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159.50_ver05 BeadRetriever User Manual

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1. 11 43 PRINCIPLE OF MAGNETIC BEAD iii 12 4 4 WORKING WITH A MAGNETIC ROD seine 12 4 4 1 0 12 4 4 2 Releasing magnetic beads icc nat es 12 449 Washingimagnetic beads iir AEE i 13 4 4 4 13 445 Changing the volume during the magnetic bead 13 5 INSTAELATIONU A AA 14 5 1 WHAT DO UPON DELIVERY 14 5 1 1 HOW 0 14 5 1 2 Checking delivery for 14 5 1 3 Checking for damage during transport 14 5 1 4 Environmental requirements 15 515 Things to AVOID yR FY LED A 15 5 1 6 Technical 222 00000 00 LLYRY ELE LLE 15 5 2 SETUPS BEFORE YOU PUT THE INSTRUMENT INTO OPERATION 15 5 2 1 How to release the transport 15 5 2 2 How 10 ensure startUD euw cw dy tante net ttes Fy GYRFA 17 5 3 OPERATIONAL 17 6 ROUTINE OPERATION nn cesse tte te NWR Y DY
2. E TG NAT een sel 17 7 33 a 14 24 30 INSTRUCTIONS ais 14 24 EE Eda bs 14 Materials uu ed 24 45 Us dti 29 33 42 44 45 48 49 51 Sens DDIAL OTDD UL 11 Protocolau ni 8 18 29 30 50 54 5 Samples esses cscs 8 11 15 19 20 23 25 29 34 40 41 43 44 46 47 48 50 54 S paration Meu Acie 29 34 35 40 43 48 24 25 28 30 A 24 24 30 ee a 30 SUE AO 21 A 7 25 26 general a ddn YD
3. 36 14 TABLE OF FIGURES ie eii YDD Wy FR 39 15 APPENDIX APPLICATION PROTOCOLS FOR BEADRETRIEVER USING DYNABEADS FOR SELECTIVE CONCENTRATION OF TARGET 5 40 16 APPENDIX BRIEF USER S 54 17 APPENDIX BEADRETRIEVER FEEDBACK FORM 55 18 APPENDIX D ADDRESSES wissiisscicsscssiscenscninssiesnssenes 56 BeadRetriever User Manual Rev 05 November 12 Table of Contents TABLE OF CONTENTS 1 SAFETY SYMBOLS AND 6 2 ABOUT THE USER MANUAL 7 3 INTRODUCTION TO THE 8 os Xsara tn AANE gudd A A E AR den in bI YDAW 8 32 8 3 3 ADVANTAGES OF PROCESSING MAGNETIC BEADS WITH BEADRETRIEVER 8 4 FUNCTIONAL DESCRIPTION 4 9 471 INSTRUMENTALAY OUT te DDd 9 4 1 1 9 4 1 2 Back WiGW 2s OR 10 4 1 3 caed chee AR 10 42 BEADRETRIEVER MAGNETIC BEAD
4. The keypad and display are shown in Fig 6 1 Fig 6 1 BeadRetriever keypad The keys available are two cursor keys or arrow keys direction keys to select the processing step to be run A To change to the previous next protocol by scrolling up Vv 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 BeadRetriever User Manual Rev 05 November 12 18 Chapter 6 Routine Operation 6 3 Howto START e First of all slide the tube strip tray out Fig 6 2 Fig 6 2 Loading the tube strip P tube strip contains one sample and the reagents for a single processing The user can select the position of the tube strip s on the tube strip tray The user must keep track of the sample order However the orientation of the tube strip is fixed The tube strips have to be positioned so that the slip ends face left Fig 6 3 and Fig 6 5 Note that the order in the tube strip of the sample and reagents is determined by the kit used Load only maximum fifteen special tube strips at a time into the tray for processing Fig 6 5 Slide the tray fully back in Push the tip comb all the way in ps z 2 GN Tubestriptray W Fig 6 3 Positioning of the tu
5. BeadRetriever User Manual Rev 05 November 12 36 Chapter 13 Index Installations ene E AE ere dd Fw DL a CN FAN SF y deters les 7 15 24 26 an eee dian eas 9 sade 8 Inverse magnetic particle processing sise 11 12 K 66 be Late Me L 8 11 18 54 4 18 5 aS 35 8 19 32 45 55 L Literature E EE AE E classed do E 35 6 ete ee 34 collectionOf ee Le WY stun WYR 12 12 12 13 8 10 11 12 13 22 25 28 29 4 Mains tn ee dda ae rte Pt dd te 23 25 26 22 24 RO dan 7 22 24 30 O
6. Further applications desired Where did you first learn about the product Would you like to receive information about other Invitrogen products BeadRetriever User Manual Rev 05 November 12 55 Appendix D Addresses 18 APPENDIX D ADDRESSES Invitrogen on the Internet For the latest information on the BeadRetriever as well as Dynabeads products and services visit our worldwide web site on the Internet at http www invitrogen com microbiology BeadRetriever manufactured by Thermo Fisher Scientific Oy P O Box 100 01621 Vantaa Finland Tel 358 9 329 100 Fax 358 9 3291 0415 www thermoscientific com Distributed by invitrogen United States European Asia Pacific Invitrogen Corporation Invitrogen Ltd Invitrogen Corporation 1600 Faraday Avenue 3 Fountain Drive Shanghai Representative Office Carlsbad California 92008 Inchinnan Business Park Room 4010 Grand Gateway Tower 1 Tel 1 760 603 7200 Paisley PA4 9RF UK No 1 Hongqiao Road Tel Toll Free 1 800 955 6288 Tel 44 0 141 814 6100 Shanghai 200030 P R C China Fax 1 760 603 7229 Fax 44 0 141 814 6260 Tel 86 21 6145 2000 Email techsupport invitrogen com Email eurotech invitrogen com Fax 86 21 6448 4077 Email sales cn invitrogen com BeadRetriever User Manual Rev 05 November 12 NOTES BeadRetriever User Manual Rev 05 November 12 57 NOTES
7. BeadRetriever User Manual Rev 05 November 12 58 NOTES BeadRetriever User Manual Rev 05 November 12 59
8. s Waste Electrical amp Electronic Equipment WEEE Directive 2012 19 EC 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 lt p gt BeadRetriever User Manual Rev 05 November 12 Chapter 2 About User Manual 2 ABOUT THE USER MANUAL This User Manual has been written for the actual user 0 laboratory technician and provides information on the BeadRetriever magnetic bead processor including the 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 Install the BeadRetriever e Use the BeadRetriever in routine jobs the processing step e Perform basic maintenance procedures This User Manual also describes all the features and specifications of the BeadRetriever hardware and on board software APPENDIX A Application Protocols for BeadRetriever using Dynabeads for selective concentration of target microorganisms 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 f
9. Human stool bovine faeces and environmental samples must be pre enriched for 24 h at 42 C Water samples 1 Filter 1 liter of water according to local standard procedures 2 Use a flat ended forceps to remove the filter and transfer directly into a wide mouthed bottle 3 Add 90 ml of BPW to the contents of the bottle and shake vigorously to dislodge bacteria from the surface of the membrane 4 Incubate at 42 C for 24 h 5 _ The use of filter aid is recommended for samples that are too turbid for membrane filtration BeadRetriever User Manual Rev 05 November 12 43 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms AIMS OPERATION PROCEDURE Users must aseptically dispense all reagents and samples into the tubes sequentially after the tube strips are fitted into the rack Users must read the nstructions for Use for each individual Dynabeads EPEC VTEC products before use as follows 1 Y E N o oo 11 12 Aseptically add 10 ul of properly mixed specific Dynabeads EPEC VTEC product into the two sample tubes 1 and 2 Aseptically add 500 ul of washing buffer PBST to sample tubes 1 and 2 Aseptically add 1 ml of washing buffer to tubes 3 and 4 within the strip Aseptically add 100 ul of washing buffer to the 5 tube Remove the desired tube from the sample rack and place in a second sample rack one meter away Add 500 ul of a test sa
10. 43 46 D DECONTAMINATION E A DF Y 2 21 23 24 30 31 Certificate A Nat EE Y FY A Gyda 24 30 31 DIMENSIONS onena Monet RE dure at dents sata 25 Directive d r a dent ner 26 20 terials ADHD 21 23 24 30 31 Disposal Of 23 5 7 29 35 40 41 43 44 46 48 49 0 DY Mal RT 5 42 44 E Environmental 15 Error message o 7 27 PAO ne EA 29 du cede 14 nL Lon 29 How install the BeadRetriever LLGC LLCC LL GL GL LLCC LU 14 Immunomagnetic separation 7 29 34 35 40 41 42 43 44 46 48 50 Hn 7 34 35 40 41 42 43 44 46 48 50 Ye EI i RR Carnac date dent da 8 12 13 25 46
11. 5 10 seconds and allow to dry Alternatively follow any standard procedure for disinfecting shell eggs Aseptically crack open the eggs and mix blend thoroughly both white and yolk Add Ferrous Sulphate FeSO4 solution to a final concentration of 35 mg L Pre incubate the egg mixture at 37 C for 6 hours After pre incubation mix the egg mixture thoroughly Dilute an aliguot five fold with wash buffer or buffered peptone water and use 1 ml of this dilution for IMS analysis Use a new pipette or a new pipette tip for each sample to avoid cross contamination Re incubate the remaining undiluted egg mixture overnight at 37 C N 8 9 AIMS OPERATION PROCEDURE Users must read the Instructions for Use for each individual Dynabeads anti Salmonella product before use as follows 1 Load one BeadRetriever sample tube strip for each sample into a sample rack 2 Dynabeads anti Salmonella by vortexing until the pellet in the bottom of the tube disappears and aseptically add 10 ul of properly mixed Dynabeads anti Salmonella into sample tubes 1 and 2 Note For the Shell Eggs method add 20 ul of properly mixed Dynabeads anti Salmonella into sample tubes 1 and 2 BeadRetriever User Manual Rev 05 November 12 46 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target N Note 11 12 13 14 Microorganisms Ase
12. can be used for both manual and automated protocols Before proceeding with automated immunomagnetic separation carefully read the operating instructions of the BeadRetriever and the nstructions for Use accompanying the Dynabeads anti Giardia and Dynabeads anti Cryptosporidium GC Combo Cat No 730 02 730 12 or the Dynabeads anti Cryptosporidium Cat No 730 01 730 11 products Details regarding buffers media other components as well as the protocols are provided with the Dynabeads anti Giardia and anti Cryptosporidium Instructions for Use SAMPLE PREPARATION This test has been designed for use with water samples produced and eluted via the standard method stated in the EPA method 1622 1623 and UK DETR Water Supply Regulation 2000 SI No 3184 England and 2001 SI No 3911 Wales For labs using the IDEXX Filta Max filtration system The protocols followed using the IDEXX Filta Max filtration system results in a 50 ml concentrate which is then centrifuged according to standard procedure After centrifugation the supernatant is removed via gentle aspiration to 3 ml volume The pellet size is recorded For labs using the Pall Envirochek or Envirochek HV filter capsule For protocols followed using the Pall Envirochek or Envirochek filter capsule filter and elute the sample as usual After centrifugation re suspend the pellet in 30 ml of buffer and transfer to a 50 ml conical tube Perform two additional rinses 10 ml each o
13. to allow adequate air circulation The BeadRetriever does not produce operating noise at a level that would be harmful No sound level measurements are required after installation Warning A 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 BeadRetriever 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 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 Warning This product contains strong permanent magnets 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 V The frequency range is 50 60 Hz 5 2 Setups before you put the instrument into operation 5 2 1 How to release the transport locks Tools 3 mm flat head screwdriver supplied with the instrument Note THERE ARE FOUR 4 TRANSPORT LOCKS SCREWS PRESENT Fig 5 2 A white tag with the red text CAUTION Remove Transport Lock Before Use is
14. LESHOOTING GUIDE 0 28 93 FREQUENTLY ASKED QUESTIONS FAQ ABOUT BEADRETRIEVER PROCESSING 29 9 4 SERVICE REQUEST 30 9 4 1 0 30 9 4 1 1 Certificate 31 10 WARRANTY CERTIFICAT Eai 32 10 1 WARRANTY CERTIFICATES Cd met Fy A dr 32 10 2 WARRANTY 32 11 ORDERING INFORMA T O N 33 11 1 LIST 2 su de date a dg YN Sd eas 33 1 1 2 MIST OF SPARE PARTS An eu Ag fre GY Yd Audit 33 11 3 LIST OF RECOMMENDED SPARE PARTS LLI LLE LL LLY YR LEL Y HEL HELI HEL LLED LA LL YF EDRYDD DH 33 12 GLOSSARY AND 34 12 1 KEYWORDS FOR WEB 35 12 2 eee 35 19 36 14 39 15 APPENDIX A APPLICATION PROTOCOLS FOR BEADRETRIEVER USING DYNABEADS FOR SELECTIVE CONCENTRATION OF TARGET MICROORGANISMS 40 AUTOMATED IMS USING DYNABEADS ANTI E COLI 157 41 AUTOMATED IMS USING DYNABEADS EPEC VTEC PRODUCT SERIES 0145 0111 0103 AND 26 43 AUTOMATED IMS U
15. NN nyw 18 61 18 6 2 HOW TO USE THE KEYPAD AND DISPLAY 18 6 9 SHOW TOMS TART 19 64 SHUTDOWN a 21 T 2 7 1 REGULAR AND PREVENTIVE 22 7 1 1 How to clean the tube strip 22 7 1 2 How to clean the magnetic rods 22 7 1 3 How to clean the processing chamber 22 1 27 DISPOSALIOR MATERIALS nm 0 23 7 3 DECONTAMINATION PROCEDURE iii 23 TA MOWTO PACK 2 24 1 5 7 SERVICE CONTRACTE 00 24 756 DISPOSAL OFTHE INSTRUMEN T mate cardan des O 24 8 25 8 1 GENERAL 8 8 4 4 4 erence 25 8 2 SAFETY SPECIFICATIONS 26 BeadRetriever User Manual Rev 05 November 12 Table of Contents 8 3 IN CONFORMITY WITH THE REQUIREMENTS iii 26 9 TROUBLESHOOTING 27 9 1 ERROR MESSAGES AND 8 27 92 TROUB
16. SING DYNABEADS ANTI SALMONELLA seen 46 AUTOMATED IMS USING DYNABEADS 48 AUTOMATED IMS USING DYNABEADS ANTI LEGIONELLA 2 YL YR FFF FL non 50 AUTOMATED IMS USING DYNABEADS ANTI GIARDIA AND 52 16 APPENDIX BRIEF USER S 11 54 17 APPENDIX BEADRETRIEVER FEEDBACK 55 18 APPENDIX 56 BeadRetriever User Manual Rev 05 November 12 Table of Contents Copyright Copyright 2012 Invitrogen Corporation First edition published in 2000 All rights reserved Reproduction of the accompanying user documentation in whole or in part is prohibited Patents The BeadRetriever has national and international patents as well as patents pending The production of Dynabeads is covered by various patents Trademarks and Dynal Dynabeads and SL are registered trademarks and BeadRetriever a trademark of Invitrogen Corporation Carlsbad California USA CHROMagar is a registered trademark of Dr A Rambach Paris France Decon is a registered trademark of Decon Laboratories Limited Envirochek is a registered trademark of Pall Co
17. TRIP TRAY an da 19 FIG 6 3 POSITIONING OF THE TUBE STRIP TRAY AND INSERTION OF THE COMB S 19 FIG 6 4 COMB SPECIALLY DESIGNED FOR THE 20 FIG 6 5 TUBE STRIP TRAY CONTAINING TUBES SPECIALLY DESIGNED FOR THE BEADRETRIEVER 20 FIG 15 1 SWAB STREAK TECHNIQUE u oss lds ie GRAU 44 FIG 16 1 BEAD RETRIEVER KEYPAD LM AY GM tn AW Y AGY eons 54 BeadRetriever User Manual Rev 05 November 12 39 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms 15 APPENDIX A Application Protocols for BeadRetriever using Dynabeads for selective concentration of target microorganisms Dynabeads based products for microbiology have been designed for both manual and automated immunomagnetic separation of specific target organisms from food feed and environmental samples Either process can selectively concentrate the specific target organisms from enriched sample matrices Before proceeding with automated immunomagnetic separation AIMS carefully read the operating instructions of the BeadRetriever and the user instructions accompanying each Dynabeads microbiology product The instrument currently supports only Dynabeads based products for the selective concentration of Salmonella Listeria Legionella speci
18. User Manual Rev 05 November 12 10 Chapter 4 Functional Description 42 BeadRetriever magnetic bead processor The BeadRetriever instrument Fig 4 5 is designed for maximum 15 tube strips which are compatible with the tip comb The tube strip 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 3 x 5 pieces and the other one for the plastic tip combs Fig 4 4 One tube strip tray contains fifteen separate tube strips and one sample uses one tube strip which is made up of five tubes One tip comb containing five tips is used for processing five samples at a time Before starting the magnetic bead processing via the keypad and display the samples and reagents are dispensed into the tubes within the tube strips and the tip comb s is are loaded into its their slot s The tube strip s is are placed into the removable tray in the correct position and the tray 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 inverse magnetic bead processing technology Fig 4 6 Rather than moving the liquids the magnetic beads are moved from tube to tube containing specific reagents in contrast to the external magnet met
19. Y GU OR 25 performances hihi ina ant en 25 Mn Hu 26 ei Ha RR a AL A MY 17 20 nnau E nm races YNA FYNY FYFYR FFF DY ADY MDF Y FN FYFYR FFY 6 T WES iea aa FF A 39 Tip 60 8 10 11 12 13 19 20 21 23 25 28 29 34 39 40 41 42 44 47 48 50 54 Transport ddai FN WD AR Rs 14 25 ydd TR GINI Y ydd 14 BeadRetriever User Manual Rev 05 November 12 37 Chapter 13 Index LOCK E EE 15 16 24 39 14 28 30 33 34 44 48 50 8 10 11 12 19 21 23 25 34 40 41 42 44 46 47 48 50 54 Tube strip tray 10 11 19 20 21 22 23 25 33 34 39 44 48 54 Un pack ahi eis 14 Fd 33 VIEW 9 10 uen dd 9 39 10 39 aH HN NET T 10 39 Voltage a
20. adRetriever User Manual Rev 05 November 12 35 Chapter 13 Index 13 INDEX A 2 34 y iio ICS 22750 2 HT run NR 14 33 Advantages of processing magnetic particles sisi 8 NI AYN I NE A HEN FN Y FYN yN 7 29 34 35 40 41 44 46 48 50 ET ER FYNY 5 6 7 40 55 Application HAR RA RA FA Y teu Re 7 40 Automated IMS using Dynabeads anti E 0157 41 Automated IMS using Dynabeads anti Legionella 50 Automated IMS using Dynabeads 2 48 Automated IMS using Dynabeads anti Salmonella 100 46 50 Automated IMS using Dynabeads EPEC VTEC product series 0145 011 0103 and O26 43 Automated immunomagnetic 7 29 34 35 40 41 43 44 46 48 50 B BeadRetriever 5 7 8 9 10 11 14 15 16 18 20 21 22 29 30 32 33 35 39 40 41 43 46 47 48 50 51 54 dd 28 15 22 24 29 magnetic rods aga eee ee 22 processing Chambers Muse then me ai ne te RAY 22 Re ES 7 12 13 39 40
21. amiliar with the contents of Chapter 7 on maintenance For warranty and ordering information refer to Chapter 10 WARRANTY and Chapter 11 ORDERING INFORMATION A CD ROM video demonstrating use of the BeadRetriever and automated IMS AIMS is available to all users Please request a CD by email environmental invitrogen com or visit www invitrogen com microbiology BeadRetriever User Manual Rev 05 November 12 Chapter 3 Introduction to BeadRetriever 3 INTRODUCTION TO THE BEADRETRIEVER 3 1 Intended use The BeadRetriever magnetic bead processor Fig 3 1 is intended for professional research and laboratory use by trained personnel The instrument is intended for automated transfer and processing of magnetic beads in a 1 ml tube scale Use for self testing is excluded 3 2 Principle of operation The principle of the BeadRetriever system patents pending is based on the use of magnetic rods covered with disposable specially designed tip combs and tube strips The instrument functions without any dispensing or aspiration parts or devices Samples and reagents including magnetic beads are dispensed into the tube strips according to the corresponding kit instructions The steps of the protocol that are selected by the user via the keypad and display have already been preloaded in the on board software Fig 3 1 BeadRetriever magnetic bead processor 3 3 Advantages of processing magnetic beads with BeadRetriever The Bea
22. are available three days after receipt of samples 1 2 Transfer the concentrated resuspended bead bacteria complexes into 10 ml of Rappaport Vassilliadis Soya peptone broth RVS and incubate at 42 C for 18 24 hours Follow standard procedure for isolation by spreading a loopful of RVS culture onto any Salmonella plating media such as Brilliant Green agar BGA Xylose Lysine Deoxycholate agar XLD Bismuth Sulphite agar BSA Hektoen agar HE etc The RAPID Method This is recommended for processed foods known to harbour none or low levels of background flora only Presumptive Sa monella positive results are available two days after receipt of samples 1 2 Transfer 50 ul of the resuspended bead bacteria complex onto each of two Salmonella selective agar plates BGA XLD BSA etc Interpretation of presumptive results depends on the skill of the user to correctly differentiate the isolated colonies based on typical Salmonella morphology Suspect colonies must be confirmed by standard biochemical and serological test methods BeadRetriever User Manual Rev 05 November 12 47 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms Automated IMS Using Dynabeads anti Listeria Dynabeads anti Listeria has been designed for use in both manual and automated immunomagnetic separation of Listeria species from food feed and environmental samples Ei
23. at a total sample volume of 2 ml would be analyzed with a resultant increase in the number of target organisms recovered This approach might be useful when performing tests where the number of target organisms might be limiting due to competition or a prolonged lag phase due to sub lethal injury In order to avoid cross contamination of the prepared tubes it is recommended that transfer of samples be performed in a designated area at least one meter from the prepared tubes It is important that filtered pipette tips are used to transfer samples into the test tubes It is recommended that the tip combs be removed not less than 10 minutes after the assay has been completed to allow for air drying before removal Washing buffer PBS Tween 0 15M NaCl 0 01M Sodium Phosphate buffer pH 7 4 with 0 05 Tween 20 Autoclavable at 121 C for 15 minutes can be stored under refrigeration Available in powder form from Sigma Aldrich No P3563 2 A filter homogenizer bag removes particulate material and fatty substances which are inhibitory to IMS For certain foods for example meat with bones or dry pasta a blender is preferred prior to using a homogenizer bag to avoid the risk of perforation After blending the contents should be transferred into a filter homogenizer bag or wide screw cap bottle BeadRetriever User Manual Rev 05 November 12 40 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentrat
24. bacteria complexes 2 Dilute further by streaking with a loop Always carry the loop back into the previously streaked quadrant several times to ensure that the beads reach a fresh unstreaked quadrant 3 Incubate the plates at 35 37 for 18 24 h Read the plates for suspect coli 0157 colourless colonies on CT SMAC and pink mauve coloured colonies on CHROMagar 0157 The choice of plating media has been based on some distinct characteristics of E coli 157 H7 It is the only E coli among clinical isolates which does not ferment sorbitol within 24 h and which is glucuronidase negative The organisms are resistant to potassium tellurite and cefixime Presumptive coli 0157 colonies must be confirmed by standard biochemical and serological testing BeadRetriever User Manual Rev 05 November 12 42 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms Automated IMS Using Dynabeads EPEC VTEC Product Series 0145 0111 0103 and O26 The Dynabeads EPEC VTEC Product Series is specifically developed for rapid selective concentration of non O157 Escherichia coli serotypes 0145 0111 0103 and O26 using the BeadRetriever from food feed and environmental samples The products may also be used to perform manual IMS Either process can selectively concentrate the targeted 157 coli serotypes from 24 pre enriched sample matrices at 42 C Before proceedi
25. be strip tray and insertion of the tip comb s 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 tube strip or one to fifteen tube strips depending on the amount of samples At least one tip comb Fig 6 4 always has to be inserted into one of the narrow slots meant for the tip comb BeadRetriever User Manual Rev 05 November 12 19 Chapter 6 Routine Operation Fig 6 4 Tip comb specially designed for the BeadRetriever e Close the front lid if you want to The front lid protects the instrument against environmental contamination Note front lid and or top lid can be left open if desired This action does not break the run Select the name of the method e Use the START key to initiate the run For startup refer to Section 6 1 Switching on Note At the end of the run make sure that the text End of run is shown on the display and then press the STOP key Note samples are collected immediately after the run and transferred into other tubes for longer storage Slip end faces left Fig 6 5 Tube strip tray containing tubes specially designed for the BeadRetriever BeadRetriever User Manual Rev 05 November 12 20 Chapter 6 Routine Operation 6 4 Shutdown Switch the BeadRetriever off by pressing the power switch Fig 4 3 at the bottom left of the back panel of the instrument into the OFF positi
26. cacia OAH dy A CA 15 17 18 25 26 W Warnin 5 2 2 15 16 17 27 39 Warranty wd FA FLY 7 14 32 15 25 BeadRetriever User Manual Rev 05 November 12 38 Chapter 14 Table of Figures 14 TABLE OF FIGURES FIG 3 1 BEADRETRIEVER MAGNETIC BEAD PROCESSOR 0 0 8 FIG 4 1 BEADRETRIEVER FRONT VIEW WITH 9 9 FIG 4 2 BEADRETRIEVER FRONT VIEW WITH LIDS OPEN ee 9 FIG 4 3 BEADRETRIEVER REAR Sah OR 10 FIG 4 4 BEADRETRIEVER INTERNAL VIEW ois LR Run tan un A AU RA AFF Fd 10 FIG 4 5 BEADRETRIEVER MAGNETIC BEAD 0 11 FIG 4 6 INVERSE MAGNETIC BEAD PROCESSING sssscsssscscsesssesesesesssecsesssesaesesssassenecececsessnscaesenececeenenens 12 FIG 4 7 A CONCENTRATION STEP DURING MAGNETIC BEAD PROCESSING scscsssscscscsesessvscsesseecseseeecacsesenaes 13 FIG 5 1 BEADRETRIEVER TRANSPORT LOCK TAG PRESENT BEFORE RELEASE OF THE 16 FIG 5 2 LOCATION OF THE TRANSPORT LOCK SCREWS AND WARNING TAG re 16 FIG 5 3 CONNECTING THE MAINS SUPPLY CABLE 17 FIG 6 1 BEADRETRIEVER KEYPAD in i ns 18 FIG 6 2 LOADING THE TUBE S
27. d 33 ul of Dynabeads anti Giardia to each tube 1 3 9 Insert the protective tip combs into the BeadRetriever 10 Insert the rack containing the tube strips into the BeadRetriever 11 Check that everything is properly aligned and close the instrument door IRON BeadRetriever User Manual Rev 05 November 12 52 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target 12 13 14 15 16 Microorganisms Select the GC Combo program by scrolling with the arrow keys and then press the START button During instrument operation the door should be kept closed at all times The display will show the time left for each stage of the automated IMS protocol The stages of the GC Combo protocol in order are Mixing stage Bead isolation Wash stage and then dissociation stage After the dissociation stage the machine will beep once finished At the end of the program run open the instrument door and carefully remove the rack containing the sample tube strips POST AUTOMATED IMS PROCESS 1 N Remove each sample tube strip one at a time and carefully remove the 100 ul 0 1 N HCL from tube 5 Spot this 100 ul onto Dynal Spot On Cat No 740 04 slides containing 10 pl 1N NaOH Repeat this for all samples processed onto separate slides per sample Then incubate the slides according to standard procedure 30 min at 41 C Once dry then fix the
28. d tube strips and discard into an appropriate biohazardous waste container and autoclave prior to disposal Safety Note During operation the BeadRetriever does not create a biohazardous aerosol and may be operated safely on the laboratory bench For further information see http Www hpa org uk srmd div_esl_su pdf_guidance2 htm ISOLATION PROCEDURE Plate the bead bacteria complexes from tube 4 onto an appropriate Legionella selective media for example GVPC agar using a standard plating technigue and according to local procedures Culture plates should then be incubated according to standard procedures CONFIRMATION Interpretation of presumptive results depends on the skill of the user to correctly differentiate the isolated colonies based on the typical Legionella morphology Suspect colonies must be confirmed by standard biochemical and serological test methods BeadRetriever User Manual Rev 05 November 12 51 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms Automated IMS using Dynabeads Anti Giardia and Anti Cryptosporidium Dynabeads anti Giardia and anti Cryptosporidium have been designed to be used for the isolation and detection of Giardia cysts and Cryptosporidium oocysts from environmental water samples These immunomagnetic beads can be used to simultaneously isolate both Giardia and Cryptosporidium or to isolate Cryptosporidium only These beads
29. dRetriever magnetic bead processor provides several advantages relating mainly to the principle of operation in that it e Enables automation of complicated manual steps e Enables simultaneous processing and purification e Enables a quicker reaction and a more efficient wash due to the simultaneous shaking during incubation and washing e Facilitates a good collection of bead bound sample due to the efficiency of the magnet e By moving the beads from tube to tube during processing contaminating background flora is reduced further e Prevents carryover due to the fact that the same tip is only used for one sample e Facilitates the whole processing with the aid of an internal program BeadRetriever User Manual Rev 05 November 12 Chapter 4 Functional Description 4 FUNCTIONAL DESCRIPTION 41 Instrument layout 4 1 1 Front view e Keypad Display Front lid Top lid ON Fig 4 1 BeadRetriever front view with lids closed Top lid Fig 4 2 BeadRetriever front view with lids open BeadRetriever User Manual Rev 05 November 12 Chapter 4 Functional Description 4 1 2 Back view 1 ON OFF switch 2 Main power supply socket 3 USB 4 Serial port RS 232C Fig 4 3 BeadRetriever rear view 4 1 3 Internal view 1 Tube strip tray with tube strips 2 comb holder 3 comb slot with tip comb 4 Magnetic rod Fig 4 4 BeadRetriever internal view BeadRetriever
30. e bag firmly and leave the instrument in the bag for at least 24 h BeadRetriever User Manual Rev 05 November 12 23 10 11 7 4 Chapter 7 Maintenance Remove the instrument from the bag Clean the instrument using a mild detergent Remove any stains using 70 ethanol After performing this decontamination procedure label the instrument with a signed and dated Certificate of Decontamination How to pack for service When you ship the instruments for service remember to Inform about the use of hazardous materials Decontaminate the instrument beforehand Install the transport locks 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 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 Enclose the return authorization number RGA given by the Invitrogen representative Indicate the fault after you have been in touch with your local Invitrogen representative See Section 8 1 for details on storage and transportation temperatures 7 5 Service contracts It is recommended that you maintain and service the instrument regularly every twelve 12 months on a contract basis by the manufacturer s trained service engineer
31. ects the magnetic rod A disposable tip comb always has to be fitted into the tip comb holder slot during processing TBST Tris buffered saline with Tween e g TBST BSA TSB Tryptone Soya Broth e g mTSB modified Tryptone Soya Broth Tube strip The disposable tubes where all the reagents and samples are located and where the processing takes place Tube strip tray Tube rack A removable tube strip tray or tube rack that holds 15 tube strips Fig 6 3 and Fig 6 5 VCC Vancomycin Cefsulodin Cefixime VTEC Verocytotoxin producing Escherichia coli XLD Xylose Lysine Desoxycholate medium BeadRetriever User Manual Rev 05 November 12 34 Chapter 12 Glossary and Abbreviations 12 1 Keywords for web pages AIMS magnetic beads automated IMS magnetic separation BeadRetriever processing Cryptosporidium processor Dynabeads purification 0157 Salmonella EPEC VTEC separation IMMUNOMAGNETIC SEPARATION separator Giardia superparamagnetic beads IMS Thermo Fisher Scientific Listeria Thermo Scientific 12 2 Literature A collection of articles illustrating the use of Dynabeads based IMS protocols for the isolation of bacteria and protozoa is available at our website These articles include protocols for E coli 0157 from Meat E coli 0157 amp other EPEC VTECs Giardia Listeria Salmonella spp Salmonella and Eggs Cryptosporidium Visit our website at www invitrogen com microbiology Be
32. einvitrogen BeadRetriever BeadRetriever User Manual User Manual Rev 05 November 12 Product No 159 50 Table of Contents BeadRetriever User Manual Rev 05 November 12 Product No 159 50 CONTENTS 1 SAFETY SYMBOLS AND 6 2 202 7 3 INTRODUCTION TO THE BEADRETRIEVER 11 1 8 4 FUNCTIONAL DESCRIPTION 222 00 Run YN ans 9 5 INSTALLATION 14 6 ROUTINE OPERATION riscssccissssscscssatssscnsassesensnccscndnansseenneisaaccuaansecnnadisansaabasaensaasiannteaaasunnaansannneanas 18 7 MAINTENANCE 05 22 8 TECHNICAL SPECIFICATIONS DYNA DAN YN 25 9 TROUBLESHOOTING 27 10 WARRANTY 6 32 11 ORDERING INFORMATION fiiscsinsstsecsasecsssninsnsasnacsnacssnuinsasnastsaassesvensssnnbinancdavunsasnsotnnacnsduanstannueneeds 33 12 GLOSSARY AND ABBREVIATIONS 77 LL LL 34 13 5 lt
33. elease of Ready magnetic particles low volume well magnetic particles suspension in a from a large volume low volume well well Fig 4 7 A concentration step during magnetic bead processing BeadRetriever User Manual Rev 05 November 12 13 Chapter 5 Installation 5 INSTALLATION 51 What to do upon delivery 5 1 1 How to unpack Move the unpacked instrument to its site of operation Unpack the BeadRetriever 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 available when you open the package the packing instructions packing list the BeadRetriever Feedback Form the Warranty Certificate card the 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 pa
34. er DO NOT expose painted surfaces to 4 Painted surfaces cleaned with most laboratory detergents Dilute the cleaning 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 solution should be used 4 DO NOT autoclave of this instrument except for the tube strip tray 7 1 1 How to clean the tube strip tray Keep the tube strip tray surface clean to avoid dust and dirt entering into the instrument Clean the tray 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 tube strip tray you can autoclave the tray 7 1 2 How to clean the magnetic rods If required wipe the magnetic rods using a soft cloth or tissue paper soaked in a mild detergent solution 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 BeadRetriever User Manual Rev 05 November 12 22 Chapter 7 Maintenance 7 2 Disposal of materials Refer to local
35. eria complexes Dilute further by streaking with a loop Always carry the loop back into the previously streaked quadrant several times to ensure that the beads are streaked into a fresh unstreaked quadrant See Fig 15 1 Incubate the plates at 35 37 C for 18 24 h Proceed to the next section 5 ww 5 1 Swab 2 Streak with a loop 3 New loop and streak Fig 15 1 Swab streak technique BeadRetriever User Manual Rev 05 November 12 44 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms PRESUMPTIVE IDENTIFICATION AND CONFIRMATION OF NON O157 VTEC 1 oo 10 Add 10 ul of physiological saline onto a glass slide placed on a dark background Two or three tests may be performed on one slide Transfer a sweep of mixed growth from the first half of the blood agar plate or a single colony onto the slide and make a smooth milky suspension Observe for autoagglutination In the absence of any autoagglutination add 10 ul of the specific OK EPEC VTEC antiserum provided in the Colony Verification kit to the suspension and mix well Observe for agglutination by tilting the slide for 1 30 seconds If autoagglutination occurs test one to five distinct individual colonies see step 7 below A visible agglutination reaction within 30 seconds is a strong
36. erials 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 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 1 Please include decontaminating solution used BeadRetriever User Manual Rev 05 November 12 31 Chapter 10 Warranty Statement 10 WARRANTY CERTIFICATE 10 1 Warranty Certificate The BeadRetriever is fully guaranteed against defective parts and materials including defects caused by poor workmanship for a period of one year from the date of delivery Invitrogen 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 Invitrogen Instructions for Use 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 Invitrogen subsidia
37. es Cryptosporidium Giardia and the verocytotoxin producing Escherichia coli VTEC serotypes 0157 0145 0111 0103 and O26 The specially designed disposable tip combs and tube strips that are needed to use the instrument must be purchased separately These disposable plastic components can be autoclaved before use It is recommended that the tube strips should be autoclaved before use The tips are designed for easy insertion into the slots in the instrument in one direction only The tubes are designed as a strip containing five tubes per test sample for easy insertion into the designated rack and subsequently into the instrument Both tip combs and tubes are designed for one time use only and should be disposed of into a biohazardous waste container Application Note Notice that only 500 ul of the enriched samples is to be added to 500 ul of washing buffer PBST in each of the two sample tubes This will result in a twofold dilution of the 1 ml sample for analysis This is done to achieve a test sample from samples that are fatty viscous and or particulate from which bead recovery would otherwise have been difficult This twofold dilution results in an optimal sample preparation to achieve optimal bead recovery while performing AIMS Samples that are otherwise not fatty viscous and or particulate could be analyzed without any such dilution In this case only 1 ml of the pre enriched sample should be added to the sample tubes This implies th
38. f the centrifuge tube to recover all remaining sediment Centrifuge the sample again at 1500 x G Allow the rotator to coast to a stop do not use the brake Re record the pellet volume on the bench sheet Proceed as described above for the Filta Max filter If the pellet is above 0 25 ml then split the sample into sub samples respectively for example if the pellet is 0 5 ml split into two sub samples of 0 25 ml pellet each The 3 ml concentrate is then vortexed thoroughly to re suspend the pellet AIMS OPERATION PROCEDURE Place one disposable sample tube strip into a BeadRetriever rack for each sample to be processed The tab on the tube strip may be used for sample labeling Users must read the nstructions for Use for Dynabeads anti Giardia and anti Cryptosporidium before use as follows To tubes 1 3 add 100 ul of 10X SL Buffer A and 100 ul of 10X SL Buffer To tube 4 add 1 ml of 1X SL Buffer A To tube 5 add 100 ul of 0 1 N HCL Split the 3 ml sample concentrate across tubes 1 3 1 ml into each tube 1 3 Add up to 300 yl RO water to the sample concentrate tube Vortex the concentrate tube to re suspend and wash any debris left behind This produces the rinsate Split the rinsate across tubes 1 3 up to 100 ul to each tube 1 3 Add 33 ul of Dynabeads anti Cryptosporidium to each tube 1 3 For Cryptosporidium only detection proceed to step 9 8 For dual Giardia and Cryptosporidium detection within the same assay also ad
39. fastened to the instrument to BeadRetriever User Manual Rev 05 November 12 15 Chapter 5 Installation 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 locks are situated Remove the tag and release the transport locks Ensure that the transport locks have been released before you put the instrument into operation Fig 5 1 BeadRetriever transport lock tag present before release of the locks 1 remove the transport locks unscrew the four 4 screws Fig 5 2 with the 3 mm flat head screwdriver supplied Make sure that the screws are fully unscrewed in the opposite direction 2 Then remove the transport lock warning tag Fig 5 1 3 Save the transport lock warning tag for future transportation Tag attached here Fig 5 2 Location of the transport lock screws and warning tag BeadRetriever User Manual Rev 05 November 12 16 Chapter 5 Installation 5 2 2 How to ensure 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 you need to use any other type of mains supply cable than supplied 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
40. fety 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 to 40 C Humidity Maximum relative humidity 80 for temperatures up to 31 C decreasing linearly to 50 relative humidity at 40 C Mains supply fluctuations 10 from nominal Installation category overvoltage category ll 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 CAT 1 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 BeadRetriever User Manual Rev 05 November 12 26 9 TROUBLESHOOTING GUIDE 9 1 Error messages and warnings E
41. he tab on the tube strip may be used for labeling samples Users must read the Instructions for Use for Dynabeads anti Legionella before use as follows Add 100 ul of Sample buffer to sample tube 1 Vortex Dynabeads anti Legionella for a minimum of 10 seconds to mix Add 50 ul of Dynabeads anti Legionella into sample tube 1 Add 1 ml of Wash buffer to tubes 2 and 3 within the strip Add 100 ul of Wash buffer to tube 4 Add 1 ml of sample aliquot to tube 1 to avoid cross contamination between samples it is recommended that sample transfer is performed at least 1 meter from the other prepared tube strips 7 Insert the sterile disposable protective tip combs into the instrument Each comb should click into position Insert the rack with filled tubes into the correct position in the instrument Check that everything is properly aligned and close the instrument door 0 Select the ENVIRONMENTAL program sequence by scrolling with the arrow key and press the START button 11 While the instrument is in operation the door must be kept closed Each processing step and the total time remaining can be followed on the LC display 12 Atthe end of the program run remove tube rack from the instrument 5 ON 0 00 BeadRetriever User Manual Rev 05 November 12 50 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms 13 Remove the tip combs an
42. here is nothing preventing the head from moving up and down 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 BeadRetriever User Manual Rev 05 November 12 28 Chapter 9 Troubleshooting Guide 93 Frequently asked questions FAQ about BeadRetriever processing Q1 What does the BeadRetriever do A1 Dynabeads based products for microbiology have been designed for automated immunomagnetic separation AIMS of these target organisms from food feed and environmental samples The process can selectively concentrate the specific pathogenic microorganisms from sample materials Invitrogen now offers a complete BeadRetriever system the magnetic bead processor and the reagents for the purification of microbial samples The processor handles beads automatically according to the preloaded protocol 02 Do need a to control the processor A2 The protocols are preloaded by Invitrogen and no PC is needed for routine use A PC program can be used to create new protocols or modify existing protocols if needed PC
43. hod Magnetic beads are transferred with the aid of magnetic rods covered with disposable specially designed plastic tip combs H 1 Fig 4 5 BeadRetriever magnetic bead processor BeadRetriever User Manual Rev 05 November 12 11 Chapter 4 Functional Description 4 3 Principle of magnetic bead processing Magnetic rod Transfer Magnetic bead Collection of Magnetic beads Release of Ready suspension before magnetic beads on the tip surface magnetic beads suspension collection Fig 4 6 Inverse magnetic bead processing 4 4 Working with a magnetic rod Working with magnetic beads can be divided into five separate processes collecting beads releasing beads washing beads incubation concentration 4 4 1 Collecting magnetic beads During the collection of the magnetic beads the magnetic rod is fully inside the tip The magnetic rods together with the tip comb s move slowly up and down in the tubes within the tube strip and the magnetic beads are collected onto the wall of the tips The magnetic rod together with the tip comb having collected the magnetic beads will lift out of the tube and be transferred into the next tube within the same tube strip 4 4 2 Releasing magnetic beads After collection of the magnetic beads the magnetic rods together with the tip comb s are lifted from the tubes the magnetic rods are lifted up and the tip comb s is are lowered into the next tube in
44. indication of a presumptive positive sample When testing a sweep of mixed growth confirm the initial presumptive result by testing one to five distinct individual colonies from the blood agar plate in a similar manner using the specific OK antiserum If no distinct colonies could be picked plate further for purity from the other plating media onto blood agar and proceed as described above The reaction is read with the naked eye by holding the slide in front of a light source against a black background i e indirect illumination A positive reaction is seen as a visible agglutination A negative reaction is persistence of the homogeneous milky turbidity A late or weak agglutination should be considered negative Plate the agglutination positive colonies further for purity and confirm them by standard biochemical serological and DNA tests e g PCR BeadRetriever User Manual Rev 05 November 12 45 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms Automated IMS Using Dynabeads anti Salmonella Dynabeads anti Salmonella has been designed for use in both manual and automated immunomagnetic separation of Salmonella from food feed and environmental samples Either process can selectively concentrate Salmonella from enriched sample matrices Before proceeding with automated immunomagnetic separation carefully read the operating instructions of the BeadRetrieve
45. ion of Target Microorganisms Automated IMS Using Dynabeads anti E coli 0157 Dynabeads anti E 0157 has been designed for use in both manual and automated immunomagnetic separation of E coli 0157 from food feed and environmental samples Either process can selectively concentrate E coli 0157 from enriched sample matrices Before proceeding with automated immunomagnetic separation carefully read the operating instructions of the BeadRetriever and the nstructions for Use accompanying the Dynabeads anti E coli 0157 product Details regarding buffers media other components as well as the protocols are provided with the Dynabeads anti E coli 0157 Instructions for Use SAMPLE PREPARATION Food samples 1 _ Weigh 25 g of food sample and place into a filter homogenizer bag 2 _ Add 225 ml of the desired enrichment medium e g BPW TSB or BGBB and homogenize Incubate for 6 18 h at 37 C or 41 5 C 3 the pre enriched sample thoroughly by homogenizing once more Human stool bovine faeces and environmental swab samples 1 For human animal stool samples prepare 10 suspension physiological saline and transfer 1 ml into 10 ml of a suitable enrichment broth 2 Human rectal swab and environmental swab samples should be transferred into 10 ml of a suitable enrichment broth 3 Enrichment broths should be incubated as described above for food samples Water samples 1 Filter 1 liter of water according
46. local Invitrogen representative will keep you up to date with the progress of service and provide you with any further details you might need e g on maintenance serviceability troubleshooting and replacement 941 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 BeadRetriever User Manual Rev 05 November 12 30 Chapter 9 Troubleshooting Guide 9 4 1 1 Certificate of Decontamination The decontamination procedure is required prior to shipping the instrument to Invitrogen e g for repair If the instrument is shipped back to Invitrogen 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 body fluids toxic carcinogenic or radioactive mat
47. magnetic beads are attached to the tip combs This happens sometimes but it will not affect the yield because the sample has been released from the beads Q8 Are the volumes of reagents in each well critical A8 It is strongly recommended that you keep the given volumes within certain limits to avoid failure in performance of the chemical reactions and the processor BeadRetriever User Manual Rev 05 November 12 29 Chapter 9 Troubleshooting Guide 9 4 Service request protocol If the BeadRetriever requires service contact your local Invitrogen representative 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 Invitrogen representative will prepare a complaint form Complaint order containing a detailed description of the fault symptom or condition based on the information you give them Check Section 7 4 How to pack for service 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 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 Your
48. ment Insert the rack with filled tubes into the instrument to lock it in place Check that everything is properly aligned and close the instrument door 0 Select the EPEC VTEC program sequence by scrolling with the arrow key and press the START button DO BeadRetriever User Manual Rev 05 November 12 41 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms 11 While the instrument is in operation the door must be kept closed Each processing step and the total time remaining can be followed on the LC display 12 At the end of the program run remove the tube s rack from the instrument and plate one half of the bead bacteria complexes from the 5 tube onto each of the appropriate plating media as recommended below 13 Remove the tip combs and discard into a biohazard waste container together with the tube strips ISOLATION PROCEDURE After IMS transfer the resuspended beads onto each internationally accepted E coli 0157 culture media plates It is recommended that two different culture media are used to increase the chances of detecting suspect colonies that have distinct differential features on each media Invitrogen recommends Sorbitol MacConkey SMAC media supplemented with CT supplement CHROMagar 0157 1 Spread the bead bacteria complexes over one half of the plate with a sterile swab This ensures the break up of the bead
49. 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 BeadRetriever 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 e g if they are mechanically damaged during runs Local technical personnel can carry out changing the magnets Q4 How strong are the magnets Can they disturb e g some sensitive equipment A4 The BeadRetriever should not be kept in close proximity to magnetic tapes computer discs or other magnetic storage systems such as credit cards as these can be damaged by the strong magnetic field of the BeadRetriever heads Do not hold the BeadRetriever heads close to a PC display since this may cause damage to the display Do not use metal tools when handling BeadRetriever heads Q5 What happens if forget to insert the tip combs into the tip comb holder and the magnetic beads stick to the magnetic rods A5 Wipe the magnetic rods using a soft cloth or tissue paper soaked in a mild detergent solution soap solution or alcohol Q6 What if the magnetic beads remained in the well A6 If the starting material is too viscose the magnetic rods will not be able to collect the beads Dilute the sample and check that the sample is properly homogenized lysed Q7 What if
50. mple to tubes 1 and 2 and transfer the inoculated tube to the first sample rack Repeat for the remaining samples Aseptically insert the sterile protective tip combs into the instrument Insert the rack with filled tubes into the instrument to lock it in place Check that everything is properly aligned and close the instrument door Select the EPEC VTEC program sequence by scrolling with the arrow key and press the START button While the instrument is in operation the door must be kept closed Each processing step and the total time remaining can be followed on the LC display At the end of the program run remove the tube rack i e tube strip tray from the instrument and plate one half of the bead bacteria complexes from the 5 tube onto each of the appropriate plating media as recommended below Remove the tip combs and discard into a biohazardous waste container together with the tube strips ISOLATION OF EPEC VTEC OTHER THAN 0157 VTEC After automated IMS transfer all the resuspended bead bacteria complexes onto blood agar or one half of the bead bacteria complexes onto blood agar and the remaining half onto anyone of the following plating media CHROMagar 0157 Invitrogen MacConkey agar modified Haemorrhagic Colitis agar mHC Eosin Methylene Blue EMB and Chromocult Coliform Agar Merck 1 Spread the bead bacteria complexes over one half of the plate with a sterile swab This is to ensure the break up of the bead bact
51. ng with automated immunomagnetic separation carefully read the operating instructions of the BeadRetriever and the Instructions for Use accompanying each Dynabeads EPEC VTEC product Details regarding buffers media other components as well as the protocols are provided with the Dynabeads EPEC VTEC product Instructions for Use SAMPLE PREPARATION Food samples 1 Weigh 25 g of food sample and place into a filter homogenizer bag 2 Add 225 ml of Buffered Peptone Water BPW 3 Incubate at 42 C for 24 h 4 Mix the pre enriched sample thoroughly by homogenizing once Human stool bovine faeces and environmental swab samples Whole stool specimens must be refrigerated as soon as possible after collection and must be examined within 1 2 h of collection If they cannot be examined within 1 2 h whole stool samples or a swab of the stool or rectal swabs should be placed in a transport medium e g Stuart s Cary Blair etc and held refrigerated until examination within 2 3 days If samples are held longer than 3 days before examination they should be frozen at 70 C Specimens in transport medium should not be left at ambient temperature 1 _ Transfer 1 ml of human liquid stool sample into 10 ml of BPW For solid human stool samples and bovine faeces prepare a 10 suspension in physiological saline and transfer 1 ml into 10 ml BPW 3 Human rectal and environmental swab samples should be transferred into 10 ml of BPW 4
52. o tube 3 within the strip Aseptically add 100 ul of washing buffer to the 5 tube For each sample remove the labelled sample tube strip from the sample rack and place in a second sample rack one meter away Add 500 ul of a test sample to tubes 1 and 2 and transfer the inoculated tube to the first sample rack Repeat for the remaining samples Aseptically insert the sterile protective tip combs into the instrument Insert the rack with filled tubes into the instrument to lock it in place Check that everything is properly aligned and close the instrument door Select the LISTERIA program sequence by scrolling with the arrow key and press the START button 10 While the instrument is in operation the door must be kept closed Each processing step and the total time remaining can be followed on the LC display 11 At the end of the program run remove the tube rack i e tube strip tray from the instrument and plate one half of the bead bacteria complexes from the 5 tube onto each of the appropriate plating media as recommended below 12 Remove the tip combs and discard into a biohazardous waste container together with the tube strips ONO BeadRetriever User Manual Rev 05 November 12 48 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms ISOLATION PROCEDURE The resuspended Dynabeads bacteria complexes are now ready for plating Tran
53. ommended spare parts Code Item 1 2 5 10 units year year 2104 0820 H motor assembly 2104 0850 assembly 2407 0740 Free wheel assembly 1 2407 2550 Magnetic plate assembly 1091910 Tooth belt T2 5 330 1 1092250 Tooth belt T2 5 265 1 2004460 PSP 01 1 2004470 CSU 02 1 2 2004480 CSU 01 1 21040811 Power supply 1 2 1 1 3 1 Contact your local Invitrogen representative BeadRetriever User Manual Rev 05 November 12 33 Chapter 12 Glossary and Abbreviations 12 GLOSSARY AND ABBREVIATIONS AIMS Automated immunomagnetic separation BGA Brilliant Green Agar BPW Buffered Peptone Water BRILA Brilliant Green Bile Lactose Broth BSA Bismuth Sulfite Agar or Bovine Serum Albumin CT Cefixime Tellurite e g CT supplement and CT SMAC EMB Eosin Methylene Blue EPEC Enterotoxin producing Escherichia coli GN Gram negative e g GN broth HC Haemorrhagic Colitis Agar e g mHC modified Haemorrhagic Colitis Agar HE Hektoen Enteric Agar IMS Immunomagnetic separation Magnetic rod The rods that are magnetic and collect magnetic beads The rods do not collect the beads on their own the magnetic rods must always be protected by a tip comb PBST PBS Tween Phosphate Buffered Saline containing 0 05 Tween 20 RVS Rappaport Vassilliadis Soya peptone broth SMAC Sorbitol MacConkey media Tip comb Prot
54. on Remove any tube strips still on the tube strip tray Dispose of all tube strips as biohazardous waste 0 Remove any combs still in the instrument Dispose of combs biohazardous waste Wipe the tray surface and the adjacent 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 tray disinfect with 70 alcohol or some other disinfectant see Section 7 3 Decontamination procedure BeadRetriever User Manual Rev 05 November 12 21 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 It is recommended that 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 tube strip tray with clean low pressure compressed air or a cloth dampened with water or a mild detergent when necessary Although the BeadRetriever 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 agent as recommended by the manufactur
55. ptically add 500 ul of wash buffer to sample tubes 1 and 2 Aseptically add 1 ml of wash buffer to tubes 3 and 4 within the strip Aseptically add 100 ul of wash buffer to tube 5 For each sample remove the labelled sample tube strip from the sample rack and place in a second sample rack one meter away Add 500 ul of the test sample to tubes 1 and 2 and return the inoculated tube to the first sample rack Repeat for the remaining samples Aseptically insert the sterile protective sample tip combs into the instrument Insert the rack with filled tubes into the instrument to lock it in place Check that everything is properly aligned and close the instrument door Select the SALMONELLA program sequence by scrolling with the arrow key and press the START button For the Shell Eggs method select the Salmonella eggs program from the BeadRetriever menu While the instrument is in operation the door must be kept closed Each processing step and the total time remaining can be followed on the LC display At the end of the program run remove the sample rack from the instrument For each sample process the bead bacteria complexes either using the RAPID or ENHANCED methods described below Remove the sample tip combs and discard into a biohazard waste container together with the tube strips ISOLATION PROCEDURE The ENHANCED Method This is the recommended method for all food and environmental samples Presumptive Salmonella positive results
56. r and the Instructions for Use accompanying the Dynabeads anti Salmonella product Details regarding buffers media other components as well as the protocols are provided with the Dynabeads anti Salmonella Instructions for Use SAMPLE PREPARATION Food feed and environmental samples 1 Weigh 25 g of sample material and place into a stomacher bag with filter and add 225 ml of pre enrichment broth Invitrogen recommends buffered peptone water as a pre enrichment broth 2 Mix well using the stomacher apparatus a stomacher bag with filter removes particulate matter as well as fatty components and allows easy pipetting of clear aliquot for analysis For certain foods for example bony meat pasta etc a blender is preferred prior to using a stomacher bag to avoid the risk of perforation However after blending the contents should be transferred into a stomacher bag with a filter For environmental samples using a swab place the swab into 10 50 ml of pre enrichment broth and incubate as described below Incubate the prepared sample in the stomacher bag for 18 24 hours at 37 C Mix the pre enriched sample thoroughly by homogenising once more Pipette 1 ml aliquot of the filtered suspension for the immunomagnetic separation procedure Change to a new pipette for each new sample o Neon Method for shell eggs Wash dirty eggs with a stiff brush under running water and dry with a paper towel Dip the eggs into 70 ethanol for
57. regulations for the disposal of infectious material The samples can be potentially infectious Dispose of all used tube strips tip combs disposable 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 It is strongly recommended that the complete decontamination procedure be performed before relocating the instrument from one laboratory to another Decontamination is not required for the proper functioning of the instrument Example of decontaminants e Ethanol 70 e Virkon solution 1 3 e Glutaraldehyde solution 4 Chloramine e Microcide SO 1 64 e Decon 90 min 4 Always use disposable gloves and protective clothing and operate in a well ventilated area 1 Prepare the decontaminating agent for example 200 4 glutaraldehyde solution or another agent recommended by your safety officer 2 the tube strip tray 3 Switch OFF the power and disconnect the mains supply cable Fig 5 3 4 Disinfect the outside of the instrument using cloth dampened with 70 ethanol 5 Place the 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 th
58. ries or distributors The guarantee is not transferable to a third party without prior written approval from Invitrogen 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 Invitrogen representative This card acts as a warranty certificate 10 2 Warranty limitations e Consumables are not included in the warranty warranty does not extend beyond the expiration date of the reagent kit BeadRetriever User Manual Rev 05 November 12 32 Chapter 11 Ordering Information 11 ORDERING INFORMATION Code Instrument System 159 50 BeadRetriever M 100 240 V 11 1 List of accessories Code Item Quantity 1508270 BeadRetriever User Manual 1 159 51 BeadRetriever Tubes and Tips 1 11 2 List of spare parts Code Item Quantity 159 52 Tube strip tray Tube Rack 1 11 3 List of rec
59. rporation Filta Max is a registered trademark of IDEXX Laboratories Inc Microside SQ is a registered trademark of Global Biotechnologies Inc Tween is a registered trademark of ICI Americas Inc USA Virkon is a registered trademark of Antec International Ltd All other trademarks and registered trademarks are the sole property of their respective holders Disclaimer Invitrogen 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 Invitrogen assumes no liability for any errors or omissions nor for any damages resulting from the application or use of this information No liability for consequential damages Invitrogen shall not be liable for any indirect or consequential damages whatsoever arising out of the use or inability to use this product BeadRetriever User Manual Rev 05 November 12 Chapter 1 Safety Symbols and Markings 1 SAFETY SYMBOLS AND MARKINGS These symbols are intended to draw your attention to particularly important information and alert you to the presence of hazards as indicated SAFETY SYMBOLS USED IN THE BEADRETRIEVER Power ON C Power OFF SN Serial number REF Catalog number Date of manufacture Consult Instructions for Use WEEE symbol This product is required to comply with the European Union
60. rror 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 Serial no already set Too high pos offset Invalid parameter Out of memory No acceleration tbl NV parameters lost Program full Time already expired BeadRetriever User Manual Rev 05 November 12 Chapter 9 Troubleshooting Guide 27 92 Troubleshooting guide Chapter 9 Troubleshooting Guide Error Cause Action e Cannot move track Head position error e Head timeout position error e Track timeout e Magnets position error e Magnets timeout parameters lost The track cannot be moved because the head is not up The calculated head position does not agree with the head home sensor signal The head has not arrived at the home sensor within the allowed time The calculated track position does not agree with the track home sensor signal The track has not arrived at the home sensor within the allowed time The calculated magnetic rod position does not agree with the rod home sensor signal The magnetic rods have not arrived at the home sensor within the allowed time The nonvolatile parameters of the instrument are lost Press STOP If the fault cannot be remedied then switch the power on and off Check that t
61. rt USB Performance specifications Tube strip tray autoclavable Special design 3 x 5 format Magnetic rods fixed 3 x 5 format Capacity Vessel volumes 15 samples run 3 x 5 50 pl 1 mi Vessel type disposable Special tube strip 1 x 5 tubes Tip comb disposable Magnetic bead size Special design 1 x 5 format ca gt 1 um Collecting efficiency 95 vessel washing buffer Incubating temperature Keyboard Display Ambient temperature no incubation START STOP two cursor keys LCD Software Space for ca 200 protocols loading through the external PC BeadRetriever User Manual Rev 05 November 12 25 Chapter 8 Technical Specifications 8 2 Safety specifications This section describes the safety specifications for the BeadRetriever instrument 8 3 conformity with the requirements BeadRetriever bears the following markings Type 701 100 240 V 50 60 Hz 40 VA CE mark BeadRetriever 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 2012 19 EC Waste of Electrical and Electronic Equipment 2011 651 EC RoHS Directive Restriction of the Use of Certain Hazardous Substances in Electrical and Electronic Equipment 2006 42 EC Machinery Directive The sa
62. rts are missing contact Invitrogen or your local Invitrogen representative 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 Invitrogen or your local Invitrogen representative BeadRetriever User Manual Rev 05 November 12 14 Chapter 5 Installation 5 1 4 Environmental requirements When you set up your BeadRetriever 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 Make sure that there is sufficiently room behind the instrument to enable disconnecting the device Make sure the ambient air is clean and free of corrosive vapors smoke and dust Make sure the ambient temperature range is between 10 C 50 F and 40 C 104 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
63. s This will ensure that the product is properly maintained and gives trouble free service Contact your local Invitrogen representative for details 7 6 Disposal of the instrument gt Decontaminate the instrument prior to disposal See Sections 7 3 9 4 1 and 9 4 1 1 on decontamination e 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 e Regarding the original packaging and packing materials use the recycling operators known to you e For more information contact your local Invitrogen office or local supplier BeadRetriever User Manual Rev 05 November 12 24 Chapter 8 Technical Specifications 8 TECHNICAL SPECIFICATIONS 8 1 General specifications Technical specifications Overall dimensions ca 290 W 290 mm 0 x 310 mm 11 4 in W x 11 4 in D x 12 2 in H Weight ca 10 kg 23 lbs Operating conditions Indoor use 10 C 40 C RH 90 max Transportation conditions 40 C 70 C packed in transport packaging Storage conditions 25 C 50 C packed in transport packaging Mains power supply 100 240 V a c 50 60 Hz nominal Automatic voltage detection Power consumption 40 VA max Heat dissipation 136 BTU max Computer interface Serial RS 232C po
64. sample concentrates spiked water samples environmental swab samples and dilute suspensions of target non target or reference organisms This protocol is also suitable for samples which have been acid or heat treated If the sample has been stored below room temperature leave sufficient time to allow the sample to equilibrate to room temperature Samples may be prepared by standard filtration and or centrifugation methods appropriate to meet local regulations This is recommended where target cell numbers are expected to be below 1000 colony forming units cfu per liter Environmental swab samples may be eluted into a suitable volume of Maximum Recovery Diluent prior to IMS Note Maximum Recovery Diluent is an isotonic diluent containing a low level of peptone used for maintaining the viability of organisms during dilution procedures Please use Oxoid Cat No CM733 Difco Cat No 218971 Dehydrated 500 g or equivalent BS ISO 11731 2 2004 BS 6068 4 18 2004 Water quality Detection and enumeration of Legionella Direct membrane filtration method for waters with low bacterial counts BS 6068 4 12 1998 ISO 11731 1998 Water quality Microbiological methods Detection and enumeration of Legionella Current Work In Hand http bsonline techindex co uk AIMS OPERATION PROCEDURE Place one disposable sample tube strip into a BeadRetriever rack for each sample to be processed and using aseptic technique dispense reagents into each tube T
65. sfer 50 ul of the resuspended bead bacteria complexes from the 5 tube onto two Listeria plating media one of which should be Palcam agar and plate by standard streaking with a loop or the swab streak technique Fig 15 1 All inoculated plating media must be incubated at 37 C The plates are read after 24 h and if necessary after 48 h for presumptive Listeria colonies Total analysis time from sample receipt to presumptive results is 48 h CONFIRMATION The presumptive Listeria colonies must be confirmed by standard biochemical and serological testing or by genetic fingerprinting to identify the species BeadRetriever User Manual Rev 05 November 12 49 Appendix A Application Protocols for BeadRetriever Using Dynabeads for Selective Concentration of Target Microorganisms Automated IMS Using Dynabeads anti Legionella Dynabeads anti Legionella has been designed for use both manual and automated immunomagnetic separation of Legionella species spp from water samples and water sample concentrates Before proceeding with automated immunomagnetic separation carefully read the operating instructions of the BeadRetriever and the Instructions for Use accompanying the Dynabeads anti Legionella product Details regarding buffers media other components as well as the protocols are provided with the Dynabeads anti Legionella Instructions for Use SAMPLE PREPARATION This test protocol has been designed for use with 1 ml of water
66. slides according to standard procedure 25 ul methanol Once the slides are fixed stain with FITC monoclonal antibody for Giardia and Cryptosporidium Follow the manufacturer s instructions for FITC staining After staining with FITC then stain the slides with 4 6 diamidino 2 phenylindole DAPI according to standard procedure 50 ul for 2 minutes Wash the slides and mount with cover slip according to standard procedure The slides are then ready for oocysts enumeration via Fluorescence assay FA microscopy and Normanski differential interference contrast DIC microscopy ADDITIONAL CONFIRMATION PROCEDURE After samples are enumerated on the slides you can further process the sample to get information on the species or genotype of the Cryptosporidium oocysts captured by AIMS Using our Cryptosporidium genotyping kit Cat No 740 05 Complete Kit Cat No 740 06 Starter Kit Cat No 740 07 Reagents the user can isolate DNA from the oocysts adhered to the slide This material is then amplified with unique PCR primers to amplify a Cryptosporidium specific target This target is then digested with three distinct restriction enzymes to produce bands of different sizes These bands are resolved by gel electrophoresis Based on the banding pattern the user can achieve a presumptive species or genotype ID for the organisms present on the slide Refer to the Cryptosporidium Genotyping kit manual for further information BeadRetriever User Man
67. the tube strip containing a reagent BeadRetriever User Manual Rev 05 November 12 12 Chapter 4 Functional Description Magnetic beads are released by moving the tip comb s up and down several times at considerably high speed until all the beads have been mixed with the substance in the next reaction 4 4 3 Washing magnetic beads Washing the magnetic beads is a frequent and an important processing phase Washing is a combination of the release and collection processes in a tube 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 bead suspension evenly mixed in long running reactions the tip comb s can be moved up and down from time to time 445 Changing the volume during the magnetic bead processing The volume of the first tube can be larger than the volume of the next tube and this is used for concentration purposes see Fig 4 7 below ransfer y y 4 s Y f y gl id y y y y y 5 4 gt f 1 11 1 7 4 4 y 2 y om 4 ve a lt y ym Collection of Transfer into R
68. ther process can selectively concentrate Listeria from enriched sample matrices Before proceeding with automated immunomagnetic separation carefully read the operating instructions of the BeadRetriever and the nstructions for Use accompanying the Dynabeads anti Listeria product Details regarding buffers media other components as well as the protocols are provided with the Dynabeads anti Listeria Instructions for Use SAMPLE PREPARATION Food samples Weigh 25 g of food sample and place into a filter homogenizer bag Add 225 ml of Half Fraser broth and homogenize 1 2 3 Incubate the prepared sample at 30 C for 24 h 4 _ Mix pre enriched samples thoroughly by homogenizing once more Environmental samples 1 Take a swab sample from any surface material or filter 10 liters of water through a membrane filter 2 Place the swab or filter into an appropriate container filled with 10 50 of Half Fraser broth Incubate for 24 h at 30 C 3 Mix by shaking vigorously AIMS OPERATION PROCEDURE Users must aseptically dispense all reagents and samples into the tubes sequentially after the tube strips are fitted into the rack Users must read the nstructions for Use for Dynabeads anti Listeria before use as follows 1 Aseptically add 10 ul of properly mixed Dynabeads anti Listeria into the two sample tubes 1 and Aseptically add 500 ul of washing buffer PBST to sample tubes 1 and 2 Aseptically add 1 ml of washing buffer t
69. to standard local procedures 2 Use a flat ended forceps to remove the filter and transfer directly into a wide mouthed bottle 3 Add 90 ml of BPW or TSB to the contents of the bottle and shake vigorously to dislodge bacteria from the surface of the membrane 4 Incubate for 6 24 h at 37 C or 41 5 C 5 _ The use of a filter aid is recommended for samples that are too turbid for membrane filtration AIMS OPERATION PROCEDURE Place one disposable sample tube strip into a BeadRetriever rack for each sample to be processed and using aseptic technigue dispense reagents into each tube The tab on the tube strip may be used for labeling samples Users must read the nstructions for Use for each individual Dynabeads coli O157 product before use as follows 1 Resuspend Dynabeads anti E coli O157 until the pellet in the bottom disappears by vortexing Aseptically add 10 ul of properly mixed Dynabeads anti E coli 0157 into the two sample tubes 1 and 2 Aseptically add 500 ul of wash buffer to sample tubes 1 and 2 Aseptically add 1 ml of wash buffer to tubes 3 and 4 within the strip Aseptically add 100 of wash buffer to the 5 tube Remove the desired tube from the sample rack and place in a second sample rack one meter away Add 500 ul of a test sample to tubes 1 and 2 and transfer the inoculated tube to the first sample rack Repeat for the remaining samples Aseptically insert the sterile protective tip combs into the instru
70. to the mains power socket Fig 5 3 at the bottom 3 Connect the instrument to a correctly installed line power outlet that has a protective conductor that is grounded Warning Never operate your instrument from a power outlet that has no ground connection 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 guickly 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 3 It is recommended that you carry out an empty run e g a demonstration to verify proper instrument operation BeadRetriever User Manual Rev 05 November 12 17 Chapter 6 Routine Operation 6 ROUTINE OPERATION 6 7 Switching on Before you switch on the BeadRetriever 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 Howto use the keypad and display The liquid crystal display LCD is a 20 character by 2 line LCD The lower line of the display can show the steps of the protocol while the processing is running The remaining time till the end of the process is shown in the top right corner Furthermore if errors occur they will be displayed on the lower line
71. ual Rev 05 November 12 53 Appendix Brief User s Guide 16 APPENDIX B BRIEF USER S GUIDE e Switch the BeadRetriever on p 17 e Pipette the samples reagents and washing buffers into the tube strips p 19 40 Load the tube strip tray s onto the instrument p 19 e Insert the tip comb s into the tip comb holder slot s p 19 e Select the desired protocol from the keypad and display p 18 e Press the START button p 19 e After the end of the run remove the tube strips and tip combs p 21 e Maintain your BeadRetriever instrument regularly p 22 Fig 16 1 BeadRetriever keypad BeadRetriever User Manual Rev 05 November 12 54 Appendix BeadRetriever Feedback Form 17 APPENDIX C BEADRETRIEVER 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 Mr Mrs Ms Job title Position Surname block capitals First name block capitals Internet e mail address Excellent Above As Below Comments expectations expected expectations Operational costs Customer support Reagent kit Instructions Instrument User manual Ll O Operational reliability Design Ll 0 Ease of use Ll Ll Ll Ll Ll Ll Ll Ll Further instrument system developments desired

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