonradioactive labeling and detection of biomolecules 1992 Kessler C ed Springer Verlag Berlin 436 pages Bioluminescence Methods and Protocols 1998 LaRossa R A ed Methods in Molecular Biology Vol 102 Humana Press Totowa New Jersey 310 pages Bioluminescence and chemiluminescence Perspectives for the 21 century 1999 Roda A Pazzagli M Kricka L J and Stanley P E eds John Wiley amp Sons Chichester U K 625 pages Bioluminescence and Chemiluminescence Part C Methods in Enzymology Vol 305 2000 Ziegler M M and Baldwin T O eds Academic Press San Diego 732 pages Bioluminescence amp Chemiluminescence 2001 Case J
86. ront view Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 13 2 2 Rear view 6 3 aq A 1 Mains power supply socket 2 Power switch ON OFF 3 Serial communication connector for the computer 4 Identification plate 5 Housing retaining screws 4 pieces 6 Cooling air outlet Fig 2 2 Fluoroskan Ascent FL Fluoroskan Ascentrear view 14 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 2 3 Internal view Light cover for the optical unit Dispensers opt Dispensing head opt Leakage tray opt Dispensing head holder opt Dummy plug Control switches in the Fluoroskan Ascent FL Cover sensor in the Fluoroskan Ascent FL o On OO Oh RK Y DY Beam selector Fig 2 3 Fluoroskan Ascent FL Fluoroskan Ascent internal view Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 15 2 4 Optical system ja gt The light source is a quartz halogen lamp The excitation filter in the excitation filter wheel The lamp reference detector kh WOW N The highly focused excitation light beam in the well to be measured The diameter of the normal beam in the sample is about 3 mm and that of the small beam about 1 5 mm 5 In the Fluoroskan Ascent FL the emission light beam is also strictly limited with a light shield to avoid crosstalk in luminometric measurements 6 The emission filter in the emission filter wheel 7 The photomultiplier tube detects the emission lig
87. ry twelve months on a contract basis by the manufacturer s trained service engineers This will ensure that the product is properly maintained and gives trouble free service Contact authorized technical service for further details Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 85 86 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 9 Disposal of the Instrument and Materials 9 1 Disposal of the instrument For the disposal of the instrument do the following Decontaminate the instrument prior to disposal See Sections 8 2 Decontamination procedure and 8 3 Certificate of Decontamination Dispose of the instrument according to the legislation stipulated by the local authorities concerning take back of electronic equipment and waste The proposals for the procedures vary by country Thermo Fisher Scientific has contracted with one or more recycling disposal companies in each EU Member State European Country and this product should be disposed of or recycled through them Further information on Thermo Fisher Scientific s compliance with these Directives the recyclers in your country and information on Thermo Scientific products which may assist the detection of substances subject to the RoHS Directive are available at www thermo com WEEERoHS Regarding the original packaging and packing materials use the recycling operators known to you For further information contact your local Therm
88. s feature is important to obtain optimal sensitivity The light path from the measurement well to the first lens is protected by a light shield Fig 4 7a and Fig 4 7b The light shield is required with 96 and 384 well plates to avoid crosstalk in luminometric measurements The light shield is not necessary but does not interfere in fluorometric measurements When plates are higher than 15 mm the light shield must be removed If the 384 well plate has a height of less than 15 mm then an adapter must be used below the plate to raise the plate to the proper height This adapter for low 384 well plates Fig 3 6 4 is included in the Fluoroskan Ascent FL instrument and can also be ordered separately Caution A luminometric measurement without a light shield may cause extra crosstalk Caution The light shield must be removed with higher plates Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 11 12 Note The emission filter slots 7 and 8 are reserved only for luminometric measurements Filter slot 7 is empty for measurement and filter slot 8 is blocked to enable measurement of the PMT photomultiplier tube dark current Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 2 Instrument Layout 2 1 Front view Instrument housing 1 2 Dispenser and optics cover 3 Instrument chassis 4 Measurement chamber door 5 Power busy and error indicator Fig 2 1 Fluoroskan Ascent FL Fluoroskan Ascent f
89. seen through the hole in the light cover of the optical unit Fig 4 5 6a When the optical unit is below the plate the measurement chamber must be lifted up to reach the selection knob see Section 4 6 Changing the measurement direction points 1 6 The position of the selection knob in the optical unit is shown in Fig 4 5 6b Filter wheel screw lr EXCITATION BEAM Normal beam 3 mm om Small beam 1 5 mm mm Push Normal beam Pull Small beam a pointer shows which beam is on Fig 4 5 6b Beam selection when carrying out bottom reading 50 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual To select the normal beam push the button firmly To select the small beam pull the button firmly Caution DO NOT leave the button in a middle position The pointer on the knob also shows the position of the selection 4 5 7 Area measurements In area measurements Fig 4 5 7 when there is more than one measurement point in a well we recommend you use the normal beam The distance between two measurement positions is 1 5 mm When you use the normal beam 3 mm the measurement areas overlap and the whole area is measured If you use the small beam 1 5 mm the small areas between the measurement points are not measured at all L A LAA AAA A Y NA NA NA Y NA LLLL LL LL LLL OLA v Area measurement with the normal Area measurement with the small beam The whole ar
90. situation is shown but transmission is on a logarithmic scale Each filter deeply blocks the other providing a pure emission signal 44 102 Excitation Emission 10 Crossover point 10 Crosstalk area 1081 450 500 550 600 nm Fig 10 3c Excitation and emission spectra on a logarithmic scale This example shows that the center wavelengths of the filters are not necessarily the same as those of the fluorescent material s excitation and emission spectra In this way the final result is better If the crossover point is located too high the background signal is high and the measurement sensitivity poor 98 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 10 4 Fluorochromes and corresponding filter pairs Fluorochrome Ex Em Filters Cat no Notes Acridine Orange 485 538 1424852 1425382 DNA alamarBlue 544 590 1425442 1425902 AlexaFluor 488 485 518 or 538 1424852 1425182 or 1425382 6 Aminoquinoline 355 544 1423552 1425442 AFC 390 510 1423902 1425102 AttoPhos 444 555 1424442 1425552 Calcein 485 538 1424852 1425382 or 518 or 1425182 Chlorophyll 444 680 1424442 1426802 Cy3 544 590 1425442 1425902 Cy5 646 678 1426462 142678210 DAPI 355 460 1423552 1424602 DNA EGFP 485 520 1424852 1425202 or 527 or 1425272 EYFP 485 527 1424852 1425272 FAM 485 518 1424852 1425182 FITC 485 538 or 518 142485
91. th band undergo an atomic change and instantaneously emit light of some longer wavelength band The most common fluorescent materials have well characterized excitation and emission spectra Fig 10 3a shows the excitation and emission spectra for a fluorescent material The excitation and emission bands are each fairly broad with a half bandwidth of approximately 40 nm and the wavelength difference Stokes shift between the excitation and emission maximums is small just 32 nm Wavelehght difference Excitation Emission Half bandwidth Relative intensity o 040 500 50 OO nm Fig 10 3a Excitation and emission spectra The requirement for any fluorometric system is to excite the fluorescent material with monochromatic light and permit observation of the subsequent emission Because the intensity of stray non absorbed excitated light is usually many tens of thousands of times greater than that of the emission light some type of spectral separation is necessary to produce the contrast required for observation Fig 10 3b shows the excitation and emission spectra for an optimized pair of excitation and emission filters Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 97 Emission Excitation o p If bandwidth Relative intensity 0 Fig 10 3b An optimized pair of excitation and emission filters In Fig 10 3c the same spectral
92. the brass tube lock 2 The red adjustment collars Fig 4 5 4b 2 should be moved from one side of the fixed collar to the other to select the correct dispenser head height 3 Fitthe dispenser head tube back into place 46 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 4 5 5 Chemical resistance of the dispenser The following table Table 4 5 5 is intended to provide guidelines for compatibility with materials used in the fluid path of the dispensers Compatibility information is based on charts provided by the material manufacturer It is recommended that each laboratory determines compatibility for their respective applications Caution Failure to determine compatibility of chemicals used in individual applications with the dispenser may result in damage to the dispenser and or test results Plastic materials used in dispensers Polysulfone Cross Flow Manifold Assembly in the aspirate syringe Teflon PTFE TFE FEP tubing valve plug and seal Kel F valve body Polypropylene fittings for tubing and dispensing tip Note Also take into account the chemical resistance of microplates Classification in the table No data available No effect excellent Minor effect good Moderate effect fair Severe effect not recommended WN O Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 47 Table 4 5 5 Compatibility chart of materials suitable with the dispenser
93. tilled water at RT 114 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 15 Index A ADICON VO Ervin dd 33 ADDI VIGtlONS i iisctisiseicccsduits active reoodudjuivae ladiaduaavvsbeweisdouQaivevvdaneardetinavaaveds odes 101 ACGESSONES uret rd 19 20 25 64 75 91 95 Adapter weenie ccs var 11 26 28 29 46 74 75 78 79 91 Application rerarararnrnrnrnrnnnnnnnnnnnannnnnnnnnnnnnnnnns 7 8 9 10 39 47 50 75 96 Autocalibration cccccccsssseeececeseensseseeeeeseeananseeeeeeeees 30 33 34 76 102 103 B Beam selecto riada ral dada ts 17 Bla A EE 30 38 Blake dd ii 35 71 119 Bottom reading cccccncncncnenenenenenenenenencncnnnnnnnononononenenenos 11 26 38 50 72 76 Brass tube holder irrin dani ii eee aa heisen dni 68 69 Brief Users G ud iii diabetes 119 C Chassis iia cda abia et 13 52 56 101 Chemical resistance mrrrrrmrrrvrvvvvvvvvvrrrvrvrrrrvrrerererererererererererererererererer 47 Clean optical SyStem iii A dan a 62 Consumables ia 108 Control switches ooononncccnncccnnonanana nana no no ER naar arc nn nn E REE EET nana 15 16 43 44 Control Sic siii nta eit dida da aida 9 30 31 119 Cover sensorusaskrudadan kem ia 15 COSSIO Brisa tallada dotada dido cidad aia aa diana ide 98 101 Crosstalkiiiaiiia id aa a a 9 11 16 33 37 77 78 101 D Decontamination rrnrnnnrannnnnrnernvnvnnnnnnnnssernnnnnnnnenr 61 81 83 84 85 87 101 Certificate OF rara a tirada
94. tion M The positions are optimized for a 96 well plate Fig 4 5 3a When the dispensers are not in use the dispensing heads may be stored in the dispensing head holder but the tip holes to the measurement chamber must be closed with dummy plugs Note Ensure that the tip position in the instrument corresponds to that defined in Ascent Software 42 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 1 M position Dispensing head directed in the measurement well position Dispensing into the well during the measurement step is possible with this position 2 Y position Dispensing head directed in the well next to the measurement position in the Y direction When this position is used for dispensing an extra plate movement is carried out before the measurement step causing minor time delays 3 X position Dispensing head directed in the well next to the measurement position in the X direction When this position is used for dispensing an extra plate movement is carried out before the measurement step causing minor time delays Fig 4 5 3a Dispenser tip positioning optimized for a 96 well plate Before inserting the dispensing head into a dispensing position prime the syringe and tubing to an external priming vessel The instrument has no internal priming vessel for priming the syringe and tubing You can find the priming instructions in the Ascent Software User s Guide The minimum priming volume needed is 700 ul and th
95. trument has found an error the error message is sent to the computer and the computer has not acknowledged it 4 1 Switching on When switched on the instrument performs the following initialization tests and adjustments 1 The measurement system is tested using an internal test signal then different gain steps are tested and adjusted 2 Both filter selection systems are tested and the filters in position 1 are selected 3 X and Y movements of the plate carrier are tested and the carrier is driven out Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 31 4 When all the initialization tests and adjustments have been performed successfully the indicator LED turns green and the instrument is ready to receive computer commands The recommended warm up time is 15 minutes but the instrument will perform commands immediately after the initialization period 4 2 Loading the microplate The microplate is loaded onto the instrument plate carrier for measurement The plate carrier is able to handle microplates of different sizes therefore the free space in the carrier is clearly larger than the standard 96 well plate The positioning lever in the plate carrier Fig 4 2 will automatically position the plate correctly into the upper left corner of the carrier when the plate is driven in A1 corner of the microplate Positioning lever Fig 4 2 Plate carrier 32 Fluoroskan Ascent FL amp Fluoroskan Ascent
96. ture for luminometry Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 9 The Fluoroskan Ascent FL can also be equipped with filters for luminometric applications Up to three reagent dispensers can be fitted on board making the reagent addition simple and highly accurate The ability of the instrument to dispense and measure simultaneously enables the detection of flash luminescence reactions fluorometric Ca measurements and other rapid kinetic applications For assays requiring temperature control these instruments have an on board incubator Built in orbital shaking speeds up reaction times and ensures effective mixing Robotic integration is simple and effective with the Fluoroskan Ascent FL and Fluoroskan Ascent The plate carrier allows convenient access for the robotic arm and Ascent Software is easy to integrate with robotic and HIS LIMS systems The Fluoroskan Ascent FL and Fluoroskan Ascent are also fully compatible with Thermo Scientific robotic plate handling devices expanding the measurement capacity For further information contact your local Thermo Fisher Scientific representative 1 1 Intended use The Fluoroskan Ascent is a high quality microplate fluorometer intended for laboratory research use by professional personnel The Fluoroskan Ascent is used to measure fluorescence from suitable 1 to 384 well plates mentioned in this manual lt also has incubation shaking and reagent dispensing capabilities
97. uminometric filters and measurements fluorometric emission filters are situated in the same filter wheel maximum 6 filters Plate types 1 to 384 well plates Can also be programmed for nonstandard configurations Maximum dimensions 90 mm x 134 mm x 25 mm Shaker Built in orbital shaker with adjustable speed and diameter Incubator Temperature range from RT 25 C 3 C to 45 C when the ambient temperature is 25 C The temperature is selected using Ascent Software Dispensers 1 3 optional dispensers Syringe volumes 1000 ul Dead volume 600 ul Metal free fluid path Exchangeable valve syringe tubing and tip Autoclavable tubing and tip 110 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 14 2 Safety specifications The Fluoroskan Ascent FL Fluoroskan Ascent FL fulfils the following requirements EN 61010 1 2001 EN 61010 2 101 2002 including CA US National Differences CSA C22 2 No 1010 1 M1992 The safety specifications are also met under the following environmental conditions in addition to or in excess of those stated in the operating conditions Altitude Up to 2000 m Temperature 5 C 40 C Mains supply 10 if larger than specified above fluctuations Installation category ll according to IEC 60664 1 see Note 1 overvoltage category Pollution degree 2 according to IEC 60664 1 see Note 2 Notes 1 The installation category overvoltage category
98. umps with valves syringes tubing and dispensing heads The dispensing heads have three alternative dispensing positions one of these dispenses into the well in the measurement position Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 17 2 9 Plate carrier A U N 18 A1 corner of the microplate Positioning lever Adjustable stoppers Fluorescence reference chips Waste strip holder opt 4 wells for the tip priming during the measurement session Plate carrier door Fig 2 9 Plate carrier Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 3 Installation 3 1 Upon delivery 3 1 1 Unpacking The Fluoroskan Ascent FL or Fluoroskan Ascent is packed in a specially designed shipping carton Move the unpacked instrument to its site of operation To prevent condensation the instrument should be left in its protective plastic wrapping until the ambient temperature has been reached Unpack the Fluoroskan Ascent FL or Fluoroskan Ascent instrument and accessories carefully with the arrows on the transport package pointing upwards Open the top of the package and lift the Fluoroskan Ascent FL or Fluoroskan Ascent out of the shipping carton Fig 3 1 The following notes and instructions are sent with the instrument and are immediately available when you open the package e the packing instructions e the packing list e the Warranty Certificate card e the performance test reports e t
99. ure ethanol in distilled water and afterwards with a lint free cloth or lens tissue Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 63 5 4 Replacing the halogen lamp Switch off the Fluoroskan Ascent FL Fluoroskan Ascent and disconnect the mains power supply cable Fig 5 4 1 Remove the instrument cover according to the instructions in Section 4 6 Changing the measurement direction points 1 4 Locate the lamp Fig 5 5 4 on top of the optical unit Fig 5 5 1 Open the two screws of the lamp connector Fig 5 5 2 and remove the connector Open the two screws of the lamp holder Fig 5 5 3 to release the lamp Replace the faulty lamp Fig 5 5 4 with the spare provided with the accessories Contact your local Thermo Fisher Scientific representative if a spare lamp is required Caution DO NOT touch the glass bulb with your bare fingertips Note that the lamp is not symmetrical The guide in the rim should point to the right 5 Refit the lamp connector 6 Replace both the light cover and the instrument cover 7 Reconnect the instrument to the mains power supply 1 Optical unit 2 Lamp connector 3 Lamp holder 4 Lamp Fig 5 4 Replacing the halogen lamp 64 Fluoroskan Ascent FL amp Fluoroskan Ascent User Manual 5 5 Routine maintenance of optional dispensers and main lens To obtain optimum performance and maximum useful life from the dispensers it is important that the recommen
100. ure that there are no items preventing the closure of open the plate carrier door during measurement 4 3 1 Validation 1 34 In the validation procedure a new filter pair is introduced to the autocalibration system The Fluoroskan Ascent FL Fluoroskan Ascent has a special automatic gain selection to obtain the optimal dynamic range without any adjustments The automatic gain selection optimizes the relation between the excitation and emission wavelengths and the anode voltage of the photomultiplier tube to compensate for the halogen lamp intensity spectra as well as PMT spectral response The validation procedure always uses the normal beam only You can find the filter pair validation procedure in Ascent Software The menu selection is Setup Filters The instrument reads the fluorescence values of all the reference chips selects the chip for which the value is suitable neither too low nor too high The selected value is saved in memory for the autocalibration procedure If the validation procedure does not find a suitable reference chip the program requests if the user still wants to use this filter pair If the answer is Yes the pair can be used in measurements The only difference is that the autocalibration procedure cannot form a correction factor to compensate for the drifts mentioned in the previous section Other compensations such as the excitation intensity compensation with the reference channel remain in the auto

Fluoroskan Ascent FL - User Manual

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