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KingFisher - User Manual

1. eese 46 18 Appendix D Addresses sce cio cee eere edi ret EKE KK KAK KA KAKA KA KA KA gen KA KA KA KA KA KA KA KA KA KA KA KAWA 47 19 Appendix E Supplementary Application Protocols LL EEE Ek kk kk kk kk kk nennen 48 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 2 Table of Contents Table of Contents 1 Safety Symbols and Marking iiocoacci iii 6 SZ About the User Manual seu errare tron bana k k ka k nav ea ea DDR ERA savannen VV W R n HEKE K ac anden bekken 7 3 Introduction to the KingFisher LL Ek Eka kk kk kk kk kk kk kK KAKA KAKA KA KA KA KA KAKA KA KAK 8 3 1 Intended Usera daa see 8 3 2 Principle of operation 8 3 3 Advantages of processing magnetic particles with the KingFisher esessss 9 dk Functional Description iere Tenere erret reete ren iei ka a Din arx eee ke ciega fen eua eg 10 4 1 Instrument T y oU nU MIS UE RUNS IRR OPE 10 4 1 1 gione ne 10 4 1 2 Back VIEW iiie neben n dense io MR Pe ED 11 4 1 3 Internal VIEWS A aa dusker da Iud tcs 11 4 2 KingFisher magnetic particle processor ener nennen KA KA KA A 12 4 3 Principle of the KingFisher magnetic particle processor Ek kk kk kK 13 4 4 Working with a magnetic rode 13 4 4 1 Collecting magnetic Darticles nn nr enne A KAKA KA KA KAK RA 13 4 4 2 Releasing magnetic particles oooccccconocinnncccncconcnnnnnnnnann cnn nnnn rca nena rr 13 4 4 3 Wa
2. Thermo Scientific KingFisher User Manual Rev 2 4 Cat no 1507730 Table of Contents Thermo Scientific KingFisher Cat no 5400000 User Manual Rev 2 4 June 2008 Cat no 1507730 Contents 1 Safety Symbols and Markings EK 6 2 About the User Manual rernnnvovnnnnnvvvnnnnnvnnnnnnnvnnnnnnnvnnnnnnnnvnnnnnnnvnnnnnnavnnnnnnsnnnnnnnennnnnnnsnvnnnnnnnnvennn 7 3 Introduction to the Kingbtsbher REENEN 8 4 Functional Description einer idet o Deere Kek kK KAKA KAKA KA KK KA KA KA neen KAK KA KA 10 5 Installation nan ninan xezay RR 15 6 Routine Operati On E E L 19 7 Maintenance ERROREM 23 8 Technical Specifications EL LL EEE kk kk eene kk kK KK A KA KAKA KA KAKA KA KA KA nenne A KA KK nnns 26 9 Troubleshooting Guide L Ek kk kk kk kak kk nennen KAKA KA KAKA KA KAKA KA KA KA KA KA KA KA KA KA KA KK A KA KA 29 10 Warranty Certificate echt edie iain ain dara 34 11 Ordering BI ee E EE 35 12 Glossary and Abbreviations rxrvrarrnnvnvnnnnnvnvnnrnnnnvnnnnnnnnnennnnnnvenennnnvnnsnnnneenennnneenennnnessennnnneenennn 36 13 DI A ERES 38 14 Table of Figures 2 20 x2 yan PE 41 15 Appendix A Application Protocols for KingFisher L L EEE ek kkkk ek 42 16 Appendix B Thermo Scientific KingFisher Brief User s Guide 45 17 Appendix C Thermo Scientific KingFisher Feedback Form
3. 3 We recommend that you carry out an empty run for example a demonstration to verify proper instrument operation Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 18 Chapter 6 Routine Operation 6 Routine Operation 6 1 Switching on Before you switch on the KingFisher ensure that the voltage on the rating label at the bottom left of the back panel Fig 4 3 corresponds to the local voltage 6 2 How to use the keypad and display The liquid crystal display is a 20 character by 2 row line LCD The lower line of the display can show the steps of the protocol while the processing is running Furthermore if errors occur they will be displayed on the lower row The keypad and display are shown in Fig 6 1 Thermo Labsystems KingFisher START Y STOP Fig 6 1 KingFisher keypad The keys available are two cursor keys or arrow keys direction keys to select the processing step to be run To change to the previous next protocol by scrolling up Y To change to the next protocol by scrolling down two function keys START To initiate the processing step STOP To pause terminate the processing step In short STOP paused START the instrument continues after a Pause step STOP paused STOP the processing is terminated Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 19 Chapter 6 Routine Operation 6 3 How to START e First of all slide the plat
4. Chapter 9 Troubleshooting Guide 9 4 Service request protocol If the KingFisher requires service contact your local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy 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 Your local Thermo Fisher Scientific representative or distributor will take care of sending a complaint form that is the Warranty Claim Technical Sheet to the Thermo Fisher Scientific technical service department The Warranty Claim Technical Sheet contains a more detailed description of the fault symptom or condition Give all the necessary information to the distributor who will fill out and forward the Warranty Claim Technical Sheet to the Thermo Fisher Scientific technical service department Check Section 7 4 How to pack for service You will find instructions on how to proceed before shipping the instrument for service to Thermo Fisher Scientific Oy Check that any necessary decontamination procedure has been carried out before packing See Sections 9 4 1 and 9 4 1 1 on decontamination Refer also to Section 7 3 Decontamination procedure Ensure that the Certificate of Decontamination as well as the return authorization number RGA are sent with the instrument The Thermo Fisher Scientific technical service departmen
5. 1 um 99 vessel washing buffer Ambient temperature i e no incubation START STOP two cursor keys LCD 1 8 fixed internal protocols loading through the external PC Thermo Scientific KingFisher Software can be used to create and modify protocols Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 26 Chapter 8 Technical Specifications 8 2 Safety specifications The KingFisher fulfills the following requirements EN 61010 1 1993 A2 1995 IEC 61010 1 1990 A1 1992 A2 1995 including CENELEC Common Modifications US and CA National differences Installation Category Overvoltage Category Il Pollution Degree 2 EN 61010 1 2001 Ed 2 The safety specifications are also met under the following environmental conditions in addition or in excess to those stated in the operating conditions Altitude Up to 2000 m Temperature 5 40 C Mains supply fluctuations t 10 from nominal Installation category overvoltage category Il according to IEC 60664 1 see Note 1 Pollution degree 2 according to IEC 60664 1 see Note 2 Notes 1 The installation category overvoltage category 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 II which is the category used for instruments in installations supplied from a supply comparable to public
6. 26 31 36 42 43 44 45 O O O NN 25 26 30 31 32 conraes EE Ee 25 A TERT 25 32 r q est protocol chai Pl 32 SOUTO WA A A dasa ta ada davai e A A tar 22 e SET ee A e a 7 26 27 genertal ud haram RU RR OPI Ge en 26 performance ico el nay eo KE ber rote tore de kaker eee ds 26 GOL G ayay a y ke n y ene I be en 27 Sj Te EIER ee een Kee P 18 19 21 Streptavidin coated magnetic particles rrrrrannrvvnnanavrvrnannrrerrrnarrenrsnnnrrenssnarrennenarveessnrveessnrveessnarvesennn 36 O pet 6 T BRUET CTT 41 MICA AA Attia 8 11 12 13 14 20 21 22 26 30 31 35 36 45 AN 15 16 17 25 26 Gama Gees a A A A Hn 15 oe A EE 16 17 25 Package A A a aa 15 25 Troubl shobtngau E 7 29 30 32 guide EE 30 U UripaGk a ERE en er a OR OE Urn pee Ur Rp Eie ERE 15 User rfiantial 2 xa yal y teet cnni dE ure decre ee Kya Ka aha UL aU nva der 7 15 35 46 V AT 10 11 FROMM a Ada 10 man EEE ir it decia 11 A Ad A Ee Ed A A AS da 11 lee E 16 18 19 26 28 Tele ET A E E 18 19 W A lists 16 17 18 29 Elise e 7 15 34 Welt icis Eis day ka y k A Eee A YER saa EE REA ETC TAY We W ex ED E RO Ex E Fc De FORE LAO ERE EX bd ue 16 26 44 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 40 Chapter 14 Table of Figures 14 Table of Figures Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig Fig 3 1 KingFisher magnetic particle Grocessor rn Ell
7. 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 dimensioning of the electrical insulation within the instrument Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 27 Chapter 8 Technical Specifications 8 3 In conformity with the requirements The KingFisher bears the following markings Type 700 100 240 Vac 50 60 Hz 80 VA CE mark CSA monogram The KingFisher conforms to the following requirements 2006 95 EC Low Voltage Directive 2004 108 EC Electromagnetic Compatibility Directive EMC 2002 96 EC Waste of Electrical and Electronic Equipment Safety performance EN 61010 1 1993 A2 1995 IEC 61010 1 1990 A1 1992 A2 1995 including CENELEC Common Modifications US and CA National differences Installation Category Overvoltage Category Il Pollution Degree 2 EN 61010 1 2001 Ed 2 EMC performance EN 50081 1 1992 Generic emission standard Residential commercial and light industry EN 50082 1 1997 Generic immunity standard Residential commercial and light industry EN 61326
8. 1 1997 A1 1998 Product family standard A2 2001 A3 2003 Test standards Performance limits EN 55022 1998 Class B 150 kHz 1 GHz EN 61000 3 2 1995 A1 1998 Class A A2 1998 A13 1997 A14 2000 EN 61000 3 3 1995 EN 61000 4 2 1995 A1 1998 4 kV CD 8 kV AD Criteria B EN 61000 4 3 1996 A1 1998 3 V m 80 MHz 2 GHz Criteria A EN 61000 4 4 1995 1 kV Criteria B EN 61000 4 5 1995 2 kV line to ground 1 kV line to line Criteria B EN 61000 4 6 1996 3 Vims 150 kHz 80 MHz Criteria A EN 61000 4 11 1994 3096 10 ms Criteria B 60 100 ms Criteria C 100 20 ms Criteria B gt 9596 5 s Criteria C Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 28 9 Troubleshooting Guide 9 1 Error messages and warnings Error message Warning Command error Cannot move track Head position error Head timeout Track position error Track timeout Magnets pos error Magnets timeout Serial number error Serialno already set Too high pos offset Invalid parameter Out of memory No acceleration tbl NV parameters lost Progmem full Time already expired Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 Chapter 9 Troubleshooting Guide 29 9 2 Troubleshooting guide Chapter 9 Troubleshooting Guide Error Cause Action e Cannot move The track cannot be moved Press STOP If the fault cannot be remedied track because the head is not up then switch
9. 9404190 Item KingFisher User Manual KingFisher tip comb RNase free KingFisher plate 100 ul RNase free KingFisher plate 200 ul RNase free KingFisher plastics 8 pack 8 plates 100 ul 8 tip combs box KingFisher plastics 200 ul 8 pack 8 plates 8 tip combs box FP Stepper 10 ul 5 ml Finntip 2 5 ml 11 2 List of spare parts Code 11272711 2305290 5186980 Item Quantity Plate tray 1 Serial cable F9 F25 for RS 232C port 1 KingFisher Software 1 11 3 List of recommended spare parts Code 1091910 1092250 2004460 2004470 2004480 21040811 21040820 21040850 24070740 24072290 ltem 1 2 unit s year Tooth belt T2 5 330 Tooth belt T2 5 265 PCB PSP 01 PCB CSU 02 1 PCB CSU 01 Power supply 1 H motor assembly M motor assembly Free wheel assembly 1 Magnetic plate assembly SU gt gt DN gt Naao Contact the Thermo Fisher Scientific technical service department Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 Quantity 1 50 pcs box 50 pcs box 50 pcs box 1 box 1 box 1 100 pcs box 10 units year 3b Chapter 12 Glossary and Abbreviations 12 Glossary and Abbreviations Frame The frame where the special microstrips are placed Magnetic rod The rods which are magnetic and collect magnetic particles The rods do not collect the particles on their own the magnetic rods must always be protected by a tip comb Microstrip The wells where all the reagents
10. DR E eda 24 7 3 Decontamination procedure ccccccccnncnnnnnonencnnnnnnnnnnnn eae ee ee kk ka kaka kk KAKA KAKA KA KA KAKA KA KA KA KA KA KAK AA KA AA A KA 24 Z4 Howto packfor Service oreet eie torpet ed E He eR ea ER REED a wO Ve WEKO VAR e v e aa 25 7 5 Service COFItFacts iiui d rk nter d 25 7 6 Disposal of the instrument seesssssesesseee esee nennen enne nnns KAKA A KAKA A KA KA KA KA KA KA nns 25 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 3 Table of Contents 8 Technical Specifications 000 ii 26 84 General SPSCifiCAtIONS eegne eege edd 26 8 2 Safety SPECIFICATIONS 2 1 rri eret ree ert Rte Fee ri Te Pd n kek k y Hl Ter Fei EA En 27 8 3 In conformity with the requirements ccoooccccconoccnccnaninnncnaninnncnanononcnnnnnnnrcrnnn ka kak ka anar ener KA KA KA KA A 28 9 Troubleshooting Guide E 29 9 1 Error messages and warnings ENEE 29 92 Troubleshooting guide timida tds e k 30 9 3 Frequently asked questions FAQ about the KingFisher esee 31 9 4 Service request DrotoCOl sesenta bale K Ked k AQ ben n 32 9 4 1 DecontaminatiOn eri rhe eie eee en B e rea Eno 32 9 4 1 1 Certificate of Decontamination occccccnnnnnonnnnnnnnnnnnnnnncnnnnnnnnnnnn nn cnn nr enn nn nn rr rre kk K k KAKE KAKA KAK KAKA KK KAKA 33 10 Warranty Certificate soiree ccce rn RE KA KAR K R H k AV ARK Lu Ra KA Rupe EE Yu KWA d w kun EEN 34 10 1 Warranty litirtati ns x cete ya
11. Deggerdal A Jakobsen K S and Larsen F 1997 Rapid universal method to isolate PCR ready DNA using magnetic beads BioTechniques 22 506 511 Sinclair B 1998 To bead or not to bead Applications of magnetic bead technology Scientist 12 16 19 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 37 Chapter 13 Index 13 Index A Padala IKENIEN oI aE AE E S AS SAA E A E E A ES 36 About the User Manitlak aie nren iaa a Een b a EF oira aa de lier 7 AGCCOSSOLF GS cci ite A EE at here alados 15 35 NN 47 Advantages of processing magnetic Darticles EE 9 A DPIC ATOR is er IEEE 5 6 7 31 37 42 46 48 Application protocol KingFisher genomic DNA purification protocol for blood esses nnne 42 KingFisher total RNA pourtftcatton ER 43 B Brief ser S Guld6 iii iia die 45 C Gable iii EN 18 24 35 Certificate of DECONTAMINATION wverravnnnnnnevvnrnnvnnnnnnnnnvenvnnnnnnenenvevvnnnnnnsnenevrnnnnnnsnenvevennnnnnsnesensenen 25 32 33 Bid KK EEE EE M bes 15 18 30 32 GIS As aii s ro aran ERE NEED EE da 16 23 24 31 33 magnetic rods os coire cert ak b W ya tree Ve y ka nye une 23 processing ke l i RC ECRIRE Rev 23 Concentration E deg aba cle ead a aa Dra elena De Ta dada TR RR 13 14 44 D DECONTAMINATION ssi oer eoi cere ee ka a Ze Rv Sa Safi 25 32 33 Certificate Oficina Pug Ua ted exe oda e ca Edd xa dea eas CERTI PESE VR ERAI 25 32 33 Blue 26 DiFe
12. and samples are located and where the or microwell strip processing takes place Plate Frame microstrips 12 pieces Tip comb Protects the magnetic rod A tip comb always has to be fitted into the tip comb holder slot during processing 12 1 Keywords for web pages affinity separation PCR amplification PCR amplification beads PCR product cloned DNA post PCR purification cloning primer DNA probes DNA complex processing DNA immobilization processor DNA purification purification DNA strand separation RNA DNA template mRNA double stranded DNA RT PCR dsDNA separation encapsulated magnetic microspheres separator genome silica coated beads genomic DNA silica coated microspheres hybride selection silica coated particles KingFisher or Kingfisher streptavidin coated beads magnetic beads streptavidin coated microspheres magnetic microspheres streptavidin coated particles magnetic particles subtraction libraries magnetic separation superparamagnetic beads microspheres superparamagnetic microspheres nucleotide template oligo dT Thermo Fisher Scientific particles Thermo Scientific Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 36 Chapter 12 Glossary and Abbreviations 12 2 Literature Aasheim H C Deggerdal A Smeland E B and Hornes E 1994 A simple subtraction method for the isolation of cell specific genes using magnetic mono disperse polymer particles Biotechniques 16 4 716 72
13. authorization number RGA given by the Thermo Fisher Scientific representative e Indicate the fault after you have been in touch with your local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy See Section 8 1 for details on storage and transportation temperatures 7 5 Service contracts We recommend you maintain and service the instrument regularly every 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 the Thermo Fisher Scientific technical service department for more details 7 6 Disposal of the instrument Decontaminate the instrument prior to disposal See Sections 7 3 9 4 1 and 9 4 1 1 on 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 th
14. available when you open the package packing instructions packing list Thermo Scientific KingFisher Feedback Form Warranty Certificate card KingFisher User Manual Caution Do not touch or loosen any screws or parts other than those specially designated in the instructions Doing so might cause misalignment and will void the instrument warranty To lift the instrument put your fingers under the bottom on either sides and lift it with your back straight Retain the original packaging for future transportation The packaging is designed to assure safe transport and minimize transit damage Use of alternative packaging materials may invalidate the warranty Also retain all instrument related documentation provided by the manufacturer for future use 5 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 5 1 3 Checking for damage during transport Visually inspect the transport package the instrument 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 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
15. easy to carry out the whole processing with the aid of an internal program Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 9 Chapter 4 Functional Description 4 Functional Description 4 1 Instrument layout 4 1 1 Front view 1 2 3 4 Keypad Display Front lid Top lid Fig 4 1 KingFisher front view with lids closed Fig 4 2 KingFisher front view with lids open Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 10 Chapter 4 Functional Description 4 1 2 Back view ON OFF switch Mains power supply socket Cooling air outlet Serial port RS 232C RON Fig 4 3 KingFisher rear view 4 1 3 Internal view Thermo Labsystems KingFisher EZ mee y STOP ER Plate carrier Tip comb holder Tip comb slot Fixed magnetic rod holder AS Q N a Fig 4 4 KingFisher internal view Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 11 Chapter 4 Functional Description 4 2 KingFisher magnetic particle processor The KingFisher magnetic particle processor Fig 4 5 is designed for maximum two 96 well plates and special microstrips which are compatible with the tip comb The plate s is are kept stationary and the only moving assembly is the processing head with tip combs and magnetic rods The head consists of two vertically moving platforms One is needed for the magnetic rods 2 x 12 pieces and the other one for the plastic tip combs Fig 4 4 One
16. for longer storage Fig 6 5 Plate containing microstrips specially designed for the KingFisher Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 21 Chapter 6 Routine Operation 6 4 Shutdown Switch the KingFisher off by pressing the power switch Fig 4 3 at the bottom left of the back panel of the instrument into the OFF position Remove any plate still on the plate carrier Dispose of all microplates as biohazardous AN waste Remove any tip combs still in the instrument Dispose of tip combs as biohazardous AN waste Wipe the plate carrier surface and the neighboring instrument surface with a soft cloth or tissue paper moistened with distilled water a mild detergent SDS sodium dodecyl sulfate or soap solution If you have spilt infectious agents on the plate carrier disinfect with 70 alcohol or some other disinfectant see Section 7 3 Decontamination procedure Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 22 Chapter 7 Maintenance 7 Maintenance 7 1 Regular and preventive maintenance For reliable daily operation keep the instrument free of dust and liquid spills Abrasive cleaning agents are not recommended because they are likely to damage the paint finish We recommend you clean the case of the instrument periodically to maintain its good appearance A soft cloth dampened in a warm mild detergent solution will be sufficient Clean the outside of the instrument and the p
17. plate contains twelve separate microstrips and one sample processing uses one microstrip containing eight wells One tip comb containing twelve tips is used for processing twelve samples at a time within one plate Before starting the magnetic particle processing via the keypad and display the samples and reagents are dispensed into the wells and the tip comb s is are loaded into its their slot s The plate s is are placed onto the moving plate carrier in the correct position and the plate carrier is pushed into the end position During the operation the front and top lids can be closed Fig 4 1 or open Fig 4 2 Closed lids protect the processing against environmental contamination The operating principle employed is MPP nverse magnetic particle processing technology Fig 4 6 Rather than moving the liquids the magnetic particles are moved from microwell to microwell containing specific reagents in contrast to the external magnet method Magnetic particles are transferred with the aid of magnetic rods covered with disposable specially designed plastic tip combs KingFisher Fig 4 5 KingFisher magnetic particle processor Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 12 Chapter 4 Functional Description 4 3 Principle of the KingFisher magnetic particle processor Magnetic rod Transfer Tip j bax SS Ss SI P i Magne
18. s fully into the frame Position the plate s correctly Push the microstrip s fully into the frame Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 30 Chapter 9 Troubleshooting Guide 9 3 Frequently asked questions FAQ about the KingFisher Q1 What does the KingFisher system do A1 Thermo Fisher Scientific now offers a complete KingFisher system the magnetic particle processor for the purification and processing of DNA RNA proteins and cells The processor handles particles automatically according to the preloaded purification protocols For more information on these applications contact Thermo Fisher Scientific Oy Q2 Do I need a PC to control the processor A2 Normally the protocol s is are preloaded by Thermo Fisher Scientific and no PC is needed for routine use KingFisher Software can be used to create new protocols or modify existing protocols if needed PC modified protocols are then downloaded into the processor for routine use Q3 Will the magnets get weaker If so how long can they be used Can they be replaced A3 The KingFisher magnets are made of material that is very stable The magnetic field will not get weaker Magnets can be replaced if needed Usually there is no need to replace them but it can be done for example if they are mechanically damaged during runs for example the plate is not correctly positioned during the run Changing the magnets can be carried ou
19. the power on and off e Head position The calculated head position does Check that there is nothing preventing the error not agree with the head home head from moving up and down sensor signal e Head timeout The head has not arrived at the home sensor within the allowed time e Track position The calculated track position does error not agree with the track home sensor signal e Track timeout The track has not arrived at the home sensor within the allowed time e Magnets position The calculated magnetic rod error position does not agree with the rod home sensor signal e Magnets timeout The magnetic rods have not arrived at the home sensor within the allowed time e Nonvolatile The nonvolatile parameters of the parameters lost instrument are lost e Clattering noise Plate s and or microstrip s is are not in place e Last release poor Plate s and or microstrip s is are not in place Check that there is nothing preventing the head from moving up and down Check that there is nothing preventing the track from moving left or right Check that there is nothing preventing the track from moving left or right Check that there is nothing preventing the magnetic rods from moving up and down Ensure that the tip comb is fully in Check that there is nothing preventing the magnetic rods from moving up and down Contact authorized service Position the plate s correctly Push the microstrip
20. 0 e Select the desired protocol from the keypad and display p 19 e Press the START button p 20 e After the end of the run remove the plate and tip combs p 22 e Maintain your KingFisher instrument regularly p 23 Thermo Labsystems KingFisher Fig 16 1 KingFisher keypad Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 45 Appendix C Thermo Scientific KingFisher Feedback Form Please send to Thermo Fisher Scientific Oy Fax 358 9 32910415 17 Appendix C Thermo Scientific KingFisher Feedback Form Cat no Serial no PURCHASED BY PURCHASED FROM Company Institute Distributor Department Address Address Tel Tel Fax Date of delivery Internet home page Date of purchase Your research area Dr L1 Mr O Mrs O Ms O Job title Position Surname block capitals First name block capitals Internet e mail address Excellent Above As Below Comments expectations expected expectations Reagent kit Instructions D O L1 a Instrument User manual D O L1 O Operational reliability n n n O Design O n O O Ease of use D O O L1 Operational costs D n O O Customer support n n O O Further instrument system developments desired Further applications desired Where did you first learn about the product Would you like to receive information about other Thermo Scientific products Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 46 18 Appendix D Addresses For the latest informati
21. 1 Coche T Dewez M and Beckers M C 1994 Generation of an unlimited supply of subtracted probe using magnetic beads and PCR Nucleic Acids Res 22 1322 1323 Cudjoe K S 1999 Immunomagnetic particle based techniques Overview In Eds Robinson R K Batt C A and Patel P D Encyclopaedia of Food Microbiology Academic Press London UK Pp 1088 1094 Hornes E and Korsnes L 1990 Magnetic DNA hybridisation properties of oligonucleotide probes attached to superparamagnetic beads and their use in the isolation of poly A mRNA from eukaryotic cells Genet Anal Tech Appl GATA 7 145 150 Jakobsen K S Breivold E and Hornes E 1990 Purification of mRNA directly from crude plant tissues in 15 minutes using oligo dT microspheres Nucleic Acids Res 18 3669 Jakobsen K S Haugen M S boe Larsen S Hollung K Espelund M and Hornes E 1994 Direct mRNA isolation using magnetic Oligo dT Beads A protocol for all types of cell cultures animal and plant tissues In Eds Uhl n M Hornes E and Olsvik Advances in Biomagnetic Separation Eaton Publishing Pp 61 71 Lambert K N and Williamson V M 1993 DNA library construction from small amounts of RNA using paramagnetic beads and PCR Nucleic Acids Res 21 775 776 Lee Y H and Vacquier V D 1992 Reusable cDNA libraries coupled to magnetic beads Anal Biochem 206 206 207 Rudi K Kroken M Dahlberg O J
22. 19 Litera tU a A E E E MS 37 M Magnetic particle ProC SSOF cccccccncncnnncnnnonononononononononononononononononnnnnnnnnnnnnnnnnnnnnnnnnnnnonons 7 8 9 12 13 31 Magnetic particles eene halan ana vorne eeu k ral aa 8 9 12 13 14 31 36 43 collection Of iecit ier eei P e dla ai aust ri 13 release fun oe detecta Ree tiere eder ete dae M KE D Ma vel s ne re Eod ad dl ld 13 Wa S N oftast EMITE 13 14 Magnetic tod LR 8 11 12 13 14 23 26 30 31 36 MAINS E cio baii T ATAA A A A 11 18 24 26 27 Manta daa 23 25 45 Maintenance EE 7 23 25 32 MiC hostii Piar ia aaa 8 12 21 26 30 36 O Operational ti tette tee Pre o ree aene b las Mesa 18 Ordering ei tege WEE 7 35 P Packing her ere ide dadas 15 25 32 CEA TD NT TNR 15 25 TE EE EE EE EEE pan Scab aloe AGNES A EE E EE 15 materials id tecti aa eber a NN dee ideal id 2b PER esa 31 36 37 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 39 Chapter 13 Index Plate e eret cic ea eda aec nde Mnd 8 11 12 20 21 22 23 24 30 31 35 36 43 44 45 CM Skiene ed 12 20 22 23 24 45 Plate carrie Piian e A usi La bid n dak b n eda 12 20 22 23 24 45 Processing head TEE 12 Protocol A AE A EE cede e eben ee tec oc d utv drake 8 19 31 32 37 42 43 45 R Releasing the transport lock meca o EE A Pe SEITE a TEA 16 p EE 36 37 44 REPCR PAN SR ege ge A ee Eege 36 S Satmples erede eden ee eden det de ide 8 9 12 16 21 24
23. 2008 5 Chapter 1 Safety Symbols and Markings 1 Safety Symbols and Markings These symbols are intended to draw your attention to essential information and alert you to the presence of hazards as indicated SAFETY SYMBOLS USED IN THE KINGFISHER Power ON Power OFF WEEE symbol This product is required to comply with the European Union s Waste Electrical amp Electronic Equipment WEEE Directive 2002 96 EC x O WARNING MARKINGS USED IN THE DOCUMENTATION Caution risk of electric shock Caution biohazard risk Caution risk of personal injury to the operator or a safety hazard to the surrounding area Caution risk of damage to the instrument other equipment or loss of performance or function in a specific application gt gt gt gt Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 6 Chapter 2 About the User Manual 2 About the User Manual This user manual has been written for the actual end user for example laboratory technician and provides information on the Thermo Scientific KingFisher magnetic particle processor including installation and operating instructions Read the manual in its entirety prior to operating the instrument This user manual has been designed to give you the information you need to e Install the KingFisher e Usethe KingFisher in routine the processing step e Perform basic maintenance procedures This user manual also describes all the features an
24. Copyright Copyright O 2008 Thermo Fisher Scientific All rights reserved First edition published in 1999 Printed in Finland Reproduction of the accompanying user documentation in whole or in part is prohibited Patents The KingFisher has national and international patents as well as patents pending Trademarks and Finntip and KingFisher are registered trademarks of Thermo Fisher Scientific Virkon is a registered trademark of Antec International Ltd All other trademarks and registered trademarks 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 as part of a continuous product development 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 P O Box 100 FI 01621 Vantaa Finland Tel 358 9 329 100 Fax 358 9 3291 0415 www thermo com Thermo Scientific KingFisher User Manual Rev 2 4 June
25. CtiVes cca xelaye ka nte code cerent te eee stew Cor eiut dar edad keya ku b Deyn an oe W Ee ve e a vty Cod su deva aya Vu saa d ad tg dra 28 Dis EI REDE 5 Disinfection Grocedure EE 22 23 24 25 32 33 Disposal of instr merbt 2 nante tet hide KE KER N keke KIN KRG y Ka R KEN ARGE 25 Disposal of materials tree eA All a a ah esi a EAR A 24 DNA a A a a eva oe 9 21 31 36 37 42 E Environmental requlrements irsmui te rere ce eto ddmd ceo rede lee e Cup ed epa t ea eek uvas 16 Etrorqmessatge 5 cire tr metre air 7 29 F FAO oe it i testo merbart etes cu ne Ded e Pt E ELE FU et oe 31 Feedbaek FORM ae Seege geegent 15 46 EMB a clade yaa hous PS aci 30 36 Frequently asked We LTE 31 G Eltere EE e E MEN MENOR 36 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 38 Chapter 13 Index H How to clean the magnetic rode EEN 23 How to clean the plate Carrier nerit RE etie ka L ee ku R Waa er an bs 23 How to clean the processing chamber enne nennen KA KAKA KA KA KA KA KA KA KA nnn nnns 23 How to disinfect the Instrument 22 23 24 32 33 How to ensure startup ete no irren aiaa a 20 How to install the King FISR l ana a k lika kaka ranna Ruh an Me Wl in dw Ain H RAN Ek ia Akan Z wa nin SU wa kn k 15 27 How to maintain the instrument E a a k lek kelaya s k s dar a dey n ak s e RA KL kav ke b we d v Weke saa rr 23 How 10 Pack for Selvi s xak yake EZ A
26. a WEKE KA RETE REN CRF S PARARE Ta d k R aa HEWA KCK e na a a E ay n 25 32 How to release the transport lock y i kaka kk kk aka kk KAKA KAKA KA KA KA KA KA KAK AK KA KA KARA KA nnns 16 How to setup the instrument before you put it into operation cnn nnnnannnnnnnos 16 How to Shut GOWN EE 22 HOW O AKA IKO N EEE PERE A E EE See diare rd 19 21 How to troubleshoot the mstrument unne 30 How to Uunpack tirer ete ATV FOIE KAUNAS AIR sene 15 How to use the keypad and display l l kk kaka kak aa ae ee aka kk kk K k ka kK A KAKA KA KA KA KA KA KA KA KAK A KA KA 19 How to work with a magnetic ro 13 INCUBATION K yc ii ee b ka ett nak sa nad Ked edat EO EE ed eta re Ted evt tui e 9 13 14 26 43 Sala ce ee at keren 7 15 16 25 27 Instrument EI sertuakvouss 10 Intended tuse niii cic ire tte ifa ree EC he bl eme E Lien Ye e Ee ben s n Sad E E EH Hm in iset 8 Inverse magnetic particle processing 4 4 kk aka kk nn nn cnn nana KAKA KA ka KA KA KA KA KA KA KA KA KA enn 12 13 K Keypad and display cin ia add side save Fata ya i kak cums EI sees Hussein 8 12 19 45 KO Sita A AA ya c da nak n sa Hak K a eae w ran D wa ee dod E avs sn 19 26 KeyWords for WED Page is xit mix hiy Reka k y ke w y can es xa A RA a BERA cdta 36 dep 8 34 46 L Liguidierystal displays ihi ere rrr three eee
27. a aka nennen nnns 8 4 1 KingFisher front view with lids closed 10 4 2 KingFisher front view with lids open 10 4 3 TS SN ee M e rasen e a ERE e Ue a En 11 4 4 KingFisher internal view rra 11 4 5 KingFisher magnetic particle Grocessor eene nennen nisse nnne 12 4 6 Inverse magnetic particle Grocessimng nnne KE kak ka E KE KA AA KA ka KAK ku ku 13 4 7 Concentration step during magnetic particle processing sssssssss een 14 5 1 KingFisher transport lock tag present before release of the jockpiece renner 17 5 2 Location of the transport lock SCreWs uLEEL ELLE rennene rr er nn tn aa aa annes kaka ka kaka ka s HA ku kaka k 17 5 3 CONNECTING ING Mans SUP PIV Cables a iti tt b t t te e p M Ana 18 6 1 KingFisher keypad i rt EUR a UR Pande eee a HEHE EH HH JMM N 6 19 6 2 Loa d1ng the plate Carriers s e Der ebi E PE ret Meet gear 20 6 3 Positioning of the plate s and insertion of the tip combtsl lle llkkekeae 20 6 4 Tip comb specially designed for the KingFisher ooo 21 6 5 Plate containing microstrips specially designed for the Kioofisber sss 21 16 1 KingFisher Keypad motto te ME eiat Mte E P PE LE beti Fro pe spo 45 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 41 Appendix A Application Protocols for KingFisher 15 Appendix A Application Protocols for KingFisher A 1 Example of a KingFisher genomic DNA purification protocol from blood The genomic DNA isolation kit based on paramagnetic parti
28. 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 Date and place Signature Name block capitals Please include decontaminating solution used PHOTOCOPIABLE Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 33 Chapter 10 Warranty Certificate 10 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
29. cles can used with the KingFisher magnetic particle processors to automate genomic DNA purification Blood can be used as starting material for the DNA purification with the KingFisher The isolated DNA can be used for several downstream applications including PCR and restriction analysis The gDNA purification has been adopted into the KingFisher microplate format The kit reagents are pipetted into the KingFisher plate After filling the KingFisher microplate insert the plate into the KingFisher instrument and choose the correct program by using the arrow keys The KingFisher program is called Genomic DNA 1 Start the program by pressing the START button Purified genomic DNA is released into a 40 ul volume into the last well H and the particles are returned into well F This genomic DNA purification protocol takes ca 30 minutes You can use lt 40 pl of blood as starting material The description of the KingFisher protocol is as follows Program 1 The sample is incubated with particles in well A for 3 minutes 2 The sample is incubated with particles in well B for 3 minutes 3 The sample is incubated with particles in well C for 3 minutes 4 The sample is incubated with particles in well D for 3 minutes 5 The particles are washed in well E 6 The particles are washed in well F 7 The particles are washed in well G 8 The purified DNA is released into distilled water in well H 9 The particles are removed and returned into wel
30. d specifications of the KingFisher hardware and on board software Chapter 15 Appendix A Application Protocols for KingFisher explains the processing principles and procedures In Chapter 9 Troubleshooting Guide you will find explanations of all error messages and a problem solving guide The user should be familiar with the contents of Chapter 7 on maintenance For warranty and ordering information refer to Chapters 10 Warranty Certificate and 11 Ordering Information In an effort to produce useful and appropriate documentation we appreciate your comments on this document to your local Thermo Fisher Scientific representative Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 7 Chapter 3 Introduction to the KingFisher 3 Introduction to the KingFisher 3 1 Intended use The KingFisher magnetic particle processor Fig 3 1 is intended for professional research use by trained personnel The instrument is intended for automated transfer and processing of magnetic particles in a microwell scale Use for self testing is excluded It is recommended that Good laboratory Practices GLP are followed to guarantee reliable analyses 3 2 Principle of operation The KingFisher magnetic particle processor Fig 3 7 is designed for the automated transfer and processing of magnetic particles in a microwell scale The principle of the KingFisher system patents pending is based on the use of magnetic rods covered with the dispo
31. e carrier out Fig 6 2 Thermo Labsystorms KingFisher Cut corner Fig 6 2 Loading the plate carrier e Load only maximum two special plates Fig 6 5 onto the plate carrier for processing at a time The plates have to be positioned so that the cut corners face left down Fig 6 2 They also have to be in their places Slide the plate carrier fully back in Thermo Labsysterns KingFisher Cut corner I Fig 6 3 Positioning of the plate s and insertion of the tip comb s Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 20 Chapter 6 Routine Operation e The tip comb s always has have to be in its their slot s and fully inserted Fig 6 3 The instrument also functions with either one microplate or one to twelve microstrips depending on the amount of samples At least one tip comb Fig 6 4 always has to be inserted into either of the narrow slots meant for the tip comb Fig 6 4 Tip comb specially designed for the KingFisher e Close the front lid if you want to The front lid protects the instrument against environmental contamination Note The front lid and or top lid can be left open if desired This action does not break the run e Select the name of the method for example Genomic DNA 1 e Use the START key to initiate the run For startup refer to Section 6 1 Switching on Note The samples are collected immediately after the run and transferred into other tubes
32. e original packaging and packing materials use the recycling operators known to you For more information contact your local Thermo Fisher Scientific representative Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 25 Chapter 8 Technical Specifications 8 Technical Specifications 8 1 General specifications Technical specifications Overall dimensions Weight Operating conditions indoor use Transportation conditions Storage conditions Mains power supply Power consumption Heat dissipation Computer interface W x D x H 290 x 290 x 310 mm 11 4 x 11 4 x 12 2 in ca 10 kg 23 lbs 10 C 40 C RH 90 max 40 C 70 C packed in transport packaging 25 C 50 C packed in transport packaging 100 240 Vac 50 60 Hz nominal Automatic voltage detection 80 VA max 50 VA standby 273 BTU max Serial RS 232C port Baud rate 9600 Character format 1 start bit 8 data bits 1 stop bit no parity Flow control XON XOFF Performance specifications Microplate type disposable Magnetic rods fixed Capacity Vessel volumes Vessel type disposable Tip comb disposable Magnetic particle size Collecting efficiency Incubating temperature Keyboard Display Software Special design 8 x 12 grid format 2 x 12 format 24 samples run 2 x 12 20 200 ul Special microstrip 1 x 8 wells Special design 1 x 12 format ca gt
33. ells or 3 10 mg of tissue for the purification Yields will vary according to the kit cell and tissue type used in the assay The name of the purification protocol is Total_RNA_1 Pipetting the reagents KingFisher Well KingFisher microplate 100 ul A Cell lysate OR tissue homogenate 100 pl magnetic particles 10 ul Washing buffer 1 100 ul Washing buffer 1 100 ul Washing buffer 1 100 ul Washing buffer 2 100 pl Washing buffer 2 100 pl Washing buffer 2 100 pl Elution buffer 40 ul T mimj oco joj u Protocol Total_RNA_1 The sample is incubated with magnetic particles in well A for 18 minutes The particles are washed in washing buffer 1 in well B The particles are washed in washing buffer 1 in well C The particles are washed in washing buffer 1 in well D The particles are washed in washing buffer 2 in well E The particles are washed in washing buffer 2 in well F The particles are washed in washing buffer 2 in well G The particles are released into the elution buffer in well H The protocol is paused for a heating step The text marked PAUSED appears on the screen 10 The plate is removed and transferred to the incubator for the heat incubation 11 After incubation the plate is returned to the KingFisher 12 The protocol will continue after you press START 13 The particles are removed and returned into well E o0 og B6 N Thermo Scientific KingFisher User Man
34. g tag for future transportation gamme A an Thermo Labsystarns her SART E Stor gt Tag attached here Fig 5 2 Location of the transport lock screws Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 17 Chapter 5 Installation 5 2 2 How to ensure the startup 1 Warning Ensure that the mains switch Fig 4 3 on the bottom left of the back panel is in the OFF position left of the back panel If any other type of mains supply cable than supplied is needed use only cables certified by the local authorities Before you plug in the power cable ensure that the voltage on the rating label at the bottom left of the back panel Fig 4 3 corresponds to the local voltage 2 Connect the mains supply cable to the mains power socket Fig 5 3 at the bottom 3 Connect the instrument to a correctly installed line power outlet which has a protective conductor that is grounded Warning Never operate your instrument from a power outlet that has no ground connection y Fig 5 3 Connecting the mains supply cable 5 3 Operational check 1 First switch the instrument ON The instrument performs initialization tests and adjustments The initialization tests are so called self tests 2 The display quickly shows the internal software version and after that the display shows one of the protocols in the internal memory This happens when the initialization tests and adjustments have been completed
35. ion SDS soap solution or alcohol 7 1 3 How to clean the processing chamber If required clean the processing chamber using a soft cloth or tissue paper soaked in a mild detergent solution SDS soap solution or alcohol Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 23 Chapter 7 Maintenance 7 2 Disposal of materials Refer to local regulations for the disposal of infectious material The samples can be potentially infectious Dispose of all used microplates disposable AN gloves syringes disposable tips etc as biohazardous waste 7 3 Decontamination procedure 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 before relocating the instrument from one laboratory to another Decontamination is not required for the proper functioning of the instrument Example of decontaminants Ethanol 7096 Virkon solution 1 396 Glutaraldehyde solution 4 Chloramine T Microcide SO 1 64 Always use disposable gloves and protective clothing and operate in a well ventilated N area 1 Prepare the decontaminant for example 200 ml 4 glutaraldehyde solution or another agent recommended by your safety officer 2 Empty the plate carrier Ensure that you are wearing disposable gloves 3 Switch OFF the power and disc
36. l ak ave b kan ada 34 11 Ordering Information nee IER enira rea ne cp ked R ken d yk YR a ENNAN ASER NR ENEE d e dee 35 11 1 List Of ACCESS ONES iste dalal etie reo re e Tie kite Fela 35 11 2 List of sp re DA P S Saya coena eene A AAA A dr ee ede eraat 35 11 3 List of recommended spare parts 35 12 Glossary and Abbreviations cies eese rain iii 36 12 1 Keywords for Web Da kya dlt edited tne ke cic ii ier d ree e b v re e x 36 12 2 A E 37 13 lI c WER 38 14 Table of e Cc EM 41 15 Appendix A Application Protocols for KingFisher rrnnvnrnnnnnvrnnnnnnvnnnnnnnvenennnnvnnnnnnnnvnnernnnenn 42 A 1 Example of a KingFisher genomic DNA purification protocol from blood 42 A 2 Example of a KingFisher total RNA purification protocol seen 43 16 Appendix B Thermo Scientific KingFisher Brief User s Guide 45 17 Appendix C Thermo Scientific KingFisher Feedback Form esses 46 18 Apperidix D Addresses uerit rentes rer ak k d wu a DERDEK A Lane ku a WARA xo M e a k WEK 47 19 Appendix E Supplementary Application Protocols rrrrnnnnvnrnnnnnvnnnnnnnvnnnnrnnnvnrennnnvnnernnnnen 48 Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 4 Table of Contents
37. l F Volumes A Blood 10 pl lysis buffer 75 ul particles 16 pl B Blood 10 pl lysis buffer 75 pl C Blood 10 pl lysis buffer 75 pl D Blood 10 pl lysis buffer 75 ul E Washing buffer 100 pl F 7096 EtOH 100 pl G 70 EtOH 100 pl H Distilled water 40 pl You can make a master mixture of the blood and lysis buffer 40 ul of blood 300 ul of lysis buffer Then you dispense 85 ul of the mixture into wells A D Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 42 Appendix A Application Protocols for KingFisher A 2 Example of a KingFisher total RNA purification protocol The total RNA purification kit based on paramagnetic particles can be used for the purification of total RNA from cultured cells or tissue homogenates The reagents and specific consumables are designed to be used with the KingFisher magnetic particle processors as part of an integrated system The paramagnetic silica particles bind RNA in the presence of chaotrophic salts After the binding step the RNA magnetic particle complex is washed extensively in washing buffers to remove any remaining contaminants Finally the purified RNA is released into the elution buffer During the elution step the samples are heated outside the KingFisher instrument With this protocol processing time for 24 samples is 40 minutes The isolated RNA can be directly used in downstream applications such as RT PCR and Northern blotting You can use 1 x 10 c
38. late carrier with clean low pressure compressed air or a cloth dampened with water or a mild detergent when necessary Although the KingFisher is constructed from high quality materials you must immediately wipe away spilt saline solutions solvents acids or alkaline solutions from outer surfaces to prevent damage Painted surfaces can be cleaned with most laboratory detergents Dilute the cleaning agent as recommended by the manufacturer Do not expose painted surfaces to concentrated acids or alcohols for prolonged periods of time as damage may occur Clean the display areas with a mild laboratory detergent The keypad has a wipe clean surface Plastic covers and surfaces can be cleaned with a mild laboratory detergent or alcohol If any surfaces have been contaminated with biohazardous material a mild sterilizing A solution should be used 2 Do not autoclave any part of this instrument 7 1 1 How to clean the plate carrier Keep the plate carrier surface clean to avoid dust and dirt entering into the instrument Clean the plate carrier surface at least once a week using a soft cloth or tissue paper soaked in a mild detergent solution SDS soap solution or alcohol If you have spilt infectious agents on the plate carrier disinfect according to Section 7 3 Decontamination procedure 7 1 2 How to clean the magnetic rods If required clean the magnetic rods using a soft cloth or tissue paper soaked in a mild detergent solut
39. local Thermo Fisher Scientific representative or Thermo Fisher Scientific Oy Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 15 Chapter 5 Installation 5 1 4 Environmental requirements When you set up your KingFisher 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 lids 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 and at the back of the unit to allow adequate air circulation The KingFisher does not produce operating noise at a level that would be harmful No sound level measurements are required after installation Warning Do not operate the instrument in an environment where potentially damaging liquids or gases are present 5 1 5 Things to avoid Do not smoke eat or drink while using the KingFisher 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 f
40. luids in or on the equipment Warning This product contains strong permanent magnets People wearing a pacemaker or metallic prostheses should not use this product A pacemaker or prostheses may be affected or damaged if it comes in very close contact with a strong magnetic field 5 1 6 Technical prerequisites Place the instrument on a normal laboratory bench The net weight of the unit is approx 10 kg 23 Ibs The instrument operates at voltages of 100 240 Vac and a frequency range of 50 60 Hz 5 2 Setups before you put the instrument into operation 5 2 1 How to release the transport lock Note THERE IS ONE TRANSPORT LOCK PRESENT Fig 5 2 A white tag with the red text CAUTION Remove Transport Lock Before Use is fastened to the instrument to attract the user s attention Fig 5 7 The tag in Fig 5 7 and the black arrows in Fig 5 2 indicate where the transport lock piece is situated Remove the tag and release the transport lock Ensure that the transport lock has been released before you put the instrument into operation Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 16 Chapter 5 Installation Fig 5 1 KingFisher transport lock tag present before release of the lock piece 1 Toremove the transport lock unscrew the three screws Fig 5 2 with the 2 5 mm hexagonal screwdriver supplied 2 Remove the transport lock piece 3 Save the transport lock piece and screws and the transport lock warnin
41. lysis buffer using the same ratio as above Tools with a diameter of 5 mm or 7 mm are suitable for small volumes a minimum of 300 ul and can be used for homogenization in microfuge tubes Refer to your homogenizer supplier for suitable handling volumes e Homogenize the sample on ice in the presence of lysis buffer for 5 90 seconds depending on the toughness of the tissue e Centrifuge the homogenate for 1 minute at 1000 g 3000 rpm in a microcentrifuge Note Ensure that you follow the centrifugation instructions Do not exceed the time and speed as this will drop the RNA yield drastically Note If a rotor stator homogenizer is not available other homogenization methods can be used instead For example soft tissues such as liver can be homogenized using for example a pestle and needle syringe method Always follow the centrifugation instructions above despite the homogenization method used e Add up to 100 ul of the supernatant into the KingFisher plate according to the instructions Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 44 Appendix B Thermo Scientific KingFisher Brief User s Guide 16 Appendix B Thermo Scientific KingFisher Brief User s Guide e Switch the KingFisher on p 18 e Pipette the samples reagents and washing buffers into the microwells of the plate s p 42 e Load the plate s onto the plate carrier p 20 e Insert the tip comb s into the tip comb holder slot s p 2
42. netic particles Washing the magnetic particles is a frequent and an important processing phase Washing is a combination of the release and collection processes in a microwell filled with washing solution To maximize washing efficiency the magnetic rods together with the tip comb s are designed to have minimized liquid carrying properties 4 4 4 Incubation To keep the magnetic particle suspension evenly mixed in long running reactions the tip comb s can be moved up and down from time to time 4 4 5 Changing the volume during the magnetic particle processing The volume of the first microwell can be larger than the volume of the next microwell and this is used for concentration purposes see Fig 4 7 below Y Transfer r v A y E gt El N j v v j o a Collection of Transfer into Release of Ready magnetic particles a low volume well magnetic particles suspension in a from a large volume low volume well well Fig 4 7 Concentration step during magnetic particle processing Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 14 Chapter 5 Installation 5 Installation 5 1 What to do upon delivery 5 1 1 How to unpack Move the unpacked instrument to its site of operation Unpack the KingFisher instrument and accessories carefully with the arrows on the transport package pointing upwards The following notes and instructions are sent with the instrument and are immediately
43. on on products and services visit our websites at http www thermo com http www thermo com kingfisher Manufactured by Distributed by Thermo Fisher Scientific Oy P O Box 100 FI 01621 Vantaa Finland Tel 358 9 329 100 Fax 358 9 3291 0415 www thermo com Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 Appendix D Addresses KingFisher 47 Appendix E Supplementary Application Protocols 19 Appendix E Supplementary Application Protocols Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 48 NOTES Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 49 NOTES Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 50 NOTES Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 51 Thermo Fisher Scientific Oy Microplate Instrumentation Ratastie 2 P O Box 100 Fl 01621 Vantaa Finland www thermo com 1507730
44. onnect the mains supply cable Fig 5 3 4 Disinfect the outside of the instrument using a cloth dampened with 70 ethanol b Placethe instrument in a large plastic bag Ensure that 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 ethanol Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 24 Chapter 7 Maintenance 11 After performing this decontamination procedure enclose a signed and dated Certificate of Decontamination both inside the transport package and attached to the outside of the package 7 4 How to pack for service When you ship the instrument for service remember to Inform about the use of hazardous materials Decontaminate the instrument beforehand Install the transport 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 e Enclose a dated and signed Certificate of Decontamination see Section 9 4 1 1 both inside and attached to the outside of the package in which you return your instrument or other items e Enclose the return
45. sable specially designed tip combs and microstrips These are provided with the reagent kit The instrument functions without any sucking or aspiration parts or devices Samples and reagents including magnetic particles are dispensed into the wells according to the corresponding kit instructions The use of microplate format allows the use of microwell scale volumes and existing liquid handling tools The steps of the protocol that are selected by the user via the keypad and display are preloaded in the on board software KingFisher START STOP Fig 3 1 KingFisher magnetic particle processor Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 8 Chapter 3 Introduction to the KingFisher 3 3 Advantages of processing magnetic particles with the KingFisher The KingFisher magnetic particle processor provides several advantages relating mainly to the principle of operation in that it Enables automation of complicated manual steps Enables simultaneous processing and purification for example of double stranded DNA Enables a quicker reaction and a more efficient wash due to the simultaneous shaking during incubation and washing Facilitates a good collection of bead bound sample due to the efficiency of the magnet Prevents carry over due to the fact that the same tip is only used for one sample and furthermore that the reagents have been pipetted into their own wells different reagents are not added into the same well Is
46. shing magnetic particles nennen nennen kad e kw s n 14 4 4 4 incubation 3522 2 Sawing ane hne Ic EIE UD 14 4 4 5 Changing the volume during the magnetic particle prorcessing 14 5 Installation sus LEE 15 5 1 Whatto do upon delivery iip RII I eg UE Ten 15 5 1 1 How tolunpaek ser 15 5 1 2 Checking delivery for completeness rrssrrrrrrnnrrvnnrrnnvrrrrrnnvnesrrnnvnrsrennrnessennvnsssennrnsssennnne 15 5 1 3 Checking for damage during transport 15 5 1 4 Environmental requirements annan nn aka teeta mn rra rre 16 5 1 5 Things to VOI race idit 16 5 1 6 Technical prerequisites smc e dan 16 5 2 Setups before you put the instrument into operation oooocccnnccnnnccononnncnananancnanannncrnnnnnnnnnnn nn 16 5 2 1 How to release the transport lock sess 16 5 2 2 How to ensure the Startup EEN EEN 18 5 3 Operational check iia 18 6 Routine Operation ecc id iii 19 d MEE reri delP 19 6 2 Howto use the keypad and depnlay anne kk nc nn enne kK KA KA 19 6 3 HOW To START a bear ect ee Hi E REP il 20 64 gn EE 22 Z7 Maintenance JERICHO 23 7 1 Regular and preventive maintenance ococccccccccnnnnnnnoncnnnnnnnnnnnnnnn cnn rn nn rn kk keka aka k kk KAKA KA KAKA KA KA KAK KA KAR 23 7 1 1 How to clean the plate carier nennen KAKA KAKA KA KA 23 7 1 2 How to clean the magnetic rode 23 7 1 3 How to clean the processing chamber 23 7 2 Disposal Of materials necne nere erre e nte enean daka kelk a Ra Haee ee
47. t by local technical service personnel O4 How strong are the magnets Can they disturb for example some sensitive equipment A4 The magnetic field is situated just around the magnetic tips so it is very local Therefore the magnetic field should not cause any problems with surrounding equipment However if you touch your floppy disk with the magnetic rod you will lose all your data Generally a normal PC screen may also detect the magnetic field with a minor distortion of the image Q5 What happens if I forget to insert the tip combs into the tip comb holder and the magnetic particles stick to the magnetic rods Ab Wipe the magnetic rods with a soft cloth or tissue paper soaked in a mild detergent solution soap solution or alcohol Q6 What if magnetic particles remained in the sample well A6 If the starting material is too viscose the magnetic rods will not be able to collect the particles Dilute the sample and check that the sample is properly homogenized lysed Q7 What if magnetic particles are attached to the tip combs A7 This happens sometimes but it will not affect the yield because the sample has been released from the particles Q8 Are the volumes of reagents in each well critical A8 We strongly recommend that you keep the given volumes within certain limits to avoid failure in performance of the chemical reactions and the processor Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 31
48. t 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 9 4 1 Decontamination 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 recommendable when infectious substances have been in direct contact with any part s of the instrument Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 32 Chapter 9 Troubleshooting Guide 9 4 1 1 Certificate of Decontamination The decontamination procedure is required prior to shipping 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 7 3 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 item s ensure that they are fully decontaminated Confirm A or B status Name Address Tel Fax Name Serial no A confirm that the returned items have not been contaminated by
49. 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 conditions 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 10 1 Warranty limitations Consumables are not included in the warranty The warranty does not extend beyond the expiration date of the reagent kit Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 34 Chapter 11 Ordering Information 11 Ordering Information Code 5400000 Instrument System KingFisher 100 240 V 11 1 List of accessories Code 1507730 97002070 97002080 97002084 97002090 97002094 4540000
50. tic particle Collection of Magnetic particles Release of Ready suspension before magnetic particles on the tip surface magnetic particles suspension collection Fig 4 6 Inverse magnetic particle processing 4 4 Working with a magnetic rod Working with magnetic particles can be divided into five separate processes collecting particles releasing particles washing particles incubation concentration 4 4 1 Collecting magnetic particles During the collection of the magnetic particles the magnetic rod bar is fully inside the tip The magnetic rods together with the tip comb s move slowly up and down in the microwells and the magnetic particles are collected onto the wall of the tips The magnetic rods together with the tip comb s having collected the magnetic particles can be lifted out of the microwells and transferred into the next microwells 4 4 2 Releasing magnetic particles After collection of the magnetic particles the magnetic rods together with the tip comb s are lifted from the microwells the magnetic rods are lifted off and the tip comb s is are lowered into the next microwells containing a reagent Magnetic particles are released by moving the tip comb s up and down several times with considerably high speed until all the particles have been mixed with the substance in the next reaction Thermo Scientific KingFisher User Manual Rev 2 4 June 2008 13 Chapter 4 Functional Description 4 4 3 Washing mag
51. ual Rev 2 4 June 2008 43 Appendix A Application Protocols for KingFisher Sample preparation from cultured cells e Transfer cultured cells suspended in PBS into a suitable centrifuge tube e Collect the cells by centrifugation for 5 minutes at 6000 rpm and discard the supernatant e If frozen cells are used thaw the cells on ice centrifuge the cells for 5 minutes at 1000 rpm and discard the supernatant e Add lysis buffer including 10 ul B ME 1000 ul buffer so that the final concentration is 1 x 10 cells 100 ul e Mix well by vigorously pipetting up and down several times until the lysate is homogenous Vortex for an additional 30 seconds at full speed Note If a lot of foam has formed keep the sample on ice while filling the KingFisher microplate with other reagents or centrifuge the sample for 1 minute at 1000 g 3000 rpm in a microcentrifuge Note Ifa centrifuge is used ensure that you follow the centrifugation instructions Do not exceed the time and speed as this will drop the RNA yield drastically e Mix the homogenate gently by pipetting and add the cell mixture into the KingFisher plate according to the instructions Sample preparation from tissue e Weigh the needed amount of tissue up to 10 mg and add 300 ul of lysis buffer including 10 ul B ME 1000 ml buffer Depending on the rotor stator homogenizer for example IKA Ultra Turrax Heidolph Diax etc and the tool used change the amount of tissue and

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