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SOLiD™ EZ Bead™ Emulsifier Getting Started Guide (PN 4441486E)

1. 13 Materials and equipment required 13 Before you begin mit A de ee dale dulce ee a e ere aia t 13 Prepare the Oil Master MIX 14 Pre rinse the SOLID EZ Bead Amplifier Pouch ee 16 M CHAPTER 2 Prepare and Run the Emulsifier E10 17 Prepare and run the Emulsifier E10 18 Preparethe P1 BeadsE10 c erai eee ea tea t R mn Bale nea ae Dens 18 Installthe oiliphase E10 i o ceto ata a eot a REIR Cini Adele 18 Prepare the aqueous phase E10 21 Set up and run the Emulsifier E10 22 Collect the emulsion E10 2 23 M CHAPTER3 Prepare and Run the Emulsifier E20 25 Prepare and run the Emulsifier E20 26 Preparethe PT BeadsE20 15 ip REL ERE a geen elis 26 Install the oil phase E20 26 Prepare the aqueous phase E20 29 Set up and run the Emulsifier E20 30 Collect the emulsion E20 32 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 3
2. t Not programmed Table 13 Covalent Declump 3 3 cycles Treatment 1 followed by Treatment 2 Treatment 1 Treatment 2 Duty Cycle 2 5 Intensity 6 9 Cycles Burst 100 100 Time 5 sec 30 sec Target wattage power 4 15 performance estimate WIT t Not programmed Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 61 Appendix C The Covaris S2 System Covaris S2 programs 62 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Instrument Warranty Information This appendix covers B Computer configuration 64 m Limited product warranty 64 m Damages claims and returns 66 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 63 Appendix D Instrument Warranty Information Computer configuration Computer configuration Applied Biosystems supplies or recommends certain configurations of computer hardware software and peripherals for use with its instrumentation Applied Biosystems reserves the right to decline support for or impose extra charges for supporting nonstandard computer configurations or components that have not been supplied or recommended by Applied Biosystems Applied Biosystems also reserves the right to require that computer hardware and software be restored to the standard configuration pri
3. T The kit includes consumables for all three instruments of the SOLiD EZ Bead System the Enricher Amplifier and Emulsifier 50 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Required kits E80 Appendix A Ordering Information Required kits E80 Required Applied Biosystems kits E80 Item part number Components Applied Biosystems SOLiD EZ Bead Emulsifier E80 Reagent Kit 4452722 Box 1 e SOLID EZ Bead Emulsifier E80 Aqueous Master Mix e SOLID EZ Bead Emulsifier E80 P1 Beads Box 2 e SOLID EZ Bead Emulsifier E80 P1 Reagent e SOLID EZ Bead Emulsifier E80 P2 Reagent e SOLID EZ Bead Emulsifier E80 BB Solution Box 3 e SOLID EZ Bead Emulsifier Reagent 1X TEX Buffer e SOLID EZ Bead Emulsifier Reagent 1X Low TE Buffer SOLIDO EZ Bead Emulsifier Oil Pack Kit 4457185 SOLID EZ Bead Emulsifier Oil Kit e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 1 e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 2 e SOLID EZ Bead Emulsifier Oil Kit Oil SOLID EZ Bead Emulsifier Oil Accessory Kit s SOLID EZ Bead Emulsifier Oil Kit Oil Mix Bottle e SOLID EZ Bead Emulsifier Oil Kit Oil Funnel SOLIDO EZ Bead Emulsifier E80 Accessories Kit 4453070 SOLID EZ Bead Emulsifier E Large Jar Pack SOLID EZ Bead Emulsifier E Large AQ Tube Pack SOLID EZ Bead Emulsifier Tubing Pack SOLIDO EZ Bead E80 System Consumables 4 pack 4453095 SOLID EZ Bead
4. Turn the knob clockwise to unlock the compartment door then open the door Slide the coupler up and away from the impeller head Gently pull the impeller head from the coupler Swing out the arm at the base then remove the impeller and the jar from the compartment Detach the tubing from the port on the cap of the jar Remove the pump cassette tubing and AQ Tube from the instrument as one unit To release the tubing from the pump cassette turn the pinch wheel clockwise to the silver zone then remove the tubing Discard the used AQ Tube and tubing Store the pump cassette on top of the Emulsifier IMPORTANT The pump cassette is part of the instrument and is not disposable Close the door of the compartment then turn the knob counter clockwise to lock the door Immediately perform emulsion PCR ePCR on the emulsion using the SOLiD EZ Bead Amplifier refer to the Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 39 Z Chapter 4 Prepare and Run the Emulsifier E80 Prepare and run the Emulsifier E80 40 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Prepare and Run the Emulsifier E120 This chapter explains how to prepare and run the Emulsifier for the E120 scale Choose the scale of preparation of templated beads on page 13 l Prepare the Oil Master Mix on page 14
5. esse n 53 R quired quipMent us sis ah o e ERI SEP uer uber de e abe 54 Optional equipment ed wie ead de eee dd 56 APPENDIX B Troubleshooting 57 APPENDIX C The Covaris S2 System 59 Operation notes ole lus ire pog gw ae go ge ru IY KR a deseo a neue 60 Covaris S2 prograrms sd eee Shee dl Hd PI eM P i a a aa n 61 APPENDIX D Instrument Warranty Information 63 Computer configurations sooo Spr US aU RPM ll Sie bee ad 64 Limited product warranty od need es aia ee US SED nana LANES EM 64 Damages claims and returns se 66 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Contents APPENDIX E Safety sc e 67 Instrumentation safety 25 o t ae Re LAS o ie RM Hit er 68 Symbols on Instruments ence vete RE a EEE a aaa 68 Location of safety labels on instruments 2 69 General instrument safety sse 69 Physical hazard safety i issu be A RA LA RE EL dee Lxx 70 Electrical safey clc A 0 ea UN RR TU 70 Workstation Safety ial ati pelt als uie hah a te Raum 71 Safety and electromagnetic compatibility EMC standards 71 Chemical safety ss runs s a o uas ete e 73 General chemical safety 220 73 SDSS eset a ca e c bl eee Mah ae M Lb L Een PE 74 Chemical Waste safely a rere b ele ERR nee nt 74 Safely alerts
6. Pe ue dm Cap the AQ Tube and swirl to mix the solution Place the AQ Tube back on ice Power on the Emulsifier Ensure that the Oil Master Mix in the Emulsifier Jar is at room temperature Vortex then pulse spin the tube of prepared P1 beads Sonicate the prepared P1 Beads using the Covalent Declump 3 program on the Covaris S2 System for program conditions see Covalent Declump 3 on page 61 then pulse spin the tube IMPORTANT Perform steps 5 and 6 below as quickly as possible before the beads settle in the aqueous phase If the beads settle before step 6 swirl the aqueous phase again then perform step 6 Immediately add the beads to the aqueous phase then swirl the AQ Tube until the suspension is uniformly brown Immediately load the AQ Tube onto the Emulsifier then attach the tubing to the cap of the AQ Tube see Figure 15 on page 37 Ensure that the beads have not settled CAUTION PINCH HAZARD Moving Parts Moving parts can crush AS pinch and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Start the run a Press the scale selection button to select the E80 scale on the instrument key panel see Figure 16 on page 39 b Press the start button Q The motor starts and runs for 10 seconds before the pump starts The middle light on the panel is lit yellow IMPORTANT Do not open the door of the emulsifier while a run is in progress Opening the door auto
7. Complete IMPORTANT To stop the run press the stop button then press the stop button again to confirm that you want to stop the run The left light on the panel is lit red and the panel says Run aborted Stop the run only if the SOLiD EZ Bead Emulsifier malfunctions If you stop the run before the run is complete then discard the emulsion because it is not properly prepared Troubleshoot if necessary see Appendix B Troubleshooting on page 57 then prepare and install the reagents again for the appropriate scale of emulsification Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Collect the emulsion E120 Chapter 5 Prepare and Run the Emulsifier E120 5 Prepare and run the Emulsifier E120 Figure 21 How to use the Emulsifier Emulsification scales Select scale of run Stop run Select speed 9 Start run Light color Meaning Green Run is complete Yellow Instrument is running Red uec co US 9 10 11 Run aborted failed run or error Ensure that the run is finished When the run is finished the clock reads 00 00 Turn the knob clockwise to unlock the compartment door then open the door Slide the coupler up and away from the impeller head Gently pull the impeller head from the coupler Swing out the arm at the base then remove the impeller and the jar from the compartment Detach the tubing from the port on the cap of the
8. accurate chemistry kit use or safe use of a chemical CAUTION Indicates a potentially hazardous situation that if not avoided may result in minor or moderate injury It may also be used to alert against unsafe practices WARNING Indicates a potentially hazardous situation that if not avoided could result in death or serious injury DANGER Indicates an imminently hazardous situation that if not avoided will result in death or serious injury This signal word is to be limited to the most extreme situations Except for IMPORTANTS each safety alert word in an Life Technologies document appears with an open triangle figure that contains a hazard symbol These hazard symbols are identical to the hazard symbols that are affixed to Applied Biosystems instruments see Safety symbols on page 68 The Safety Data Sheets SDSs for any chemicals supplied by Applied Biosystems or Ambion are available to you free 24 hours a day For instructions on obtaining SDSs see SDSs on page 74 IMPORTANT For the SDSs of chemicals not distributed by Life Technologies or Ambion contact the chemical manufacturer The following CAUTION WARNING and DANGER statements may be displayed on Life Technologies instruments in combination with the safety symbols described in the preceding section Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide About This Guide Safety information Hazard symbol English Fran
9. s SOLID EZ Bead Emulsifier Oil Kit Oil Mix Bottle e SOLID EZ Bead Emulsifier Oil Kit Oil Funnel SOLIDG EZ Bead Emulsifier E120 Accessories Kit 4465569 SOLID EZ Bead Emulsifier E Large Jar Pack SOLID EZ Bead Emulsifier E Large AQ Tube Pack SOLID EZ Bead Emulsifier Tubing Pack SOLIDO EZ Bead E120 System Consumables 4 pack 4465571 e SOLID EZ Bead Emulsifier E120 Reagent Kit e SOLID EZ Bead Emulsifier E120 Accessories Kit e SOLID EZ Bead Amplifier E120 Accessories Kit e SOLID EZ Bead Enricher E120 Reagent Kit e SOLID EZ Bead Enricher Buffer Kit e SOLID EZ Bead Enricher Accessories Kit t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance T The kit includes consumables for all three instruments of the SOLID EZ Bead System the Emulsifier Amplifier and Enricher Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Required consumables Appendix A Ordering Information Required consumables Required consumables Item t Source Ethylene glycol American Bioanalytical AB00455 01000 1 5 mL DNA LoBind Tubes Eppendorf 022431021 Falcon high clarity polypropylene conical centrifuge tube 9400 RCF rating sterile 50 mL Becton Dickinson 352070 BD slip tip disposable tuberculin syringe 1mL Becton Dickinson 309602 BD syringe Luer Lok tip 3 mL Becton
10. y Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 Prepare the P1 Beads E120 on page 42 l Install the oil phase E120 on page 42 Prepare the aqueous phase E120 on page 45 Set up and run the Emulsifier E120 on page 46 Collect the emulsion E120 on page 47 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 41 5 Chapter 5 Prepare and Run the Emulsifier E120 Prepare and run the Emulsifier E120 Prepare and run the Emulsifier E120 Note You can thaw the Emulsifier reagents on ice as you prepare the P1 Beads and install the oil phase see Prepare the aqueous phase E120 on page 45 Prepare the P1 1 Beads E120 10 11 12 Install the oil phase 1 E120 5 42 If you are amplifying the library with the SOLiD EZ Bead Amplifier then we recommend pre rinsing the pouches used with the Amplifier before emulsifying the library see Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 Vortex and pulse spin one tube of Emulsifier E120 P1 Beads Ensure that the P1 Beads are completely resuspended then pipet 1430 uL of P1 Beads into each of TWO 2 0 mL DNA LoBind Tubes for a total of 2860 uL of P1 Beads Place both tubes of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend EACH tube of beads in 1430 uL Emulsifier E120 B
11. Contents M CHAPTER Prepare and Run the Emulsifier E80 33 Prepare and run the Emulsifier E80 34 Prepare the P1 Beads E80 34 Install the oll phase E80 2 sies e et is autre sante tes 34 Prepare the aqueous phase E80 37 Set up and run the Emulsifier E80 38 Collect the emulsion E80 39 CHAPTER 5 Prepare and Run the Emulsifier E120 41 Prepare and run the Emulsifier E120 42 Prepare the P1 Beads E120 42 Iastalliihe oil phase E120 i oxo viet ven bee a ee n ag ce td ee ee 42 Prepare the aqueous phase E120 45 Set up and run the Emulsifier E120 46 Collect the emulsion E120 47 APPENDIX A Ordering Information 49 Required kits E10 73 n actes d oec o ac ae Se acs tbt ets 49 Required kits E20 ree A eroe A sie 50 R g iredkits E80 2 sides ERR Den Rene I REM Gavel ghee dH M 51 Required kits E120 vaccine eM REA ae ae cR e Me ale od 52 Required consumables
12. E80 7 Prepare and run the Emulsifier E80 10 Close the door of the compartment then turn the knob counter clockwise to lock the door see Figure 15 Figure 15 Placement of Emulsifier E Large AQ Tube Tubing and E Large Jar Tubing tab SOLID EZ Bead Emtlsifier SOLID EZ Bead Emulsifier SOLIiD EZ Bead Emulsifier E Large AQ Tube Tubing in pump cassette E Large Jar Prepare the 1 Thaw on ice aqueous phase Emulsifier E80 P1 reagent then vortex the reagent for a few seconds and E80 pulse spin Emulsifier E80 P2 reagent then vortex the reagent for a few seconds and pulse spin Library template Place on ice Emulsifier E80 Aqueous Master mix Do not vortex 2 Place the Emulsifier E Large AQ Tube on ice for filling 3 Prepare a 1 10 dilution of P1 Reagent by adding 21 uL of P1 reagent to 189 uL of Low TE Buffer in a 1 5 mL DNA LoBind Tube 4 Add the appropriate volume to the AQ Tube in this order E80 Reagent Volume pL Aqueous Master Mix 48000 Xt 1 10 diluted P1 Reagent 200 Undiluted P2 Reagent 300 Library template X t X Volume of library template Use a Pipet Aid pipette fitted with a 50 mL serological pipette to dispense Aqueous Master Mix into the E Large AG Tube Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 37 4 Chapter 4 Prepare and Run the Emulsifier E80 Prepare and run the Emulsifier E80 Set up and run the Emulsifier E80 38
13. Emulsifier E80 Reagent Kit SOLID EZ Bead Emulsifier E80 Accessories Kit SOLID EZ Bead Amplifier E80 Accessories Kit SOLID EZ Bead Enricher E80 Reagent Kit SOLID EZ Bead Enricher Buffer Kit SOLID EZ Bead Enricher Accessories Kit t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance T The kit includes consumables for all three instruments of the SOLID EZ Bead System the Emulsifier Amplifier and Enricher Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 51 Appendix A Ordering Information Required kits E120 Required kits E120 Required Applied Biosystems kits E120 Item part number Components Applied Biosystems SOLIDE EZ Bead Emulsifier E120 Reagent Kit 4465555 Box 1 e SOLID EZ Bead Emulsifier E120 Aqueous Master Mix e SOLID EZ Bead Emulsifier E120 P1 Beads Box 2 e SOLID EZ Bead Emulsifier E120 P1 Reagent e SOLID EZ Bead Emulsifier E120 P2 Reagent e SOLID EZ Bead Emulsifier E120 BB Solution Box 3 e SOLID EZ Bead Emulsifier Reagent 1X TEX Buffer e SOLID EZ Bead Emulsifier Reagent 1X Low TE Buffer SOLIDO EZ Bead Emulsifier Oil Pack Kit 4457185 SOLID EZ Bead Emulsifier Oil Kit e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 1 e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 2 e SOLID EZ Bead Emulsifier Oil Kit Oil SOLID EZ Bead Emulsifier Oil Accessory Kit
14. discretion defective accessories during this warranty period After this warranty period Applied Biosystems will pass on to the buyer to the extent that it is permitted to do so the warranty of the original manufacturer for such accessories With the exception of consumable and maintenance items replaceable products or components used on or in the instrument are themselves warranted to be free of defects in materials and workmanship for a period of ninety 90 days Applied Biosystems warrants that chemicals and other consumable products will be free of defects in materials and workmanship when received by the buyer but not thereafter unless otherwise specified in documentation accompanying the product Applied Biosystems warrants that for a period of ninety 90 days from the date the warranty period begins the tapes diskettes or other media bearing the operating software of the product if any will be free of defects in materials and workmanship under normal use If there is a defect in the media covered by the above warranty and the media is returned to Applied Biosystems within the ninety 90 day warranty period Applied Biosystems will replace the defective media Applied Biosystems does not warrant that the operation of the instrument or its operating software will be uninterrupted or error free Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Appendix D Instrument Warranty Information Limited product wa
15. does not leak Place the Amplifier Pouch on the bench top Massage the pouch for 2 to 3 minutes to ensure that the Oil Master Mix coats the inner surface of the pouch Over a waste container unscrew the cap s then drain the Oil Master Mix into the container Gently squeeze the Amplifier Pouch to expel any remaining Oil Master Mix from the pouch Drain the Amplifier Pouch upside down for at least 20 minutes on a layer of clean lint free wipes Kimwipes Cleaning Wipes After draining the pouch the pouch is ready for use IMPORTANT Do not re use Oil Master Mix from pre rinsing Screw the cap s tightly on the nozzle s until the Amplifier Pouches are ready for use Use the pre rinsed Amplifier Pouches the same day Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Prepare and Run the Emulsifier E10 This chapter explains how to prepare and run the Emulsifier for the E10 scale Choose the scale of preparation of templated beads on page 13 l Prepare the Oil Master Mix on page 14 l Pre rinse the SOLID EZ Bead Amplifier Pouch on page 16 l Prepare the P1 Beads E10 on page 18 Install the oil phase E10 on page 18 Prepare the aqueous phase E10 on page 21 Set up and run the Emulsifier E10 on page 22 l Collect the emulsion E10 on page 23 Applied Biosystems SOLID EZ Bead Emulsifier Getting S
16. jar Remove the pump cassette tubing and AQ Tube from the instrument as one unit To release the tubing from the pump cassette turn the pinch wheel clockwise to the silver zone then remove the tubing Discard the used AQ Tube and tubing Store the pump cassette on top of the Emulsifier IMPORTANT The pump cassette is part of the instrument and is not disposable Close the door of the compartment then turn the knob counter clockwise to lock the door Immediately perform emulsion PCR ePCR on the emulsion using the SOLiD EZ Bead Amplifier refer to the Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 47 5 Chapter 5 Prepare and Run the Emulsifier E120 Prepare and run the Emulsifier E120 48 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Ordering Information Order one or more of the kits according to the scale of emulsion preparation Required kits E10 Required Applied Biosystems kits E10 Item part number Components Applied Biosystems SOLiD9 EZ Bead Emulsifier E10 Reagent Kit 4452720 Box 1 e SOLID EZ Bead Emulsifier E10 Aqueous Master Mix e SOLID EZ Bead Emulsifier E10 P1 Beads Box 2 e SOLID EZ Bead Emulsifier E10 P1 Reagent e SOLID EZ Bead Emulsifier E10 P2 Reagent e SOLID EZ Bead Emulsifier E10 BB Solution Box 3 e SOLID EZ Bead Emulsi
17. of the chemicals in the waste container before you store handle or dispose of chemical waste Provide primary and secondary waste containers A primary waste container holds the immediate waste A secondary container contains spills or leaks from the primary container Both containers must be compatible with the waste material and meet federal state and local requirements for container storage Minimize contact with chemicals Wear appropriate personal protective equipment when handling chemicals for example safety glasses gloves or protective clothing For additional safety guidelines consult the SDS Minimize the inhalation of chemicals Do not leave chemical containers open Use only with adequate ventilation for example fume hood For additional safety guidelines consult the SDS Handle chemical wastes in a fume hood After emptying a waste container seal it with the cap provided Dispose of the contents of the waste tray and waste bottle in accordance with good laboratory practices and local state provincial or national environmental and health regulations Waste disposal If potentially hazardous waste is generated when you operate the instrument you must Characterize by analysis if necessary the waste generated by the particular applications reagents and substrates used in your laboratory Ensure the health and safety of all personnel in your laboratory Ensure that the instrument waste is stored tran
18. on how to choose the scale of preparation of templated beads and describes how to prepare Oil Master Mix and how to pre rinse the SOLiD EZ Bead Amplifier Pouch Review the general workflow to prepare and sequence a library on page 12 Choose the scale of preparation of templated beads on page 13 Before you begin on page 13 Prepare the Oil Master Mix on page 14 y Pre rinse the SOLIDO EZ Bead Amplifier Pouch on page 16 y Chapter 2 Prepare and Run the Emulsifier E10 on page 17 Chapter 3 Prepare and Run the Emulsifier E20 on page 25 Chapter 4 Prepare and Run the Emulsifier E80 on page 33 Chapter 5 Prepare and Run the Emulsifier E120 on page 41 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 11 Chapter 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch Review the general workflow to prepare and sequence a library Steps Quantitate Titrate the library l Emulsify the beads l Amplify the beads L Enrich the templated beads Modify the 3 ends of the DNA Deposit beads perform bead sequencing References 5500 Series SOLiD Systems Fragment Library Preparation User Guide Part no 4460960 5500 Series SOLIDO Systems Mate Paired Library Preparation User Guide Part no 4460958 Prepare and Run the Emulsifier E10 on page 17 Prep
19. port of the cap 8 Insert the 3 stop tubing with one stop in the pump cassette see Table 3 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 19 Chapter 2 Prepare and Run the Emulsifier E10 Prepare and run the Emulsifier E10 Table 3 How to insert the tubing into the pump cassette 1 Using your right hand hold the pump cassette with your thumb and forefinger 2 Using your left hand grasp the tubing with your thumb and forefinger above the black stop 3 Pull the tubing slightly to slide the tubing into the pump cassette 4 Slide the tubing until the stop is aligned with the stop fixture on the outside edge of the pump cassette 5 Pull the tubing slightly towards the inside of the pump cassette to fit the stop snugly into the fixture a Repeat step 8 to insert the tubing with the second stop at the other end of the pump cassette Ensure that the tubing is in the inner groove opposite the second stop see Table 3 and that the tubing is in the correct orientation see Figure 3 Figure 3 Correct orientation of the Emulsifier Tubing b Install the pump cassette into the roller head of the pump see Figure 5 on page 21 c Turn the pinch wheel in the pump cassette counter clockwise so that the mark is in the blue zone see Figure 4 on page 20 This adjustment maintains correct tubing tension for normal pump operation Figure 4 Turn the pinch wheel in the pump
20. seconds and pulse spin Library template Place on ice Emulsifier E20 Aqueous Master mix Do not vortex 2 Place the Emulsifier E Small AQ Tube on ice for filling 3 Prepare a 1 10 dilution of P1 Reagent by adding 5 uL of P1 reagent to 45 uL of Low TE Buffer in a 1 5 mL DNA LoBind Tube 4 Add the appropriate volume to the AQ Tube in this order E20 Reagent Volume pL Aqueous Master Mix 11060 Xt 1 10 diluted P1 Reagent 46 Undiluted P2 Reagent 69 Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 29 Set up and run the Emulsifier E20 30 Chapter 3 Prepare and Run the Emulsifier E20 Prepare and run the Emulsifier E20 E20 Reagent Volume pL Library template X t X Volume of library template Use a Pipet Aid pipette fitted with a 25 mL serological pipette to dispense Aqueous Master Mix into the E Small AQ Tube SA Ev p D Cap the AQ Tube and swirl to mix the solution Place the AQ Tube back on ice Power on the Emulsifier Ensure that the Oil Master Mix in the Emulsifier Jar is at room temperature Vortex then pulse spin the tube of prepared P1 beads Sonicate the prepared P1 Beads using the Covalent Declump 1 program on the Covaris S2 System for program conditions see Covalent Declump 1 on page 61 then pulse spin the tube IMPORTANT Perform steps 5 and 6 below as quickly as possible before the beads settle in the aqueous phase If the b
21. 8 Symbols on instruments s RR sk ea aa eek Uae Se dense E E 68 Location of safety labels on instruments 69 General instrument safety xe sek eet e ei E RE aaa Y LE det 69 Physical hazard safety 70 Electrical safety 2 c sauts as dunk RR MES Tes Aa De eae data 70 Workstation safety zie cha nn Anar Ged nails eon Ce EA 71 Safety and electromagnetic compatibility EMC standards 71 B Chemical safety ests nt mima ae ai ed ane Eee EH at PR ee da 73 General chemical safety 73 A A Mah et edet 74 Chemical waste safety 74 B Safety alerts hr Lee Moiese an aaa acts deed Mann Ea aa e 76 Specific alerts for instrumentation 76 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 67 E Appendix E Safety Instrumentation safety Instrumentation safety Symbols on instruments 68 Note The products accompanying this manual are for demonstration purposes only and are not for sale Electrical symbols on instruments The following table describes the electrical symbols that may be displayed on Life Technologies instruments Symbol Description Indicates the On position of the main power switch Indicates the Off position of the main power switch Indicates a standby switch b
22. Applied Biosystems SOLiD EZ Bead Emulsifier E20 Reagent Kit 4452721 Box 1 e SOLID EZ Bead Emulsifier E20 Aqueous Master Mix e SOLID EZ Bead Emulsifier E20 P1 Beads Box 2 e SOLID EZ Bead Emulsifier E20 P1 Reagent e SOLID EZ Bead Emulsifier E20 P2 Reagent e SOLID EZ Bead Emulsifier E20 BB Solution Box 3 e SOLID EZ Bead Emulsifier Reagent 1X TEX Buffer e SOLID EZ Bead Emulsifier Reagent 1X Low TE Buffer SOLIDO EZ Bead Emulsifier Oil Pack Kit 4457185 SOLID EZ Bead Emulsifier Oil Kit e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 1 e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 2 e SOLID EZ Bead Emulsifier Oil Kit Oil SOLID EZ Bead Emulsifier Oil Accessory Kit e SOLID EZ Bead Emulsifier Oil Kit Oil Mix Bottle e SOLID EZ Bead Emulsifier Oil Kit Oil Funnel SOLIDO EZ Bead Emulsifier E20 Accessories Kit 4453076 SOLID EZ Bead Emulsifier E Small Jar Pack SOLID EZ Bead Emulsifier E Small AQ Tube Pack SOLID EZ Bead Emulsifier Tubing Pack SOLIDO EZ Bead E20 System Consumables 4 pack 4453094 SOLID EZ Bead Emulsifier E20 Reagent Kit SOLID EZ Bead Emulsifier E20 Accessories Kit SOLID EZ Bead Amplifier E20 Accessories Kit SOLID EZ Bead Enricher E20 Reagent Kit SOLID EZ Bead Enricher Buffer Kit SOLID EZ Bead Enricher Accessories Kit t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance
23. B Solution Sonicate EACH tube of beads using the Bead Block Declump program on the Covaris S2 System for program conditions see Bead Block Declump on page 61 then pulse spin the tubes Place BOTH tubes of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in EACH tube in 715 uL of 1X TEX Buffer Combine the two tubes of beads into one tube Use 300 uL of 1X TEX Buffer to rinse out any remaining beads in the empty tube then add them into the first tube Discard the empty tube Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in 1430 uL of 1X TEX Buffer Store the beads at 4 C until they are ready for use Unscrew and retain the cap with impeller of the Emulsifier E Large Jar Tare the E Large Jar on an analytical balance Pour 67 9 g of Oil Master Mix into the E Large Jar see Prepare the Oil Master Mix on page 14 Screw the cap with the impeller on the jar Install the jar on the Emulsifier a Turn the knob clockwise to open the compartment door b Pull the arm back at the base towards the opening of the compartment c Slide the jar until it stops against the two posts at the rear of the compartment d Allow the arm to rest against the jar see Figure 17 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Gu
24. Description Indicates a pinch hazard and to proceed with appropriate caution Indicates the presence of a hot surface or other high temperature hazard and to proceed with appropriate caution pr Environmental symbols on instruments The following symbol applies to all Applied Biosystems electrical and electronic products placed on the European market after August 13 2005 Symbol Description Do not dispose of this product as unsorted municipal waste Follow local municipal waste ordinances for proper disposal provisions to reduce Y the environmental impact of waste electrical and electronic equipment WEEE 3 European Union customers Call your local Applied Biosystems Customer Service office for equipment pick up and recycling See www appliedbiosystems com for a list of customer service offices in the European Union Location of safety The SOLiD EZ Bead Emulsifier contains warnings at the locations shown below labels on The pinch hazard label is to the right of the pump head instruments The remaining labels are near the power input General T WARNING PHYSICAL INJURY HAZARD Use this product only as specified instrument safety in this document Using this instrument in a manner not specified by Life Technologies may result in personal injury or damage to the instrument T WARNING PHYSICAL INJURY HAZARD Using the instrument in a manner not specified by Applied Biosystems may resul
25. Dickinson 309585 Portable Pipet Aid with 110V recharger Portable Pipet Aid with 220V recharger Drummond 4 000 100 Drummond 4 000 200 Corning Stripette serological pipette Corning H 4488 Corning Stripette serological pipette Corning AME 4489 Corning Stripette serological pipette Corning emt 4490 Filtered pipettor tips MLS t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance T For the SDS of any chemical not distributed by Applied Biosystems contact the chemical manufacturer Before handling any chemicals refer to the SDS provided by the manufacturer and observe all relevant precautions Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 53 A Appendix A Ordering Information E Required equipment Required equipment Required equipment Item Source SOLID EZ Bead Emulsifier Applied Biosystems The system includes 4448419 e SOLID EZ Bead Emulsifier e Applied Biosystems SOLID EZ Bead Emulsifier Getting Started Guide and Applied Biosystems SOLIDO EZ Bead Emulsifier Quick Reference Card Covaris S2 System Applied Biosystems 4387833 110 V 110 V for U S customers Applied Biosystems 220 V for international customers 4392718 220 V or The system includes Covaris e Covaris S2 sonicator s Latitude laptop from Dell Inc s MultiTemp lll Thermostatic Circul
26. E80 34 Note You can thaw the Emulsifier reagents on ice as you prepare the P1 Beads and install the oil phase see Prepare the aqueous phase E80 on page 37 If you are amplifying the library with the SOLiD EZ Bead Amplifier then it is recommended to pre rinse the pouches used with the Amplifier before emulsifying the library see Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 Vortex and pulse spin one tube of Emulsifier E80 P1 Beads Ensure that the P1 Beads are completely resuspended then pipet 1430 uL of P1 Beads into a 2 0 mL DNA LoBind Tube Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in 1430 uL Emulsifier E80 BB Solution Sonicate the beads using the Bead Block Declump program on the Covaris 52 System for program conditions see Bead Block Declump on page 61 then pulse spin the tube Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in 1430 uL of 1X TEX Buffer Store the beads at 4 C until ready for use Unscrew and retain the cap with impeller of the Emulsifier E Large Jar Tare the E Large Jar on an analytical balance Pour 67 9 g of Oil Master Mix into the E Large Jar see Prepare the Oil Master Mix on page 14 Screw the ca
27. Emulsifier E80 Table 7 How to insert the tubing into the pump cassette 1 Using your right hand hold the pump cassette with your thumb and forefinger 2 Using your left hand grasp the tubing with your thumb and forefinger above the black stop 3 Pull the tubing slightly to slide the tubing into the pump cassette 4 Slide the tubing until the stop is aligned with the stop fixture on the outside edge of the pump cassette 5 Pull the tubing slightly towards the inside of the pump cassette to fit the stop snugly into the fixture a Repeat step 8 to insert the tubing with the second stop at the other end of the pump cassette Ensure that the tubing is in the inner groove opposite the second stop see Table 7 and that the tubing is in the correct orientation see Figure 13 Figure 13 Correct orientation of the Emulsifier Tubing b Install the pump cassette into the roller head of the pump see Figure 15 on page 37 c Turn the pinch wheel in the pump cassette counter clockwise so that the mark is in the blue zone see Figure 14 This adjustment maintains correct tubing tension for normal pump operation Figure 14 Turn the pinch wheel in the pump cassette so that the mark is in the blue zone 9 Position the tubing so that it threads through the tubing tab see Figure 15 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 4 Prepare and Run the Emulsifier
28. HAZARD All chemicals in the instrument including liquid in the lines are potentially hazardous Always determine what chemicals have been used in the instrument before changing reagents or instrument components Wear appropriate eyewear protective clothing and gloves when working on the instrument A WARNING CHEMICAL HAZARD Four liter reagent and waste bottles can crack and leak Each 4 liter bottle should be secured in a low density polyethylene safety container with the cover fastened and the handles locked in the upright position Wear appropriate eyewear clothing and gloves when handling reagent and waste bottles A WARNING CHEMICAL STORAGE HAZARD Never collect or store waste in a glass container because of the risk of breaking or shattering Reagent and waste bottles can crack and leak Each waste bottle should be secured in a low density polyethylene safety container with the cover fastened and the handles locked in the upright position Wear appropriate eyewear clothing and gloves when handling reagent and waste bottles Chemical safety guidelines To minimize the hazards of chemicals Read and understand the SDSs provided by the chemical manufacturer before you store handle or work with any chemicals or hazardous materials See About SDSs on page 74 Minimize contact with chemicals Wear appropriate personal protective equipment when handling chemicals for example safety glasses gloves or protec
29. Lies Lese LEA E REEMe Satine ag ohne ed ee awn b M dtd 76 Specific alerts for instrumentation 76 Documentation and Support 77 Related documentati nz finesse di Pad a ted aed ERA ata have 77 Obtaining support td ere ieee a eee beer e dite esed 77 nde woh nest aa cubs educa ants di ound Ce bane cng hax eL T 79 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 5 Contents 6 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide About This Guide Purpose This getting started guide provides the information that you need to prepare emulsions with the SOLIDE EZ Bead Emulsifier To use the SOLiD System DNA libraries to be sequenced require amplification on P1 Beads during emulsion PCR ePCR followed by enrichment of the templated beads Templated bead preparation can be performed using the SOLiD EZ Bead System s SOLiD EZ Bead Emulsifier To prepare emulsions containing library template and P1 Beads see this guide SOLiD EZ Bead Amplifier To perform emulsion PCR ePCR refer to the Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 s SOLiD EZ Bead Enricher To wash and enrich the templated beads After enrichment perform a Terminal Transferase reaction to modify the 3 ends of the library refer to the Applied Biosystems SOLiD EZ Bead Enrich
30. PPLIED BIOSYSTEMS IS ADVISED OF THE POSSIBILITY OF SUCH DAMAGES INCLUDING WITHOUT LIMITATION DAMAGES ARISING FROM OR RELATED TO LOSS OF USE LOSS OF DATA FAILURE OR INTERRUPTION IN THE OPERATION OF ANY EQUIPMENT OR SOFTWARE DELAY IN REPAIR OR REPLACEMENT OR FOR LOSS OF REVENUE OR PROFITS LOSS OF GOOD WILL LOSS OF BUSINESS OR OTHER FINANCIAL LOSS OR PERSONAL INJURY OR PROPERTY DAMAGE Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 65 Appendix D Instrument Warranty Information Damages claims and returns NO AGENT EMPLOYEE OR REPRESENTATIVE OF APPLIED BIOSYSTEMS HAS ANY AUTHORITY TO MODIFY THE TERMS OF THIS LIMITED WARRANTY STATEMENT OR TO BIND APPLIED BIOSYSTEMS TO ANY AFFIRMATION REPRESENTATION OR WARRANTY CONCERNING THE PRODUCT THAT IS NOT CONTAINED IN THIS LIMITED WARRANTY STATEMENT AND ANY SUCH MODIFICATION AFFIRMATION REPRESENTATION OR WARRANTY MADE BY ANY AGENT EMPLOYEE OR REPRESENTATIVE OF APPLIED BIOSYSTEMS WILL NOT BE BINDING ON APPLIED BIOSYSTEMS UNLESS IN A WRITING SIGNED BY AN EXECUTIVE OFFICER OF APPLIED BIOSYSTEMS THIS WARRANTY IS LIMITED TO THE BUYER OF THE PRODUCT FROM APPLIED BIOSYSTEMS AND IS NOT TRANSFERABLE Some countries or jurisdictions limit the scope of or preclude limitations or exclusion of warranties of liability such as liability for gross negligence or willful misconduct or of remedies or damages as or to the extent set forth above In such countries and jurisdiction
31. Part 1 General Requirements Canadian EMC standard This instrument has been tested to and complies with ICES 001 Issue 3 Industrial Scientific and Medical Radio Frequency Generators European safety and EMC standards CE Safety This instrument meets European requirements for safety Low Voltage Directive 2006 95 EC This instrument has been tested to and complies with standards EN 61010 1 2001 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use Part 1 General Requirements EMC This instrument meets European requirements for emission and immunity EMC Directive 2004 108 EC This instrument has been tested to and complies with standard EN 61326 Group 1 Class B Electrical Equipment for Measurement Control and Laboratory Use EMC Requirements Australian EMC Standards C This instrument has been tested to and complies with standard AS NZS 2064 Limits and Methods Measurement of Electromagnetic Disturbance Characteristics of Industrial Scientific and Medical ISM Radio frequency Equipment Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Appendix E Safety E Chemical safety Chemical safety General chemical Chemical hazard warning safety T WARNING CHEMICAL HAZARD Before handling any chemicals refer to the Safety Data Sheet SDS provided by the manufacturer and observe all relevant precautions WARNING CHEMICAL
32. Select scale of run Stop run Start run Light color Meaning Green Run is complete Yellow Instrument is running Red Run aborted failed run or error Ensure that the run is finished When the run is finished the clock says 00 00 Turn the knob clockwise to unlock the compartment door then open the door Slide the coupler up and away from the impeller head Gently pull the impeller head from the coupler Swing out the arm at the base then remove the impeller and the jar from the compartment Detach the tubing from the port on the cap of the jar Remove the pump cassette tubing and AQ Tube from the instrument as one unit To release the tubing from the pump cassette turn the pinch wheel clockwise to the silver zone then remove the tubing Discard the used AQ Tube and tubing Store the pump cassette on top of the Emulsifier IMPORTANT The pump cassette is part of the instrument and is not disposable Close the door of the compartment then turn the knob counter clockwise to lock the door Immediately perform emulsion PCR ePCR on the emulsion using the SOLiD EZ Bead Amplifier refer to the Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 23 Chapter 2 Prepare and Run the Emulsifier E10 Prepare and run the Emulsifier E10 24 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Gu
33. Small Jar E10 2 Tare the E Small Jar on an analytical balance 3 Pour 10 4 g of Oil Master Mix into the E Small Jar see Prepare the Oil Master Mix on page 14 4 Screw the cap with the impeller on the E Small Jar 5 Install the jar on the Emulsifier a Turn the knob clockwise to open the compartment door b Pull the arm back at the base towards the opening of the compartment c Slide the jar into the groove of the base of the compartment until the jar stops against the back of the groove d Allow the arm to rest against the jar see Figure 2 e Rotate the jar so that the port on the cap is to the left of the impeller shaft 18 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Chapter 2 Prepare and Run the Emulsifier E10 2 Prepare and run the Emulsifier E10 Figure 2 The jar rests on the back of the groove and the arm is against the jar CH E 6 Fit the impeller head into the locking slot Slide the coupler down until the coupler clicks to a stop to ensure that the impeller head is firmly in place in the coupler see Table 2 Table 2 How to fit the impeller head into the locking slot Locking slot Coupler Impeller head Coupler Impeller 1 Align the locking slot with the impeller head 2 Fit the impeller head into the groove below the coupler 3 Slide the coupler over the impeller head 7 Attach the tubing with the Luer Lok to the
34. USER GUIDE applied biosystems oy Life technologies Applied Biosystems SOLiD EZ Bead Emulsifier Catalog Number 4448419 Publication Part Number 4441486 Rev E Revision Date October 2011 Emulsion Emulsion Cleanup and PCR Bead Enrichment o technologies For Research Use Only Not intended for any animal or human therapeutic or diagnostic use Information in this document is subject to change without notice LIFE TECHNOLOGIES AND OR ITS AFFILIATE S DISCLAIM ALL WARRANTIES WITH RESPECT TO THIS DOCUMENT EXPRESSED OR IMPLIED INCLUDING BUT NOT LIMITED TO THOSE OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE IN NO EVENT SHALL LIFE TECHNOLOGIES AND OR ITS AFFILIATE S BE LIABLE WHETHER IN CONTRACT TORT WARRANTY OR UNDER ANY STATUTE OR ON ANY OTHER BASIS FOR SPECIAL INCIDENTAL INDIRECT PUNITIVE MULTIPLE OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR ARISING FROM THIS DOCUMENT INCLUDING BUT NOT LIMITED TO THE USE THEREOF NOTICE TO PURCHASER LIMITED USE LABEL LICENSE Research Use Only The purchase of this product conveys to the purchaser the limited non transferable right to use the purchased amount of the product only to perform internal research for the sole benefit of the purchaser No right to resell this product or any of its components is conveyed expressly by implication or by estoppel This product is for internal research purposes only and is not for use in commercial applications of any kind includin
35. are and Run the Emulsifier E20 on page 25 Prepare and Run the Emulsifier E80 on page 33 Prepare and run the Emulsifier E120 on page 42 Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 Applied Biosystems SOLiD EZ Bead Enricher Getting Started Guide Part no 4443494 Applied Biosystems SOLiD EZ Bead Enricher Getting Started Guide Part no 4443494 5500 Series SOLiD Sequencers Operation Guide Part no 4456991 12 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch Choose the scale of preparation of templated beads You can prepare templated beads according to the amount of library that you want to amplify Scale of preparation Average yield of templated beads E10 125 million E20 250 million E80 1 billion E120 2 2 billion Materials and equipment required See Ordering Information on page 49 for a list of equipment kits and consumables necessary for this procedure Before you begin Construct the DNA library of interest refer to 5500 Series SOLiD Systems Fragment Library Preparation User Guide Part no 4460960 and 5500 Series SOLiD Systems Mate Paired Library Preparation User Guide Part no 4460958 Prepare and quantitate the DNA library before proceeding with emulsion preparation ref
36. aster Mix into the E Small AQ Tube Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 21 Chapter 2 Prepare and Run the Emulsifier E10 Prepare and run the Emulsifier E10 Set up and run the Emulsifier E10 22 AeA Q N Screw on the cap to the AQ Tube then swirl to mix the solution Place the AQ Tube back on ice Power on the Emulsifier Ensure that the Oil Master Mix in the Emulsifier Jar is at room temperature Vortex then pulse spin the tube of prepared P1 beads Sonicate the prepared P1 Beads using the Covalent Declump 1 program on the Covaris S2 System for program conditions see Covalent Declump 1 on page 61 then pulse spin the tube IMPORTANT Perform steps 5 and 6 below as quickly as possible before the beads settle in the aqueous phase If the beads settle before step 6 swirl the aqueous phase again then perform step 6 Immediately add the beads to the aqueous phase then swirl the AQ Tube until the suspension is uniformly brown Immediately load the AQ Tube onto the Emulsifier then attach the tubing to the cap of the AQ Tube see Figure 5 on page 21 Ensure that the beads have not settled CAUTION PINCH HAZARD Moving Parts Moving parts can crush A pinch and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Start the run a Press the scale selection button to select the E10 scale on the instrument k
37. ator e Covaris 2 series Machine Holder for one 1 5 mL microcentrifuge tube e Covaris 2 series Machine Holder for one 0 65 mL microcentrifuge tube e Covaris 2 series Machine Holder for one 13 mm x 65 mm tube e Covaris 2 Series Machine Holder for one microTUBE e Covaris microTUBE Prep Station e Covaris Water Tank Label Kit s Covaris microTUBEs 1 pack of 25 For system materials summary refer to Covaris S2 System Materials Summary in the SOLIDO 4 System Site Preparation Guide Part no 4448639 6 Tube Magnetic Stand Applied Biosystems AM10055 Analytical balance 250 g capacity Major Laboratory Supplier MLS Vortexer MLS Pipettors 20 uL MLS 54 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Appendix A Ordering Information A Required equipment Required equipment Item Source Pipettors 200 uL MLS Pipettors 1000 uL MLS Ice bucket MLS Picofuge MLS t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance t For the SDS of any chemical not distributed by Applied Biosystems contact the chemical manufacturer Before handling any chemicals refer to the SDS provided by the manufacturer and observe all relevant precautions Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 55 Appendix A Ordering Information Optional equipme
38. cais CAUTION Hazardous chemicals Read the Safety Data Sheets SDSs before handling ATTENTION Produits chimiques dangereux Lire fiche technique associ e au produit avant toute manipulation CAUTION Hazardous waste Refer to SDS s and local regulations for handling and disposal ATTENTION D chets dangereux Lire fiche technique associ e et prendre connaissance de la r gulation locale associ es la manipulation et l limination des d chets DANGER High voltage DANGER Haute tension WARNING To reduce the chance of electrical shock do not remove covers that require tool access No user serviceable parts are inside Refer servicing to Life Technologies qualified service personnel AVERTISSEMENT Pour viter les risques d lectrocution ne pas retirer les capots dont l ouverture n cessite l utilisation d outils L instrument ne contient aucune pi ce r parable par Uutilisateur Toute intervention doit tre effectu e par le personnel de service qualifi venant de chez Life Technologies CAUTION Pinch hazard ATTENTION Risque de pincement Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide About This Guide Safety information 10 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch This chapter provides information
39. cassette so that the mark is in the blue zone 9 Position the tubing so that it threads through the tubing tab see Figure 5 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Chapter 2 Prepare and Run the Emulsifier E10 2 Prepare and run the Emulsifier E10 10 Close the door of the compartment then turn the knob counter clockwise to lock the door see Figure 5 Figure 5 Placement of Emulsifier E Small AQ Tube Tubing and E Small Jar Tubing tab SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier E Small AQ Tube Tubing in pump cassette E Small Jar Prepare the 1 Thaw on ice aqueous phase e Emulsifier E10 P1 reagent then vortex the reagent for a few seconds and E10 pulse spin Emulsifier E10 P2 reagent then vortex the reagent for a few seconds and pulse spin Library template Place on ice Emulsifier E10 Aqueous Master mix Do not vortex 2 Place the Emulsifier E Small AQ Tube on ice for filling 3 Prepare a 1 10 dilution of P1 Reagent by adding 4 uL of P1 reagent to 36 uL of Low TE Buffer in a 1 5 mL DNA LoBind Tube 4 Add the appropriate volume to the AQ Tube in this order E10 Reagent Volume pL Aqueous Master Mix 7400 xt 1 10 diluted P1 Reagent 30 8 Undiluted P2 Reagent 46 2 Library template X t X Volume of library template Use a Pipet Aid pipette fitted with a 10 mL serological pipette to dispense Aqueous M
40. documents Obtain information about customer training Download software updates and patches Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 77 Documentation and Support Obtaining support 78 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide C CAUTION description 8 chemical safety 73 chemical waste safety 74 75 D DANGER description 8 documentation related 77 E electrical safety 70 electromagnetic compatibility standards See EMC standards EMC standards 71 ergonomics safety 71 G guidelines chemical safety 73 chemical waste disposal 74 chemical waste safety 75 H hazard icons See safety symbols on instruments 68 hazard symbols See safety symbols on instruments hazards See safety IMPORTANT description 8 installation category 71 instrument operation safety 70 M moving and lifting safety instrument 70 moving parts safety 70 MSDS See SDS Index 0 Ordering Information 49 overvoltage category rating 71 P physical hazard safety 70 pressurized fluids safety 70 R radioactive waste handling 75 repetitive motion safety 71 S safety before operating the instrument 69 chemical 73 chemical waste 74 electrical 70 ergonomic 71 guidelines 73 74 75 instrument operation 70 moving and lifting instrument 70 moving parts 70 physical hazard 70 pressurized fluids See also compressed gases safety repetitive m
41. eads settle before step 6 swirl the aqueous phase again then perform step 6 Immediately add the beads to the aqueous phase then swirl the AQ Tube until the suspension is uniformly brown Immediately load the AQ Tube onto the Emulsifier then attach the tubing to the cap of the AQ Tube see Figure 10 on page 29 Ensure that the beads have not settled CAUTION PINCH HAZARD Moving Parts Moving parts can crush AK pinch and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Start the run a Press the scale selection button to select the E20 scale on the instrument key panel see Figure 11 on page 31 b Press the start button G The motor starts and runs for 10 seconds before the pump starts The middle light on the panel is lit yellow IMPORTANT Do not open the door of the emulsifier while a run is in progress Opening the door automatically aborts the run and the emulsion is not properly prepared If you open the door in error set up and prepare the emulsion again see Prepare the P1 Beads E20 on page 26 The run time is 10 minutes After the run is complete the Emulsifier stops automatically the right light on the panel is lit green and the panel says Complete Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 3 Prepare and Run the Emulsifier E20 D Prepare and run the Emulsifier E20 IMPORTANT To stop the run pr
42. er Getting Started Guide Part no 4443496 The enriched beads are deposited on a slide and the library sequenced see Figure 1 Figure 1 The SOLiD9 EZ Bead System workflow Templated Beads Lib Salad for sequencing Amplifier Enricher Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 7 About This Guide Prerequisites Prerequisites This service guide is intended for service personnel who have been specifically trained by Applied Biosystems Applied Biosystems is not liable for damage or injury that results from the use of this manual by unauthorized or untrained parties This guide uses conventions and terminology that assume a working knowledge of the Microsoft Windows operating system the Internet and Internet based browsers Safety information Safety alert words SDSs Safety labels on instruments Note For general safety information see this section and Appendix E Safety on page 67 When a hazard symbol and hazard type appear by an instrument hazard see the Safety Appendix for the complete alert on the instrument Four safety alert words appear in Life Technologies user documentation at points in the document where you need to be aware of relevant hazards Each alert word IMPORTANT CAUTION WARNING DANGER implies a particular level of Observation or action as defined below IMPORTANT Indicates information that is necessary for proper instrument operation
43. er information that appears in the SDS of interest Select the language of your choice then click Search 3 Find the document of interest right click the document title then select any of the following e Open To view the document Print Target To print the document Save Target As To download a PDF version of the document to a destination that you choose Note For the SDSs of chemicals not distributed by Applied Biosystems contact the chemical manufacturer Chemical waste hazards T CAUTION HAZARDOUS WASTE Refer to Safety Data Sheets and local regulations for handling and disposal WARNING CHEMICAL WASTE HAZARD Wastes produced by Applied Biosystems instruments are potentially hazardous and can cause injury illness or death A WARNING CHEMICAL STORAGE HAZARD Never collect or store waste in a glass container because of the risk of breaking or shattering Reagent and waste bottles can crack and leak Each waste bottle should be secured in a low density polyethylene safety container with the cover fastened and the handles locked in the upright position Wear appropriate eyewear clothing and gloves when handling reagent and waste bottles Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Appendix E Safety E Chemical safety Chemical waste safety guidelines To minimize the hazards of chemical waste Read and understand the Safety Data Sheets SDSs provided by the manufacturers
44. er to 5500 Series SOLiD Systems Fragment Library Preparation User Guide Part no 4460960 and 5500 Series SOLiD Systems Mate Paired Library Preparation User Guide Part no 4460958 Optional Titrate the library The optimal titration point of most libraries is between 0 6 pM and 1 0 pM We recommend starting with a titration point of 0 8 pM for a new library Increase or decrease the titration as needed Install the SOLiD EZ Bead Emulsifier in a different location than the location used to enrich the emulsion Emulsification and enrichment in different locations prevent contamination of the sample To prevent contamination do not install the Emulsifier in the same room as the SOLiD EZ Bead Amplifier and SOLiD EZ Bead Enricher For more information refer to the Applied Biosystems SOLiD EZ Bead Emulsifier Site Preparation Guide Part no 4452499 and the Applied Biosystems SOLiD EZ Bead Enricher Site Preparation Guide Part no 4454036 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 13 Pouch Chapter 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Prepare the Oil Master Mix Note A single Emulsifier Oil Pack Kit Part no 4457185 yields approximately 240 mL of Oil Master Mix which is six 50 mL Falcon tubes containing 40 mL of Oil Master Mix each The 240 mL of Oil Master Mix is enough for E10 scale 8 runs E10 emulsions E10 bag ri
45. eractions with instrument or software that detrimentally affects normal operations Without limiting the abovementioned and for avoidance of doubt computer hardware monitors accessories software or other products not purchased from or supplied by Applied Biosystems Non AB Product are not covered under the foregoing warranty even if such Non AB Product is integral to functional use of an Applied Biosystems product Warranty limitations THE FOREGOING PROVISIONS SET FORTH APPLIED BIOSYSTEMS SOLE AND EXCLUSIVE REPRESENTATIONS WARRANTIES AND OBLIGATIONS WITH RESPECT TO ITS PRODUCTS AND APPLIED BIOSYSTEMS MAKES NO OTHER WARRANTY OF ANY KIND WHATSOEVER EXPRESSED OR IMPLIED INCLUDING WITHOUT LIMITATION WARRANTIES OF MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE WHETHER ARISING FROM A STATUTE OR OTHERWISE IN LAW OR FROM A COURSE OF DEALING OR USAGE OF TRADE ALL OF WHICH ARE EXPRESSLY DISCLAIMED THE REMEDIES PROVIDED HEREIN ARE THE BUYER S SOLE AND EXCLUSIVE REMEDIES WITHOUT LIMITING THE GENERALITY OF THE FOREGOING IN NO EVENT SHALL APPLIED BIOSYSTEMS BE LIABLE WHETHER IN CONTRACT TORT WARRANTY OR UNDER ANY STATUTE INCLUDING WITHOUT LIMITATION ANY TRADE PRACTICE UNFAIR COMPETITION OR OTHER STATUTE OF SIMILAR IMPORT OR ON ANY OTHER BASIS FOR DIRECT INDIRECT PUNITIVE INCIDENTAL MULTIPLE CONSEQUENTIAL OR SPECIAL DAMAGES SUSTAINED BY THE BUYER OR ANY OTHER PERSON OR ENTITY WHETHER OR NOT FORESEEABLE AND WHETHER OR NOT A
46. ess the stop button then press the stop button again to confirm that you want to stop the run The left light on the panel is lit red and the panel says Run aborted Stop the run only if the Emulsifier malfunctions If you stop the run before the run is complete then discard the emulsion because it is not properly prepared Troubleshoot if necessary see Appendix B Troubleshooting on page 57 then prepare and install the reagents again for the appropriate scale of emulsification Figure 11 How to use the Emulsifier Emulsification scales Select scale of run Stop run Start run Light color Meaning Green Run is complete Yellow Instrument is running Red Run aborted failed run or error Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 31 3 Chapter 3 Prepare and Run the Emulsifier E20 Prepare and run the Emulsifier E20 Collect the emulsion E20 32 Qus 5 0 D 10 11 Ensure that the run is finished When the run is finished the clock says 00 00 Turn the knob clockwise to unlock the compartment door then open the door Slide the coupler up and away from the impeller head Gently pull the impeller head from the coupler Swing out the arm at the base then remove the impeller and the jar from the compartment Detach the tubing from the port on the cap of the jar Remove the pump cassette tubing and AQ Tube from the instrument as one unit To release the tub
47. ey panel see Figure 6 on page 23 b Press the start button O The motor starts and runs for 10 seconds before the pump starts The middle light on the panel is lit yellow IMPORTANT Do not open the door of the emulsifier while a run is in progress Opening the door automatically aborts the run and the emulsion is not properly prepared If you open the door in error set up and prepare the emulsion again see Prepare the P1 Beads E10 on page 18 The run time is 10 minutes After the run is complete the Emulsifier stops automatically the right light on the panel is lit green and the panel says Complete IMPORTANT To stop the run press the stop button then press the stop button again to confirm that you want to stop the run The left light on the panel is lit red and the panel says Run aborted Stop the run only if the Emulsifier malfunctions If you stop the run before the run is complete then discard the emulsion because it is not properly prepared Troubleshoot if necessary see Appendix B Troubleshooting on page 57 then prepare and install the reagents again for the appropriate scale of emulsification Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Collect the 1 emulsion E10 e 10 11 SL A CES M Chapter 2 Prepare and Run the Emulsifier E10 Prepare and run the Emulsifier E10 Figure 6 How to use the Emulsifier Emulsification scales
48. fier Reagent 1X TEX Buffer e SOLID EZ Bead Emulsifier Reagent 1X Low TE Buffer SOLiD EZ Bead Emulsifier Oil Pack Kit 4457185 SOLID EZ Bead Emulsifier Oil Kit e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 1 e SOLID EZ Bead Emulsifier Oil Kit Stabilizer 2 e SOLID EZ Bead Emulsifier Oil Kit Oil SOLID EZ Bead Emulsifier Oil Accessory Kit e SOLID EZ Bead Emulsifier Oil Kit Oil Mix Bottle e SOLID EZ Bead Emulsifier Oil Kit Oil Funnel SOLIDO EZ Bead Emulsifier E10 Accessories Kit 4453065 SOLID EZ Bead Emulsifier E Small Jar Pack SOLID EZ Bead Emulsifier E Small AQ Tube Pack SOLID EZ Bead Emulsifier Tubing Pack SOLIDO EZ Bead E10 System Consumables 4 pack 4453093 t SOLID EZ Bead Emulsifier E10 Reagent Kit SOLID EZ Bead Emulsifier E10 Accessories Kit SOLID EZ Bead Amplifier E10 Accessories Kit SOLID EZ Bead Enricher E10 Reagent Kit SOLID EZ Bead Enricher Buffer Kit SOLID EZ Bead Enricher Accessories Kit t Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 49 Appendix A Ordering Information Required kits E20 T The kit includes consumables for all three instruments of the SOLiD EZ Bead System the Enricher Amplifier and Emulsifier Required kits E20 Required Applied Biosystems kits E20 Item part number Components
49. g without limitation quality control and commercial services such as reporting the results of purchaser s activities for a fee or other form of consideration For information on obtaining additional rights please contact outlicensing alifetech com or Out Licensing Life Technologies 5791 Van Allen Way Carlsbad California 92008 TRADEMARKS The trademarks mentioned herein are the property of Life Technologies Corporation or their respective owners Microsoft and Windows are registered trademarks of Microsoft Corporation in the United States and other countries Covaris is a registered trademark of Covaris Inc Latitude is a registered trademark of Dell 2011 Life Technologies Corporation All rights reserved Part Number 4441486 Rev E October 2011 Contents About This Guide iaa 7 PURPOSE A ETT 7 Prerequisites cats totes nt iub te enr ntt cee heb rte Gr nr Auteur de 8 Safety Informations sista ea ace ti ia al cele nt ete eR NEP E e ad 8 Safety alert Words rss arriere e phe wink A ag Boe es E A RES 8 SDSS cides ae le ee ire ned uites ba dotes oe Bet ne PO be ee 8 Safety labels on instruments 0 000 cc cece cece cece eee en 8 B CHAPTER 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch 11 Review the general workflow to prepare and sequence a library 12 Choose the scale of preparation of templated beads
50. he jar rests on the back of the groove and the arm is against the jar 6 Fit the impeller head into the locking slot Slide the coupler down until the coupler clicks to a stop to ensure that the impeller head is firmly in place in the coupler see Table 4 Table 4 How to fit the impeller head into the locking slot Locking slot Impeller ead Coupler Impeller head Coupler Impeller 1 Align the locking slot with the impeller head 2 Fit the impeller head into the groove below the coupler 3 Slide the coupler over the impeller head 7 Attach the tubing with the Luer Lok to the port of the cap 8 Insert the 3 stop tubing with one stop in the pump cassette see Table 5 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 27 Chapter 3 Prepare and Run the Emulsifier E20 Prepare and run the Emulsifier E20 Table 5 How to insert the tubing into the pump cassette 1 Using your right hand hold the pump cassette with your thumb and forefinger 2 Using your left hand grasp the tubing with your thumb and forefinger above the black stop 3 Pull the tubing slightly to slide the tubing into the pump cassette 4 Slide the tubing until the stop is aligned with the stop fixture on the outside edge of the pump cassette 5 Pull the tubing slightly towards the inside of the pump cassette to fit the stop snugly into the fixture a Repea
51. ide Chapter 5 Prepare and Run the Emulsifier E120 5 Prepare and run the Emulsifier E120 e Rotate the jar so that the port on the cap is to the left of the impeller shaft Figure 17 Rest the arm against the jar 6 Fit the impeller head into the locking slot Slide the coupler down until the coupler clicks to a stop to ensure that the impeller head is firmly in place in the coupler see Table 8 Table8 How to fit the impeller head into the locking slot Coupler Coupler Locking slot Impeller head Locking slot Locking slot Impeller head Impeller head 1 Align the locking slot with the impeller 2 Fit the impeller head into the groove 3 Slide the coupler over the impeller head below the coupler head 7 Attach the tubing with the Luer Lok to the port of the cap 8 Insert the 3 stop tubing with one stop in the pump cassette see Table 9 on page 44 Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 43 Chapter 5 Prepare and Run the Emulsifier E120 Prepare and run the Emulsifier E120 Table 9 How to insert the tubing into the pump cassette 1 Using your right hand hold the pump cassette with your thumb and forefinger 2 Using your left hand grasp the tubing with your thumb and forefinger above the black stop 3 Pull the tubing slightly to slide the tubing into the pump cassette 4 Slide the tubing until the stop is aligned with the stop fixtu
52. ide Prepare and Run the Emulsifier E20 This chapter explains how to prepare and run the Emulsifier for theE20 scale Choose the scale of preparation of templated beads on page 13 l Prepare the Oil Master Mix on page 14 Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 l Prepare the P1 Beads E20 on page 26 l Install the oil phase E20 on page 26 Prepare the aqueous phase E20 on page 29 l Set up and run the Emulsifier E20 on page 30 l Collect the emulsion E20 on page 32 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 25 3 Chapter 3 Prepare and Run the Emulsifier E20 Prepare and run the Emulsifier E20 Prepare and run the Emulsifier E20 Prepare the P1 Beads E20 Install the oil phase E20 26 Note You can thaw the Emulsifier reagents on ice as you prepare the P1 Beads and install the oil phase see Prepare the aqueous phase E20 on page 29 If you are amplifying the library with the SOLID EZ Bead Amplifier then it is recommended to pre rinse the pouches used with the Amplifier before emulsifying the library see Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 Vortex and pulse spin one tube of Emulsifier E20 P1 Beads Ensure that the P1 Beads are completely resuspended then pipet 330 uL of P1 Beads into a 1 5 mL DNA LoBind Tube Place the tube of beads
53. ifier has an installation overvoltage category of II and is classified as portable equipment Correct ergonomic configuration of your workstation can reduce or prevent effects such as fatigue pain and strain Minimize or eliminate these effects by configuring your workstation to promote neutral or relaxed working positions CAUTION MUSCULOSKELETAL AND REPETITIVE MOTION HAZARD These hazards are caused by potential risk factors that include but are not limited to repetitive motion awkward posture forceful exertion holding static unhealthy positions contact pressure and other workstation environmental factors To minimize musculoskeletal and repetitive motion risks Use equipment that comfortably supports you in neutral working positions and allows adequate accessibility to the keyboard monitor and mouse Position the keyboard mouse and monitor to promote relaxed body and head postures This section provides information on e U S and Canadian safety standards Canadian EMC standard European safety and EMC standards Australian EMC Standards U S and Canadian safety standards c UL us The Emulsifier has been tested to and complies with standard Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 71 72 Appendix E Safety Instrumentation safety UL 61010 1 2nd Edition CSA C22 2 No 61010 1 Safety Requirements for Electrical Equipment for Measurement Control and Laboratory Use
54. in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in 330 uL Emulsifier E20 BB Solution Sonicate the beads using the Bead Block Declump program on the Covaris 52 System for program conditions see Bead Block Declump on page 61 then Ipulse spin the tube Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant Resuspend the beads in 330 uL of 1X TEX Buffer Store the beads at 4 C until ready for use Unscrew and retain the cap with impeller of the Emulsifier E Small Jar Tare the E Small Jar on an analytical balance Pour 15 7 g of Oil Master Mix into the E Small Jar see Prepare the Oil Master Mix on page 14 Screw the cap with the impeller on the jar Install the jar on the Emulsifier a Turn the knob clockwise to open the compartment door b Pull the arm back at the base towards the opening of the compartment c Slide the jar into the groove of the base of the compartment until the jar stops against the back of the groove d Allow the arm to rest against the jar see Figure 7 e Rotate the jar so that the port on the cap is to the left of the impeller shaft Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Chapter 2 Prepare and Run the Emulsifier E20 D Prepare and run the Emulsifier E20 Figure 7 T
55. ing from the pump cassette turn the pinch wheel clockwise to the silver zone then remove the tubing Discard the used AQ Tube and tubing Store the pump cassette on top of the Emulsifier IMPORTANT The pump cassette is part of the instrument and is not disposable Close the door of the compartment then turn the knob counter clockwise to lock the door Immediately perform emulsion PCR ePCR on the emulsion using the SOLiD EZ Bead Amplifier refer to the Applied Biosystems SOLiD EZ Bead Amplifier Getting Started Guide Part no 4443494 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Prepare and Run the Emulsifier E80 This chapter explains how to prepare and run the Emulsifier for the E80 scale Choose the scale of preparation of templated beads on page 13 Prepare the Oil Master Mix on page 14 l Pre rinse the SOLID EZ Bead Amplifier Pouch on page 16 l Prepare the P1 Beads E80 on page 34 Install the oil phase E80 on page 34 l Prepare the aqueous phase E80 on page 37 l Set up and run the Emulsifier E80 on page 38 Collect the emulsion E80 on page 39 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 33 4 Chapter 4 Prepare and Run the Emulsifier E80 Prepare and run the Emulsifier E80 Prepare and run the Emulsifier E80 Prepare the P1 Beads E80 Install the oil phase
56. ly position pinch wheel 4 Swirl the AQ Tube until the suspension is uniformly brown then immediately load the AQ tube in instrument 5 Restart run Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 57 B Appendix B Troubleshooting 58 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide The Covaris S2 System This appendix covers m Operation notes ssssss 60 Bill the tank ip Ie a ek mI REIR OS ghey ee SU IEEE 60 De gas the Water ssh see me ese eh as di e Magma ns 60 Set the chillers aE e eR ELE AERE IA A 60 Perform required maintenance of the Covaris S2 System 60 R Covaris S2 programs 25 uen otek Boece ta he Eu ue t atat a Eu ge 61 bead Dlo ck Declurmp i eso gets Gea Du aee e c e ctetu 61 Covalent Decl mp lisse reete eter d nere bt at ete 61 Covalent Declump Bet iore aa e ta a cr PER nt de 61 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 59 Appendix C The Covaris S2 System Operation notes Operation notes Note For important instrument safety information refer to the Covaris 2 System manual Fill the tank Fill the tank with fresh deionized water to the proper fill line fill line level 15 is recommended The water should cover the visible part of the tube De gas the water De gas the water for 30 minutes To maintain de gassed water keep the pump continuo
57. matically aborts the run and the emulsion is not properly prepared If you open the door in error set up and prepare the emulsion again see Prepare the P1 Beads E80 on page 34 The run time is 15 minutes After the run is complete the SOLiD EZ Bead Emulsifier stops automatically the right light on the panel is lit green and the panel says Complete IMPORTANT To stop the run press the stop button e then press the stop button again to confirm that you want to stop the run The left light on the panel is lit red and the panel says Run aborted Stop the run only if the SOLiD EZ Bead Emulsifier malfunctions If you stop the run before the run is complete then discard the emulsion because it is not properly prepared Troubleshoot if necessary see Appendix B Troubleshooting on page 57 then prepare and install the reagents again for the appropriate scale of emulsification Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Collect the emulsion E80 Chapter 4 Prepare and Run the Emulsifier E80 7 Prepare and run the Emulsifier E80 Figure 16 How to use the Emulsifier Emulsification scales Select scale of run Stop run Start run Light color Meaning Green Run is complete Yellow Instrument is running Red Run aborted failed run or error QU ev E dh 9 10 11 Ensure that the run is finished When the run is finished the clock says 00 00
58. ns see Covalent Declump 3 on page 61 b Pulse spin the tube IMPORTANT Perform steps 5 and 6 below as quickly as possible before the beads settle in the aqueous phase If the beads settle before step 6 swirl the aqueous phase again then perform step 6 Immediately add the beads to the aqueous phase then swirl the AQ Tube until the suspension is uniformly brown Immediately load the AQ Tube onto the Emulsifier then attach the tubing to the cap of the AQ Tube see Figure 20 on page 45 Ensure that the beads have not settled CAUTION PINCH HAZARD Moving Parts Moving parts can crush AS pinch and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Start the run a Press the scale selection button to select the E120 scale on the instrument key panel see Figure 21 on page 47 b Press the start button G The motor starts and runs for 10 seconds before the pump starts The middle light on the panel is lit yellow IMPORTANT Do not open the door of the emulsifier while a run is in progress Opening the door automatically aborts the run and the emulsion is not properly prepared If you open the door in error set up and prepare the emulsion again see Prepare the P1 Beads E120 on page 42 The run time is 15 minutes After the run is complete the SOLiD EZ Bead Emulsifier stops automatically the right light on the panel is lit green and the panel says
59. nses _OR E20 scale 6 runs E20 emulsions E20 bag rinses _OR E80 scale 2 runs E80 emulsions E80 bag rinses _OR E120 scale 2 runs E120 emulsions E120 bag rinse 1 Prepare the Oil Master Mix in a 50 mL Falcon tube Item Volume Stabilizer 1 1 8 mL Stabilizer 2 400 uL Mineral oil 37 8 mL Total volume 40 mL a Use a 3 mL syringe to dispense 1 8 mL of Stabilizer 1 into the 50 mL Falcon tube see Figure 1 To ensure no air bubbles are in the syringe aspirate some reagent into the syringe and fully dispense it back into the reagent bottle Then slowly aspirate the correct volume Use a 1 mL syringe to dispense 400 uL of Stabilizer 2 into the 50 mL Falcon tube see Figure 1 Make sure no air bubbles are in the syringe follow the procedure in step 1a Table 1 Syringes with Stabilizers 1 and 2 Stabilizer 1 1 8 mL e srl T E C Stabilizer 2 400 uL 14 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch 2 c Pour mineral oil approximately 37 8 mL into the tube that has Stabilizers 1 and 2 so that the final volume is 40 mL Cap the Falcon tube Use a vortexer to mix the solution so that a whirlpool down to the bottom of the tube is created see Figure Figure table titles are not recommended in procedu
60. nt Optional equipment Optional equipment Item Source SOLID EZ Bead Amplifier Applied Biosystems The system includes 4448418 SOLID EZ Bead Amplifier 220 V power cable Pouch filling station Pouch frame Netbook and cables USB cord connecting cord Applied Biosystems SOLID EZ Bead AmplifierGetting Started Guide and Applied Biosystems SOLiD EZ Bead Amplifier Quick Reference Card SOLID EZ Bead Enricher Applied Biosystems The system includes 4448420 SOLID EZ Bead Enricher 220 V power cable Waste bottlet Small bottle racks with bottles attached Large bottle racks with bottles attached Laptop and cables USB cord connecting cord Applied Biosystems SOLiD9 EZ Bead Enricher Getting Started Guide and Applied Biosystems SOLiD9 EZ Bead Enricher Quick Reference Card Applied Biosystems has validated this protocol using this specific material Substitution may adversely affect system performance Supplied inside the instrument 56 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Troubleshooting Observation Possible cause Recommended action Aqueous phase not pumped into oil Tubing or pump cassette not properly If no aqueous phase has reached the phase installed oil phase 1 Press stop button twice to stop run 2 Correctly install tubing in pump cassette see the Install the oil phase procedure on page 18 page 26 or page 34 3 Correct
61. or to providing service or technical support For systems that have built in computers or processing units installing unauthorized hardware or software may void the Warranty or Service Plan Limited product warranty 64 Limited warranty Applied Biosystems warrants that all standard components of its SOLID EZ Bead Emulsifier SOLiD EZ Bead Amplifier or SOLiD EZ Bead Enricher will be free of defects in materials and workmanship for a period of one 1 year from the date the warranty period begins Applied Biosystems will repair or replace at its discretion all defective components during this warranty period After this warranty period repairs and replacement components may be purchased from Applied Biosystems at its published rates Applied Biosystems also provides service agreements for post warranty coverage Applied Biosystems reserves the right to use new repaired or refurbished instruments or components for warranty and post warranty service agreement replacements Repair or replacement of products or components that are under warranty does not extend the original warranty period Applied Biosystems warrants that all optional accessories supplied with its SOLID EZ Bead Emulsifier SOLiD EZ Bead Amplifier or SOLiD EZ Bead Enricher such as peripherals will be free of defects in materials and workmanship for a period of ninety 90 days from the date the warranty begins Applied Biosystems will repair or replace at its
62. otion 71 solvents 70 standards 71 workstation 71 safety labels on instruments 8 69 safety standards 71 safety symbols on instruments 68 SDSs about 8 description 74 obtaining 74 77 Set up the EZ Bead Emulsifier 11 17 25 33 41 solvents safety 70 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 79 Index standards EMC 71 safety 71 symbols safety 68 T training information on 77 W WARNING description 8 waste disposal guidelines 75 waste profiles description 75 workstation safety 71 80 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Headquarters 5791 Van Allen Way Carlsbad CA 92008 USA Phone 1 760 603 7200 Toll Free in USA 800 955 6288 For support visit www appliedbiosystems com support technologies www lifetechnologies com
63. p with the impeller on the jar Install the jar on the Emulsifier a Turn the knob clockwise to open the compartment door b Pull the arm back at the base towards the opening of the compartment c Slide the jar until it stops against the two posts at the rear of the compartment d Allow the arm to rest against the jar see Figure 12 e Rotate the jar so that the port on the cap is to the left of the impeller shaft Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide Chapter 4 Prepare and Run the Emulsifier E80 7 Prepare and run the Emulsifier E80 Figure 12 Rest the arm against the jar 6 Fit the impeller head into the locking slot Slide the coupler down until the coupler clicks to a stop to ensure that the impeller head is firmly in place in the coupler see Table 6 Table 6 How to fit the impeller head into the locking slot Coupler Coupler Locking slot Impeller head Locking slot Locking slot Impeller head Impeller head 1 Align the locking slot with the impeller 2 Fit the impeller head into the groove 3 Slide the coupler over the impeller head below the coupler head 7 Attach the tubing with the Luer Lok to the port of the cap 8 Insert the 3 stop tubing with one stop in the pump cassette see Table 7 on page 36 Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 35 Chapter 4 Prepare and Run the Emulsifier E80 Prepare and run the
64. phase e Emulsifier E120 P1 reagent then vortex the reagent for a few seconds and E120 pulse spin e Emulsifier E120 P2 reagent then vortex the reagent for a few seconds and pulse spin Library template Place on ice Emulsifier E120 Aqueous Master mix Do not vortex 2 Place the Emulsifier E Large AQ Tube on ice for filling 3 Prepare a 1 10 dilution of P1 Reagent by adding 21 uL of P1 reagent to 189 uL of Low TE Buffer in a 1 5 mL DNA LoBind Tube 4 Add the appropriate volume to the AQ Tube in this order E120 Reagent Volume pL Aqueous Master Mix 48000 Xt 1 10 diluted P1 Reagent 200 Undiluted P2 Reagent 300 Library template X t X Volume of library template Use a Pipet Aid pipette fitted with a 50 mL serological pipette to dispense Aqueous Master Mix into the E Large AG Tube Applied Biosystems SOLIDO EZ Bead Emulsifier Getting Started Guide 45 5 Chapter 5 Prepare and Run the Emulsifier E120 Prepare and run the Emulsifier E120 Set up and run the Emulsifier E120 46 mv e dm Cap the AQ Tube and swirl to mix the solution Place the AQ Tube back on ice Power on the Emulsifier Ensure that the Oil Master Mix in the Emulsifier Jar is at room temperature Vortex then pulse spin the tube of prepared P1 beads Sonication of P1 beads a Sonicate the prepared P1 Beads two times using the Covalent Declump 3 program on the Covaris S2 System for program conditio
65. re on the outside edge of the pump cassette 5 Pull the tubing slightly towards the inside of the pump cassette to fit the stop snugly into the fixture a Repeat step 8 to insert the tubing with the second stop at the other end of the pump cassette Ensure that the tubing is in the inner groove opposite the second stop see Table 9 and that the tubing is in the correct orientation see Figure 18 Figure 18 Correct orientation of the Emulsifier Tubing b Install the pump cassette into the roller head of the pump see Figure 20 on page 45 c Turn the pinch wheel in the pump cassette counter clockwise so that the mark is in the blue zone see Figure 19 This adjustment maintains correct tubing tension for normal pump operation Figure 19 Turn the pinch wheel in the pump cassette so that the mark is in the blue zone 9 Position the tubing so that it threads through the tubing tab see Figure 20 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 5 Prepare and Run the Emulsifier E120 5 Prepare and run the Emulsifier E120 10 Close the door of the compartment then turn the knob counter clockwise to lock the door see Figure 20 Figure 20 Placement of Emulsifier E Large AQ Tube Tubing and E Large Jar Tubing tab SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier E Large AQ Tube Tubing in pump cassette E Large Jar Prepare the 1 Thaw on ice aqueous
66. res After the whirlpool has been created continue to whirlpool vortex the mixture for another 30 seconds 45 40 as x 2 wd zii _ HS dl Whirlpool created Whirlpool not created during vortexing Incorrect method Correct method 3 4 Pour the mixture into the Oil Mix Bottle in the Emulsifier Oil Pack Kit Repeat steps 1 through 3 FIVE additional times Use a new 50 mL Falcon tube each time After performing steps 1 through 3 a total of SIX times you should have approximately 240 mL of Oil Master Mix in your Oil Mix Bottle Shake by hand the bottled mix for 30 seconds If the Oil Master Mix will be used the same day De gas at room temperature for at least 30 minutes in a dark location such as in a cupboard or drawer If the Oil Master Mix will NOT be used the same day De gas and store the Oil Master Mix at 4 C in a 4 C refrigerator STOPPING POINT The Oil Master Mix can be stored at 4 C for up to two weeks Before using the stored Oil Master Mix shake by hand the bottled mix for 30 seconds Let the Oil Master Mix reach room temperature and de gas for 1 hour in a dark location such as in a cupboard or drawer Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 15 Chapter 1 Choose the Scale of Preparation of Templated Beads Prepare the Oil Master Mix and Pre rinse the Amplifier Pouch You have completed preparation of the SOLiD EZ Bead Oil Mas
67. rranty Warranty period Any applicable warranty period under these sections begins on the earlier of the date effective date of installation or ninety 90 days from the date of shipment for hardware and software installed by Applied Biosystems personnel For all hardware and software installed by the buyer or anyone other than Applied Biosystems and for all other products the applicable warranty period begins the date the product is delivered to the buyer Warranty claims Warranty claims must be made within the applicable warranty period or for chemicals or other consumable products within thirty 30 days after receipt by the buyer Warranty The above warranties do not apply to defects resulting from misuse neglect or exceptions accident including without limitation operation with incompatible solvents or samples in the system operation outside of the environmental or use specifications or not in conformance with the instructions for the instrument system software or accessories improper or inadequate maintenance by the user installation of software or interfacing or use in combination with software or products not supplied or authorized by Applied Biosystems modification or repair of the product not authorized by Applied Biosystems relocation or movement of the instrument by Customer or any third party not acting on behalf of AB or intrusive activity including without limitation computer viruses hackers or other unauthorized int
68. s the limitation or exclusion of warranties liability remedies or damages set forth above shall apply to the fullest extent permitted by law and shall not apply to the extent prohibited by law Damages claims and returns Damages Claims Returns 66 If shipping damage to the product is discovered contact the shipping carrier and request inspection by a local agent Secure a written report of the findings to support any claim Do not return damaged goods to Applied Biosystems without first securing an inspection report and contacting Applied Biosystems Technical Support for a Return Authorization RA number After a damage inspection report is received by Applied Biosystems Applied Biosystems will process the claim unless other instructions are provided Do not return any material without prior notification and authorization If for any reason it becomes necessary to return material to Applied Biosystems contact Applied Biosystems Technical Support or your nearest Applied Biosystems subsidiary or distributor for a return authorization RA number and forwarding address Place the RA number in a prominent location on the outside of the shipping container and return the material to the address designated by the Applied Biosystems representative Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Safety This appendix covers B Instrumentation safety 6
69. sferred transported and disposed of according to all local state provincial and or national regulations IMPORTANT Radioactive materials may require special handling and disposal limitations may apply Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 75 E Appendix E Safety Safety alerts Safety alerts Specific alerts for instrumentation 76 For the definitions of the alert words IMPORTANT CAUTION WARNING and DANGER see Safety alert words on page 8 CAUTION PINCH HAZARD Moving Parts Moving parts can crush pinch EEN and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Applied Biosystems SOLID EZ Bead Emulsifier Getting Started Guide Documentation and Support Related documentation For the latest documentation on the SOLiD EZ Bead System and SOLiD System go to http www3 appliedbiosystems com AB_Home Support index htm Obtaining support For the latest services and support information for all locations go to www lifetechnologies com At the Life Technologies web site you can e Access worldwide telephone and fax numbers to contact Life Technologies Technical Support and Sales facilities Search through frequently asked questions FAQs Submit a question directly to Technical Support Order Life Technologies user documents SDSs certificates of analysis and other related documents Download PDF
70. t in personal injury or damage to the instrument T WARNING PHYSICAL INJURY HAZARD Using this product in a manner not specified by Applied Biosystems may result in personal injury or damage to the instrument Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide 69 E Appendix E Safety Instrumentation safety Physical hazard safety Electrical safety 70 Moving and lifting the instrument CAUTION PHYSICAL INJURY HAZARD The instrument is to be moved and positioned only by the personnel or vendor specified in the applicable site preparation guide If you decide to lift or move the instrument after it has been installed do not attempt to lift or move the instrument without the assistance of others the use of appropriate moving equipment and proper lifting techniques Improper lifting can cause painful and permanent back injury Depending on the weight moving or lifting an instrument may require two or more persons Operating the instrument Ensure that anyone who operates the instrument has Received instructions in both general safety practices for laboratories and specific safety practices for the instrument Read and understood all applicable Safety Data Sheets SDSs See About SDSs on page 74 Cleaning or decontaminating the instrument T CAUTION Before using a cleaning or decontamination method other than those recommended by the manufacturer verify with the manufacturer that
71. t step 8 to insert the tubing with the second stop at the other end of the pump cassette Ensure that the tubing is in the inner groove opposite the second stop see Table 5 and that the tubing is in the correct orientation see Figure 8 b Install the pump cassette into the roller head of the pump see Figure 10 on page 29 Figure 8 Correct orientation of the Emulsifier Tubing c Turn the pinch wheel in the pump cassette counter clockwise so that the mark is in the blue zone see Figure 9 on page 28 This adjustment maintains correct tubing tension for normal pump operation Figure 9 Turn the pinch wheel in the pump cassette so that the mark is in the blue zone 9 Position the tubing so that it threads through the tubing tab see Figure 10 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Chapter 2 Prepare and Run the Emulsifier E20 D Prepare and run the Emulsifier E20 10 Close the door of the compartment then turn the knob counter clockwise to lock the door see Figure 10 Figure 10 Placement of Emulsifier E 5mall AQ Tube Tubing and E Small Jar Tubing tab SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier SOLID EZ Bead Emulsifier E Small AQ Tube Tubing in pump cassette E Small Jar Prepare the 1 Thaw on ice aqueous phase e Emulsifier E20 P1 reagent then vortex the reagent for a few seconds and E20 pulse spin e Emulsifier E20 P2 reagent then vortex the reagent for a few
72. tarted Guide 17 Chapter 2 Prepare and Run the Emulsifier E10 Prepare and run the Emulsifier E10 Prepare and run the Emulsifier E10 Note You can thaw the Emulsifier reagents on ice as you prepare the P1 Beads and install the oil phase see Prepare the aqueous phase E10 on page 21 Prepare the P1 1 If you are amplifying the library with the SOLiD EZ Bead Amplifier then it is Beads E10 recommended to pre rinse the pouches used with the Amplifier before emulsifying the library see Pre rinse the SOLiD EZ Bead Amplifier Pouch on page 16 2 Vortex and pulse spin one tube of Emulsifier E10 P1 Beads 3 Ensure that the P1 Beads are completely resuspended then pipet 220 uL of P1 Beads into a 1 5 mL DNA LoBind Tube 4 Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant 5 Resuspend the beads in 220 uL Emulsifier E10 BB Solution 6 Sonicate the beads using the Bead Block Declump program on the Covaris S2 System for program conditions see Bead Block Declump on page 61 then pulse spin the tube 7 Place the tube of beads in a magnetic rack for at least 1 minute After the solution clears remove and discard the supernatant 8 Resuspend the beads in 220 uL of 1X TEX Buffer 9 Store the beads at 4 C until ready for use Install the oil phase 1 Unscrew and retain the cap with impeller of the Emulsifier E
73. ter Mix Proceed to bead emulsification according to the scale of preparation that you want Chapter 2 Prepare and Run the Emulsifier E10 on page 17 Chapter 3 Prepare and Run the Emulsifier E20 on page 25 Chapter 4 Prepare and Run the Emulsifier E80 on page 33 Chapter 5 Prepare and Run the Emulsifier E120 on page 41 Pre rinse the SOLiD9 EZ Bead Amplifier Pouch IMPORTANT The Oil Master Mix can be stored at 4 C for up to two weeks Before using the stored Oil Master Mix shake by hand the bottled mix for 30 seconds Let the Oil Master Mix reach room temperature and de gas for 1 hour in a dark location such as in a cupboard or drawer IMPORTANT If you are amplifying the library with the SOLiD EZ Bead Amplifier then pre rinse the pouches used with the Amplifier before emulsifying the library 1 2 16 Unscrew the cap from the nozzle of the SOLiD EZ Bead Amplifier Pouch Use a Pipet Aid pipette and a serological pipette of the appropriate volume to add Oil Master Mix to the Amplifier Pouch see Volume of Emulsifier Reagents E10 E20 E80 or E120 Oil Master Mix to add to an Amplifier E Small or E Large Pouch Volume of Oil Master Scale Pouch to use Mix to add mL E10 E20 SOLID EZ Bead Amplifier E Small Pouch 10 E80 E120 SOLID EZ Bead Amplifier E Large Pouch 20 Tightly screw the cap onto the nozzle of the Amplifier Pouch to ensure that the pouch
74. the proposed method will not damage the equipment Pinch hazard CAUTION PINCH HAZARD Moving Parts Moving parts can crush pinch AS and cut Keep hands clear of moving parts while operating the instrument Disconnect power before servicing Solvents and pressurized fluids WARNING PHYSICAL INJURY HAZARD Always wear eye protection when working with solvents or any pressurized fluids WARNING ELECTRICAL SHOCK HAZARD Severe electrical shock can result from operating the SOLiD EZ Bead Emulsifier without its instrument panels in place Do not remove instrument panels High voltage contacts are exposed when instrument panels are removed from the instrument Power WARNING ELECTRICAL HAZARD Grounding circuit continuity is required for the safe operation of equipment Never operate equipment with the grounding conductor disconnected Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Workstation safety Safety and electromagnetic co mpatibility EMC standards Appendix E Safety E Instrumentation safety WARNING ELECTRICAL HAZARD Use properly configured and approved line cords for the voltage supply in your facility f WARNING ELECTRICAL HAZARD Plug the system into a properly grounded receptacle with adequate current capacity Fuses T CAUTION Replace only with the same TYPE and RATING FUSE Use only 250V 2A SB fuses when replacing the fuse Overvoltage rating The Emuls
75. tive clothing For additional safety guidelines consult the SDS Minimize the inhalation of chemicals Do not leave chemical containers open Use only with adequate ventilation for example fume hood For additional safety guidelines consult the SDS Check regularly for chemical leaks or spills If a leak or spill occurs follow the manufacturer s cleanup procedures as recommended in the SDS Comply with all local state provincial or national laws and regulations related to chemical storage handling and disposal Applied Biosystems SOLiD9 EZ Bead Emulsifier Getting Started Guide 73 E Appendix E Safety Chemical safety SDSs Chemical waste safety 74 About SDSs Chemical manufacturers supply current Safety Data Sheets SDSs with shipments of hazardous chemicals to new customers They also provide SDSs with the first shipment of a hazardous chemical to a customer after an SDS has been updated SDSs provide the safety information you need to store handle transport and dispose of the chemicals safely Each time you receive a new SDS packaged with a hazardous chemical be sure to replace the appropriate SDS in your files Obtaining SDSs The SDS for any chemical supplied by Life Technologies is available to you free 24 hours a day To obtain SDSs 1 Go to www appliedbiosystems com click Support then select SDS 2 In the Keyword Search field enter the chemical name product name SDS part number or oth
76. usly on during operation and sample processing Set the chiller Set the chiller temperature to between 2 C to 5 C to ensure that the temperature reading in the water bath displays 5 C The circulated water chiller should be supplemented with 20 ethylene glycol Perform required The Covaris S2 System requires regular maintenance to work properly Perform the maintenance of the tasks in the table below see Table 10 Covaris S2 System Table 10 Required maintenance of the Covaris S2 System Required maintenance task Frequency to perform task De gas water for 30 minutes prior to use Before every use Change water Daily Clean with bleach Every two weeks 60 Applied Biosystems SOLiD EZ Bead Emulsifier Getting Started Guide Covaris 62 programs Bead Block Declump Covalent Declump 1 Covalent Declump 3 Appendix C The Covaris S2 System Covaris S2 programs Table 11 Bead Block Declump 1 cycle Treatment 1 followed by 1 cycle Treatment 2 Treatment 1 Treatment 2 Duty Cycle 1 5 Intensity 5 5 Cycles Burst 50 100 Time 5 sec 60 sec Target wattage power 2 10 performance estimate W t Not programmed Table 12 Covalent Declump 1 1 cycle Treatment 1 followed by 1 cycle Treatment 2 Treatment 1 Treatment 2 Duty Cycle 2 5 Intensity 6 9 Cycles Burst 100 100 Time 5 sec 30 sec Target wattage power 4 15 performance estimate W
77. y which the instrument is switched on to the Standby condition Hazardous voltage may be present if this switch is on standby Indicates the On Off position of a push push main power switch Indicates a terminal that may be connected to the signal ground reference of another instrument This is not a protected ground terminal Indicates a protective grounding terminal that must be connected to earth ground before any other electrical connections are made to the instrument Indicates a terminal that can receive or supply alternating current or voltage Indicates a terminal that can receive or supply alternating or direct current or voltage IIDA OGO Safety symbols The following table describes the safety symbols that may be displayed on Life Technologies instruments Each symbol may appear by itself or with text that explains the relevant hazard see Safety labels on instruments on page 8 These safety symbols may also appear next to DANGERS WARNINGS and CAUTIONS that occur in the text of this and other product support documents Symbol Description Indicates that you should consult the manual for further information and to proceed with appropriate caution gt Indicates the presence of an electrical shock hazard and to proceed with appropriate caution Applied Biosystems SOLID EZ Bead Emulsifier Getting Started Guide Appendix E Safety E Instrumentation safety Symbol

SOLiD™ EZ Bead™ Emulsifier Getting Started Guide (PN 4441486E)

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SOLiD™ EZ Bead™ Emulsifier Getting Started Guide (PN 4441486E)