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BioTek Synergy HT Operator`s Manual

1. dO dO 0 0T gt 9q ISNW AD Ue W uonejnaq puepueys JO dO 0 0Z gt 3q SNW 10419 Adeunsoy b o09T 0 4 l Mm paypeadxy 6 yBiem 71s H 9 d gJ a D d Vv CT TT OT 6 wu 0SL 0 90 sao eed su ds q 1 S a4nqeUuBis 4O dO 4O d O 6 00r9 0 g pease 0 L gt 2g ISNW AD Ue W uonelnaq puepueis 0 S gt 9g ISNW 10449 AdDeinsoy 71uBlam paypedxy 3yB1am qn OZ 8 L 9 S wu 0SZ 0 90 sao 23a su ds q 1n oz SJUSLWWILWOD aqyeq Bulpeay N S Jepeoy 4O dO 30O d O 009S 2 2poW Japeoy O T gt aq ISNW AI Ue W uoiqelAaq puepueis 0 C gt 9g ISNW 10119 Adeinsoy 1uBlam paypedxy 1461am qn 08 qjo O aO wyHh4 O IE v E c T wu 0S Z SOvV sao 232a su ds q 1 os Jasuodsig S S L UOISID Jd 9 ADeAndoy su ds d LH ABsasuASs 122 Chapter 4 Instrument Qualification BioTek Instruments Inc Chapter 5 Preventive Maintenance This chapter provides step by step instructions for maintaining the Synergy HT and external dispense module if used in top condition to ensure that they continue to perform to specification Recommended Maintenance Schedule
2. Figure 2 Removing the shipping panel Figure 3 Storage pocket on the rear of the instrument wrench carrier shipping screw and warning tag shown BioTek Instruments Inc 3 Remove the Microplate Carrier Shipping Screw 13 3 Remove the Microplate Carrier Shipping Screw I mportant Remove the microplate carrier shipping screw before turning on the Synergy HT Perform these steps to remove the carrier shipping screw 1 Pull down the microplate loading door on the front of the reader 2 Using the supplied wrench remove the carrier shipping screw with its o ring and warning tag 3 Place the wrench screw o ring and tag in the plastic tool storage bag that you attached to the back of the reader Shipping screw with o ring Shipping screw warning tag Figure 4 Removing the microplate carrier shipping screw Important Replace the microplate carrier shipping screw before repackaging the Synergy HT for shipment Please contact BioTek if you have misplaced the screw PN 7092071 and or its o ring PN 49259 Synergy HT Operator s Manual 14 Chapter 2 Installation 4 Install the Fluorescence Lamp Assembly Important Do not touch the glass lenses Fingerprints on the condenser lens or heat absorber may negatively affect performance Warning The fluorescence lamp assembly is hot when the instrument is powered on If
3. cceeeeeee eee eee eae 124 OVOP WCW eaaa a EEO AEEA O Oaa e EACT As 124 Dispense Module ssssssssrrrrrssssreesrrrrrrrrrrrrrnnrrrnnsssrreeerrrt 124 Sched lE eraa en E E E E AEE ENAERE ES 124 Warnings amp Precautions cccceeee cece eee eee eee eee eee rnnt 126 Cleaning Exposed Surfaces 0c eceeeee eee eee ener teen eee ee eeaee ees 127 Inspecting Cleaning Excitation and Emission Filters 55 128 Flushing Purging the Fluid Path cceeceeeee eects eee eee eens 129 Running a Dispense Protocol Optional eceeeeeee eee eae 130 Emptying Cleaning the Tip Priming Trough ccce 132 Cleaning the Priming Plate c eeceeee eee e eee eee neces eeee ees 132 Cleaning the Internal Components cceeee cece eect teeta ees 133 Required Materials 2 cccecee cece eee ee teen eee eee eeeeee seen ees 134 Removing the Reader s Shroud ccceeeeee cece test eae eae eas 135 Removing the Internal Tubes and Injector Heads 137 Cleaning the Internal Tubes and Injector Heads 140 Cleaning the Optical Probes ceceeee eee eee ee renee enone 141 Cleaning the Reader s Internal Surface cceeeeeeeeeeee enue 149 Reassembling the Components eceeeeeeee eee e eae e eee eas 150 Performance Che cK 20 cc ccccccecececeeeeeeeeeeecneeeeeeanueeeneetannneers 151 1
4. ccceeeeeeee eee e ee ee seen tee eaeeeeees 133 Required MaterialS ccccccscecsceeeeeeeeeseeegeesgeeseeeseeeseeaeenanenanenags 134 Removing the Reader s SHroud ccceeceeeeee eee ee eee eee eee eeeeeeaeeae ed 135 Removing the Internal Tubes and Injector Heads eeeeeee 137 Cleaning the Internal Tubes and Injector Heads c eeeeeeee es 140 Cleaning the Optical Probes cccece cece eens eee eee eee eee e teeta ened 141 Cleaning the Reader s Internal Surface ccece eee eee eee eeeeee es 149 Reassembling the COMPONents eeceee eee ee eee tees eee teeta sean ea eed 150 Performance CHECK i sii cece siamania ianen EEEE NA outa ered aide caters 151 Appendix A Decontamination c cccscsscseeeeeeceeneeceeeeeeseuseanensensessenees 153 PUP DOS CSc ige gee ecstatic enna aliens aterm iain eel alee AEE KAR AE E AA aan eae ete eae 154 Required Material S iraire aE AA ARA N det veces eaves es 155 Procedure for Models without InjectorS ssssssssrrssssrnrrresurrrnrrsrsrrrrns 156 Synergy HT Operator s Manual vi Preface Routine Procedure for Models with Injectors cseceeeeeeeeeeeeeeeee ees 157 Clean Exposed Surfaces sssssssssessessessernsrrorrrerrnrrerrennnnnnnnnernes 157 Decontaminate the Fluid LINGS cece cee eee eee e eee eeeeeeeeeeeeeeeaeenas 158 Rinse the FIUId LINES cecceceecee cece sees a ea N AAA AAR 159 Clean the Internal Tu
5. 2 Note Perform this Routine Procedure when all systems are functioning normally on the Synergy HT with Injectors If you are unable to prime the Synergy HT due to a system failure perform the Alternate Procedure described on page 161 If disinfecting with sodium hypochlorite bleach be sure to flush repeatedly with deionized water to ensure that no bleach is carried over After disinfecting with sodium hypochlorite perform the rinse procedure provided on page 159 If disinfecting with alcohol do not immediately prime with deionized water because the drying effect of the alcohol is an important aspect of its disinfectant properties Clean Exposed Surfaces Turn off and unplug the instrument Prepare an aqueous solution of 0 50 sodium hypochlorite bleach As an alternative 70 isopropyl alcohol may be used if the effects of bleach are a concern Be sure to check the percent NaClO of the bleach you are using this information is printed on the side of the bottle Commercial bleach is typically 10 0 NaClO if this is the case prepare a 1 20 dilution Household bleach is typically 5 0 NaClO if this is the case prepare a 1 10 dilution Manually open the plate carrier door slide out the plate carrier Moisten a cloth with the bleach solution or alcohol Do not soak the cloth Wipe the plate carrier and the exposed surfaces of the external dispense module Wait 20 minutes Moisten a cloth with de
6. BioTek Instruments Inc Fluorescence Tests 97 Plate Format Dimensions To create a plate format in KC4 for the Greiner SensoPlate select System gt Plate Formats click New and add the information shown below Mate Descniphion Name Greiner SensoPlate Manufacturer Greiner Catalogue 655892 Length 127750 um Width 85350 pm Top Left x fi 4600 um Top Left Y f 1400 um Bottom Right fi 13650 pm Bottom Right Y 74200 um Nb Columns fi 2 Nb Rows Well Diameter 661 0 um Height um Cancel Once the plate format is created you can reference it in a protocol s Reading Parameters dialog Synergy HT Operator s Manual 98 Chapter 4 Instrument Qualification Fluorescence Tests Using Methylumbelliferone As an alternative to using Sodium Fluorescein Methylumbelliferone MUB can be used to test the top optics for the fluorescence system Required Materials e Anew clean 96 well solid black plate such as Corning Costar Mfr 3915 e Excitation filter 360 40 nm Emission filter 460 40 nm e Deionized or distilled water e Carbonate Bicarbonate buffer CBB capsules BioTek PN 98158 e 10mg vial of Methylumbelliferone MUB BioTek PN 98156 e 100 methanol BioTek PN 98161 e Aluminum foil e Various beakers graduated cylinders and pipettes e Optional but recommended 0 45 micron filter e Gen5 or KC4 Synergy HT FI_MUB prt proto
7. 1 Gently insert the stylus PN 2872304 into each injector head pipe to clear any blockages The stylus should be stored in a plastic cylinder affixed to the rear of the dispense module or reader 2 Stream water from a faucet through the pipe to be sure it is clean If the water does not stream out try soaking the heads in hot soapy water and then reinserting the stylus O ring do not remove BioTek Instruments Inc Cleaning the Internal Components 141 Cleaning the Optical Probes The optical probes should be cleaned at least quarterly They should also be cleaned if reagent has spilled and or if an unusually high background signal has been flagged by the assay controls typically blanks or negative controls Contaminated probes can lead to a loss of sensitivity e g instead of being able to meet the 10 pg ml concentration detection limit the instrument may only be able to meet 20 pg ml Another indicator is the CV in the Corners liquid test it may increase due to the Noise in the chamber from any spilled fluorescing compounds e To access the optical probes the first step is to unplug the reader and remove its shroud cover If you haven t already done this turn to page 135 now for instructions Note For models without injectors the internal chamber and probes are not customer accessible Contact BioT ek s Technical Assistance Center with any questions about your particular model e We
8. BioTek Instruments Inc
9. BioTek Instruments Inc KC4 Software 57 Purge Utility KC4 provides a special utility to purge fluid from the dispense tubing and syringe by pumping the fluid in reverse back into the supply bottle To purge the dispense module 1 InKC4 select System Reader Control and click the Dispenser tab 2 Select the Dispenser 1 or 2 associated with the supply bottle 3 Enter the desired purge Volume from 5 to 5000 uL 4 Selecta prime Rate in uL second 5 Click Purge to start the process Emptying the Tip Priming Trough Read amp Dispense protocols can specify atip prime to compensate for any fluid loss at the dispense tip due to evaporation since the last dispense Tip priming is performed in a small removable priming trough located in the left rear of the carrier see page 45 The trough holds up to 1 5 ml of liquid and must be periodically emptied and cleaned by the user Ifthe Tip Priming Trough overflows you should clean the microplate carrier and possibly the internal surface beneath the carrier See Chapter 5 Preventive Maintenance for instructions See also Appendix A for decontamination instructions KC4 must be told when the trough has been emptied To empty the tip priming trough 1 If the microplate carrier is inside the reader press the carrier eject button 2 Carefully remove the trough from the carrier and empty it Clean or decontaminate the trough if necessary 3 In KC4 se
10. Clean the priming plate regularly to prevent bacteria growth and residue buildup Wash the plate in hot soapy water using a small brush to clean in the corners if necessary Rinse thoroughly and allow it to dry completely BioTek Instruments Inc Cleaning the Internal Components 133 Cleaning the I nternal Components Applies only to Synergy HT modds with injectors The Synergy HT s internal components that require routine cleaning include e Optical probes e Surface beneath the microplate carrier e Internal dispense tubes and injector heads The internal components should be cleaned at least quarterly In addition if fluid has spilled inside the instrument and or if an unusually high background signal has been flagged by the assay controls typically blanks or negative controls the optical probes and the surface beneath the microplate carrier should be cleaned The procedures in this section should be performed in succession Start with Removing the Reader s Shroud and execute the procedures that meet your needs in the order in which they are presented Finish with Reassembling the Components We recommend running a System Test via Gen5 or KC4 before and after performing these cleaning procedures This will verify that all systems are functioning properly and allow you to compare results before and after maintenance Caution The buildup of deposits left by the evaporation of spilled fluids within
11. If using BioTek s sodium fluorescein powder PN 98155 be sure to hold the vial upright and open it carefully the material may be concentrated at the top If a centrifuge is available spin down the tube before opening When diluting the sodium fluorescein powder in buffer it takes time for the powder to completely dissolve Allow the solution to dissolve for 4 to 5 minutes with intermittent vortexing before preparing the titration dyes Wrap the vial containing the SF stock solution in foil to prevent exposure to light Discard the unused solution after seven days Discard any open unused buffer solution after seven days 1 The Sodium Borate solution does not require further preparation proceed to step 2 If you are using PBS prepare the solution now e Optional but recommended Using a 0 45 micron filter filter 200 mL of deionized or distilled water e Follow the manufacturer s instructions on the PBS packaging to create 200 mL dissolving the necessary amount of PBS into the filtered water e Stir the solution preferably using a stir table until the PBS is completely dissolved e Check the pH it should be between 7 2 and 7 6 at 25 C 2 Prepare the sodium fluorescein stock solution Add 2 0 mL of the PBS solution to the 1 mg Sodium Fluorescein SF vial This yields a 1 32 nM stock solution Ensure that the dye has completely dissolved and is well mixed 3 Carefully prepare the dilutions Label each
12. To create the Dispense protocol in Gen5 I 2 Select File New Protocol and then Protocol Procedure Add a Dispense step with the following parameters e Select Dispenser 1 e Set Tip Priming to Before this dispense step and Volume to 10 uL e Set the Dispense Volume to 100 uL or an amount to match your assay protocol e Select a Rate adjust the rate to support the dispensing volume e Click OK to close the dialog and add the Dispense step to the list Add another Dispense step with the same parameters selecting Dispenser 2 Add a quick Read step with the following parameters Gen5 requires that a Read step follow the Dispense step e Definea partial plate read on just one well eg A1 e Set the Detection Method to Absorbance e Set the Read Type to Endpoint e Set the Read Speed to Normal e Select any wavelength Click OK to close the dialog and add the Read step to the list Click OK to close the Procedure Select File Save As and give the protocol an identifying name such as Dispense Observation Select File New Experiment to run the Dispense Observation protocol Click the Read button and follow the prompts BioTek Instruments Inc 10 Running a Dispense Protocol Optional 131 When the procedure is complete visually assess the fluid level in the wells for accuracy If the well volume appears to be unevenly distributed clean the internal dispense tubes and injector heads as descri
13. Code Description and Probable Causes 1101 Failed configuration checksum test This error indicates that during self test or at the end of a plate read the checksums calculated for configuration flash memory page 1 do not match the saved checksums Probable Causes e The flash memory on the 7080400 PCB is defective or corrupt The basecode software may need to be re downloaded 1200 Autocalibration data missing for the Fluorescence top bottom and absorbance reads This error indicates that no autocal data for any of the three read locations bottom probe top probe absorbance Probable Causes e The 7080400 PCB was changed and the Flash memory does not have the calibration values loaded Performing the autocalibration procedure will correct this 130x lt Motor gt not homed successfully This error indicates that the lt motor gt is not at home At the beginning of the motor_position function the basecode checks to see if the motor is homed If it is not homed an error is displayed on the controlling PC and the function is terminated Probable Causes e If an error 0200 is ignored see the Probable Causes for 0200 Error Motor See Probable Causes for 1300 X axis 0200 1301 Y axis 0201 1302 EX motor 0202 1303 EM motor 0203 1304 Order sorting filter wheel 0204 1305 Monochromator 0405 1306 Probe Height 0206 1307 Probe changer 0207 1308 Syringe 1 0208 1309 Syringe 2 0209 Syner
14. Run two prime cycles for a total of 10000 pl Pause for 20 to 30 minutes to allow the solution to disinfect the tubing Remove the inlet tube from the beaker of disinfectant solution From the Reader Control dialog change the Volume to 1000 ul Run one prime cycle to flush the disinfectant out of the fluid lines Empty the beaker containing the outlet tubes Put the tubes back in Important If sodium hypochlorite bleach was used perform Rinse the Fluid Lines on the next page Otherwise or after performing the Rinse procedure repeat steps 1 13 for SYRINGE 2 Dispenser 2 BioTek Instruments Inc Routine Procedure for Models With Injectors 159 Rinse the Fluid Lines Perform this procedure only if decontamination was performed using sodium hypochlorite 1 Place a beaker containing at least 30 ml of deionized water on the dispense module Place the SYRINGE 1 or 2 inlet tube in the beaker If you have not already done so place the outlet tubes in an empty beaker From the Reader Control dialog select Dispenser 1 or 2 set the Volume to 5000 ul and keep the default dispense Rate Run five prime cycles for a total of 25000 ul Pause for 10 minutes and then run one prime cycle with 5000 pl This delay will allow any residual sodium hypochlorite to diffuse into the solution and be flushed out with the next prime Empty the beaker containing the outlet tubes Wipe all surfaces with deionized water
15. Warning risk of crushing or pinching Attention risque d crasement et pincement Warnen Gefahr des Zerquetschens und Klemmen Precauci n riesgo del machacamiento y sejecion Attenzione rischio di schiacciare ed intrappolarsi Warning hot surface Attention surface chaude Vorsicht heie Oberfl che Precauci n superficie caliente Attenzione superficie calda Separate collection for electrical and electronic equipment Les quipements lectriques et lectroniques font l objet d une collecte s lective Getrennte Sammlung von Elektro und Elektronikger ten Recogida selectiva de aparatos el ctricos y electr nicos Raccolta separata delle apparecchiature elettriche ed elettroniche Consult instructions for use Consulter la notice d emploi Gebrauchsanweisung beachten Consultar las instrucciones de uso Consultare le istruzioni per uso H amp P BP In vitro diagnostic medical device Dispositif m dical de diagnostic in vitro Medizinisches In Vitro Diagnostikum Dispositivo m dico de diagn stico in vitro Dispositivo medico diagnostico in vitro Z J Synergy HT Operator s Manual xx Preface BioTek Instruments Chapter 1 Introduction This chapter introduces the Synergy HT describes its key features and lists its package contents Page 6 contains information on contacting BioTek Instruments Inc for product support and service Synergy HT Multi Mode Microplate REadef cccc
16. viii Preface Revision History Rev Date Changes 04 2002 First issue B 08 2002 Added Time Resolved Mode 08 2003 Added Dual Fluid Dispense Feature Ch 1 Introduction Updated specifications accessories and technical support Ch 2 Instrument Description Updated component descriptions and added drawings Ch 3 Installation Revised Dispenser Module setup instructions and KC4 launch procedure Revised unpacking repackaging instructions Added new Chapter 4 Getting Started With KC4 Ch 5 Performance Verification Qualification Tests formerly Chapter 4 Revised test procedures Reformatted Appx A Decontamination and Appendix B Computer Control Updated Appx C Error Codes Renamed Appx D Microplate Location Dimensions to Instrument Dimensions D 12 2003 Preface Updated safety symbols and text p ix and x Updated Intended Use Statement p xi Revised Warranty p xii Chapter 1 Modified Introduction p 1 3 Updated list of optional accessories p 1 4 Revised absorbance reading speed information p 1 5 Clarified fluorescence specifications p 1 8 and 1 9 Added specifications for injector model p 1 10 Chapter 2 Clarified description of external and internal components of the injector model and updated drawings Moved procedure for replacing the lamp assembly to Ch 6 Chapter 5 Added note regarding the availability of the Installation Operational Performa
17. 0601 0606 0610 0660 0611 0616 Description and Probable Causes Filter gain out of range Fail if PMT saturation is not met within 3 ms This error indicates that during fluorescence autocalibration the gain was increased to an unsafe level and the PMT analog PCB did not reach saturation Probable Causes e The autocalibration jig is incorrectly placed in the carrier The mirrors are not facing in the correct direction e The EM autocalibration jig is not connected to the PMT analog PCB Time Resolved function reference channel gain out of range for the selected wavelength Fail if reference signal gt 40000 and gain 1 This error indicates that the absorbance reference channel gain for a specific wavelength during Time resolved readings is out of the range necessary to ensure the lambda performance to specifications The second to last number is the lambda table position number Probable Causes e The absorbance reference channel PCB is defective e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is lined up allowing white light to pass e Monochromator is defective e Flash lamp alignment or Flash lamp power supply is defective e Lamp is too bright Time Resolved function reference channel gain out of range for the selected wavelength Fail if Reference signal gt 40000 and Gain 1 Note The order of the last two digits is lt Filter g
18. Click the Absorbance tab to specify and calibrate 6 wave lengths to be made available as default selections within a protocol s Reading Cancel Help Parameters dialog To change the settings and download them to the instrument Excitation Emission Center Bandwidth Center Bandwidth Send Filters i 1 Enter wavelength values in the Center fields or use the drop down boxes to select PLUG or HOLE 2 For each Center wavelength value enter its accompanying Bandwidth The Bandwidth is printed on the side of each filter 3 When finished click Send Filters to download the information to the reader Get Filters uploads information from the reader 4 Click OK to save the settings and close this dialog The settings become available for selection in the protocol s Reading Parameters dialog BioTek Instruments Inc KC4 Software 53 Creating Protocols In KC4 a protocol contains instructions for controlling the reader and optionally instructions for analyzing the data retrieved from the reader At a minimum a protocol must specify the Reading parameters for the assay you wish to run Reading parameters fo Bee even Tecultg b gt Betore first reauing Figure 29 KC4 s Reading parameters dialog software version 3 4 The instructions on the following pages briefly describe how to create protocols in KC4 Be sure to refer to KC4 s Help syst
19. Do not slide the two forks on the housing s right side under the fixed foam housing Replace the groundwire and its thumbscrew p 143 Reconnect the heater and thermistor wires p 143 Be sure to connect wires of the same color Attach the two internal dispense tubes to the tubing ports taking care to align the correct port with the correct injector head p 138 Slide the two internal dispense tubes into the cable clamp and close the clamp p 138 Review the steps you just performed to make sure the components have been properly reassembled Slide the shroud onto the instrument p 136 Replace the four Phillips head screws to securely attach the shroud to the base p 135 Performance Check After reassembling the instrument perform the following to verify that the instrument is functioning properly Plug the instrument in and turn it on allow its run time system test to complete Run a System Test through Gen5 or KC4 Run any required OQ PQ tests Synergy HT Operator s Manual 152 Chapter 5 Preventive Maintenance BioTek Instruments Inc Appendix A Decontamination This appendix contains procedures for decontaminating all models of the Synergy HT RUEDOSCr ai m na ce ete adie a DEAT ANa 154 Required MaterialS s ssusussssssssnenrnrsnenessesenssnunnnnnnenresessane 155 Procedure for Models without INjectOrs cccccceeseeeeeeeeeeeeaneeaas 156 Routine Procedure for Mo
20. If the four corner wells are within the accuracy range the reader is in alignment Synergy HT Operator s Manual 86 Chapter 4 Instrument Qualification Liquid Test 3 This procedure verifies operation of the Synergy HT at 340 nm and is provided for sites requiring proof of linearity at wavelengths lower than those attainable with the Absorbance Test Plate This test is optional because the reader has good front end linearity throughout its wavelength range Required Materials New 96 well flat bottom microplate Corning Costar 3590 is recommended Calibrated hand pipette s Beakers and graduated cylinder Precision balance with readability to 0 01 g Buffer Solution below Read a sample of the buffer solution at 340 nm This solution should have an optical density of approximately 0 700 to 1 000 This value is not critical but it should be within this range If low adjust up by adding B NADH powder until the solution is at least at the lower end of this range Do not adjust if slightly high Buffer Solution Sigma PBS Solution Deionized water Phosphate Buffered Saline PBS pH 7 2 7 6 Sigma tablets P4417 or equivalent B NADH Powder 6 Nicotinamide Adenine Dinucleotide Reduced Form Sigma bulk catalog number N 8129 or preweighed 10 mg vials Sigma number N6785 10VL or BioTek PN 98233 Note Manufacturer part numbers are subject to change over time Store the B NADH Powder
21. 152D Zone 1 3 and 4 152E Zone 2 3 and 4 152F Zone 1 2 3 and 4 Probable Causes e Ato D is defective for that zone e Motor power supply PCB is defective Kinetic interval not correct for selected options This error indicates one of the following scenarios e Kinetic interval in the current assay is too short e Kinetic interval for a fluorescence plate read is impossible for the given parameters e Kinetic interval for an absorbance read the total time or kinetic interval 0 e Kinetic interval for a fluorescence kinetic interval is too big gt 99999 to transmit Probable Causes e User programming error e Increase or decrease the kinetic interval Too many kinetic intervals This error code indicates old basecode Recommend upgrading This error indicates that the combination of assay parameters results in more kinetic reads than supported by the software e Change the assay parameters to reduce the number of total kinetic reads Note The Synergy HT supports kinetics through computer control only BioTek Instruments Inc Code 1900 1C00 1C01 1F01 1F02 2100 2200 2201 Error Codes 191 Description and Probable Causes Memory allocation failed This error indicates that the process failed while saving or moving data If this occurs try turning the instrument off waiting for 30 seconds and then turning the instrument back on Probable Causes e The memory is corrupt Replace the process
22. 2532 See self test Voltage Reference Max This tests the voltage across a sense resistor in series of the 6 00 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 2908 8 00V Flash Power Supply reference voltage is out of range 3251 3417 TR only See self test Voltage Reference HTTR This tests the voltage across a sense resistor in series of the 8 00 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 2911 2918 VRef instability This error indicates that one of the channels voltage reference feeding the A D converter is unstable i e A D max A D min gt 8 after 8 attempts See the table below for more information for each channel Note If multiple channels are intermittently failing a 291x x channels error it is possible that the A D converter is unstable Error Displayed See Error for More Information 2911 2901 2912 2902 2913 2903 2914 2904 2915 2905 2916 2906 2917 2907 2918 2908 Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB Synergy HT Operator s Manual 196 Appendix C Error Codes O
23. Chapter 4 Instrument Qualification Dispense Module Tests This section applies to Synergy HT models with injectors only BioTek Instruments Inc has developed a set of tests that you can perform to ensure that the dispense module performs to specification initially and over time We recommend that you perform these tests before first use e g during the Initial OQ and then every three months The Accuracy Test is a measure of the mean volume per well for multiple dispenses The actual weight of the dispensed fluid is compared to the expected weight and must be within a certain percentage to pass Pass Fail criteria depends on the per well volume dispensed 2 0 for 80 uL 5 0 for 20 uL and 20 0 for 5 uL It is assumed that one gram is equal to one milliliter The test uses a single green dye test solution and one 96 well microplate per injector to test the three different volumes The balance is tared with the empty plate and then the 80 uL dispense is performed for columns 1 4 The fluid is weighed and the balance is tared again with the plate on the balance This process is repeated for the 20 uL and 5 uL dispenses It is assumed that the solutions used are at room temperature A precision balance 3 place is used to weigh the plate The Precision Test is a measure of the variation among volumes dispensed to multiple wells For each volume dispensed 80 uL 20 uL and 5 uL to four columns the CV coefficient of variatio
24. Discard the used gloves and cloths using a Biohazard trash bag and an approved Biohazard container Clean the Internal Tubing and Injector Heads Turn to Chapter 5 Preventive Maintenance and perform the following procedures to access remove and clean the internal tubing and injector heads Required Materials Removing the Reader s Shroud Removing the Internal Tubes and Injector Heads Cleaning the Internal Tubes and Injector Heads When finished replace the internal components and the reader s shroud Synergy HT Operator s Manual 160 Appendix A Decontamination Clean the Tip Priming Trough and Priming Plate 1 Remove the tip priming trough from the left rear of the instrument s microplate carrier see below Wash the tip priming trough and priming plate in hot soapy water Use a small brush or cloth to clean the corners of the trough and plate To decontaminate soak the trough and plate in a container of 0 5 sodium hypochlorite or 70 isopropyl alcohol for 20 to 30 minutes If decontaminating in bleach solution remove the trough and plate and thoroughly rinse with DI water If decontaminating with alcohol remove the trough and plate and let them air dry Discard the used gloves and cloths using a Biohazard trash bag and an approved Biohazard container Figure 39 Tip priming trough and priming plate BioTek Instruments Inc Alternate Procedure for Models with Injectors 161 Alternate
25. Min 36 iis Range Thermistor PASS Zone 3 37 Min 36 4 37 Range Thermistor PASS Zone 4 36 Min 36 z SUs Range Thermistor PASS AUTOCAL ANALYSIS PROBE TOP Upper Left Corner x Lower Left Corner x Lower Right Corner x Upper Right Corner x Delta 1 9732 9720 Delta 2 1056 1044 Delta 3 356 360 Delta 4 5888 5896 PROBE BOTTOM Upper Left Corner x Lower Left Corner x Lower Right Corner x Upper Right Corner x Delta 1 9724 9716 Delta 2 1040 1040 Delta 3 1852 1844 Delta 4 7380 7376 PROBE ABSORB Upper Left Corner x Lower Left Corner x Lower Right Corner x Upper Right Corner x Delta 1 11244 11232 Delta 2 2552 2544 8 Delta 3 1852 1856 4 Delta 4 7376 7376 0 Probe Height 32 48 Middle Sensor 11976 Tested 11972 Delta 4 Back Sensor 11588 Tested 11580 Delta 8 Figure 30 2 Sample output for the System Test Sheet 2 of 3 The format varies depending on the software used BioTek Instruments Inc System Test 69 SYSTEM TEST PASS 0000 Reviewed Approved By For Technical Support In the U S In Europe BioTek Instruments Inc BioTek Instruments GmbH Tel 800 242 4685 Tel 49 0 7136 9680 Fax 802 655 3399 Fax 49 0 7136 968 111 All Others Tel 802 655 4040 Fax 802 655 3399 email TAC biotek com Product support center www biotek com service Figure 30 3 Sample output for the System Test Sh
26. Part numbers are subject to change over time Please contact BioTek Customer Care with any questions e Microplate reader with Excitation and Emission filter wheels installed e Operator s Manual PN 7091000 e Documents including but not limited to Warranty Statement Certificate of Compliance and Calibration Unpacking Packing Instructions e Power supply PN 76061 models with injectors or PN 76053 all other models e Power cord set specific to installation environment gt PN 75010 Schuko Europe gt PN 75012 UK gt PN 75011 USA International gt PN 75013 Australia New Zealand e RS 232 serial cable PN 75034 e USB cable PN 75108 with Virtual COM Driver Software PN 7090204 e Wrench PN 48576 e Fluorescence lamp assembly PN 7080501 Note The part number for a replacement lamp is 7080500 e Filter plugs 2 PN 7082073 also referred to as dummy filters e Plastic storage bag and Velcro strips e Time Resolved Fluorescence cartridge assembly PN 7090523 T models only e Models with injectors SIAFRTD SIAFRTD CUSTOM an external dispense module PN 7090568 with the following accessories gt Outlet tubes 2 plus 2 spare from dispense module to instrument PN 7082120 Inlet tubes 2 from supply bottles to syringe drives PN 7082121 250 uL syringes 2 PN 7083000 Syringe thumbscrews 2 PN 19511 Priming plate PN 7132158 and injector tip priming trough PN 7132169
27. Read already in progress Motor not available This error indicates that a motor is not available but it does not identify which motor was requested Real time clock not available Display device not available TO device not available Timer 0 in addition to being used by the motors is also used in microsecond timing for time resolved measurements Failed code checksum test on power up DR steps alloc free error lt assay number gt This error indicates old basecode recommend upgrading 24V power dropped below safe level Data flash write timed out Data flash read back did not match write Code flash write timed out Memory allocation heap corrupted Absorbance A D converter did not see the Ready signal This error indicates old basecode recommend upgrading Fluorescence A D Converter did not see the Ready signal This error indicates old basecode recommend upgrading Probable Causes e PMT detector PCB AA02 e Analog PCB AA01 e Motor Power PCB AA01 or AA02 Synergy HT Operator s Manual 168 Appendix C Error Codes Non Fatal Errors Non fatal errors indicate non fatal conditions that require attention The last digit of some error codes gives specific information related to the type of error For example in the Home Sensor Initial Find Errors table that follows the last digit indicates which sensor was in error 2 Note The errors presented in the following tables are common to multiple reader ins
28. column to toggle Yes No e Note You need to select only those wavelengths most appropriate for your use of the reader e If N A appears for a wavelength you wish to test update KC4 s Wavelengths table to include it System Readers Filters Wavelengths Enter or select the Operator and enter any Comments Click Run Test to begin e A sample test report is shown on the next two pages e KC4stores the results in a database and they can be retrieved and printed at any time We recommend you print and save the reports to document that the test was performed Performing Peak Absorbance Tests Using Spectral Scan Reads To create and run a KC4 spectral scan protocol to perform an independent Peak Absorbance Test Note Create one protocol per Peak Wavelength value to be tested 1 on Die SON ates O Open the Reading Parameters dialog and select the Spectrum Reading Type and the Absorbance Detection Method Set the Start and Stop wavelength values to define the test range specified in the Test Plate s Peak Wavelength certificate e g perform the scan from 580 to 590 nm Set the Step increment value to 1 nm Set the Plate Type to 96 WELL PLATE and read well C6 Save the protocol Perform a read of the Absorbance Test Plate using this protocol Locate the peak wavelength on the graph the wavelength value with the highest OD Verify that it is within the allowed tolerance see Results amp Troubleshootin
29. detected is outside of acceptable limits at 0 gain when blocking the light Probable Causes If the value is larger than 2450 e Too much light has saturated the PMT Turn the unit off and wait 24 hours e A faulty PMT analog PCB or faulty internal grounding may cause internal electronic noise or the motor power supply PCB is defective or both e PMT or PMT base is defective e There may be an ambient light leak Ensure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and properly fastened If the value is less than 700 e PMT base cable lost ground connection Rebuild connector e PMT or PMT base defective or too quiet e PMT analog PCB and or Motor power supply PCB are defective Absorbance reference channel failed offset range See noise Max lt 20000 and noise Min gt 10 on the system test This error indicates that during the system test the background electronic signal that was detected is outside of acceptable limits at maximum gain when blocking the light Probable Causes If noise Max is gt 20000 e The photodetector is too noisy and is defective e Absorbance channel analog PCB is defective e A faulty analog PCB or faulty internal grounding may cause internal electronic noise e There may be an ambient light le
30. e Manage your equipment inventory e Access documentation on your products e Download user manuals and software e Check the status of your instrument s service e Track your order BioTek Instruments Inc 1 Unpack and Inspect the Reader 9 1 Unpack and Inspect the Reader Important Save all packaging materials If you need to ship the reader to BioTek for repair or replacement you must use the original materials Using other forms of commercially available packaging or failing to follow the repackaging instructions may void your warranty During the unpacking process inspect the packaging reader and accessories for shipping damage If the reader is damaged notify the carrier and your BioTek representative Keep the shipping boxes and the packaging materials for the carrier s inspection BioTek will arrange for repair or replacement of your reader immediately before the shipping related claim is settled Perform these steps to unpack and inspect the reader and accessories 1 Open the outer shipping box Remove the foam blocks to access the inner shipping box Corner shipping block Inner shipping box Outer shipping box Figure 1 A Unpacking the reader s outer shipping box Synergy HT Operator s Manual 10 Chapter 2 Installation 2 Carefully open the inner shipping box Remove the accessories box and set it aside Remove the vertical supports 3 TheSynergy HT is attached to a wooden
31. the dispense module according to the instructions provided in Appendix A BioTek Instruments Inc 10 11 12 13 14 Repackaging and Shipping Instructions 33 If you will also be shipping the dispense module perform these steps now If using Gen5 If using KC4 a With the reader on start Gen5 a With the reader on start KC4 and and select System Reader select System Readers Control Synergy Com lt gt b Click the Dispenser tab Ensure b Click the Configuration button that Dispenser is set to 1 and select the Dispenser 1 tab c Click the Maintenance button c Click the Move Syringe to maintenance position button d The Syringe 1 bracket will lower Remove the thumbscrew from underneath the bracket Carefully unscrew the top of the syringe from the syringe valve Lift out the syringe and store it in its original box e Set the Dispenser number to 2 Repeat steps c and d for Syringe 2 f Fully detach the dispense module from the reader Replace the two nylon screws into the Syringe 1 and 2 tubing ports on the rear of the reader The screws should be stored in the plastic bag attached to the back of the module Set the module aside for the moment If you have not already done so retract the microplate carrier and then turn off and unplug the reader Remove the lamp assembly and pack it in bubble wrap see p 14 Replace the microplate carrier shipping screw seep 13 Tip the rea
32. the wavelength of the maximum absorbance is compared with the peak wavelength value entered in the software which comes from the Peak Wavelength Certificate supplied with the Test Plate The accuracy of the wavelength should be 3 nm 2 nm instrument 1 nm filter allowance If the reader fails this test review the following possible problems and solutions gt Verify that the Test Plate actually has a filter in location C6 Test Plates with the part number 9000547 do not have this filter gt Check the C6 filter to make sure it is clean If needed clean it with lens paper Important Do not remove the filter from the Test Plate and do not use alcohol or other cleaning agents gt Make sure the information entered into Gen5 or KC4 matches the information on the Test Plate s Peak Wavelength Certificate gt Make sure the Test Plate is within its calibration certification period The calibration sticker is affixed directly to the plate If it is out of date contact BioTek to schedule a recertification gt Check the microplate carrier to ensure it is clear of debris Alignment This portion of the test measures the alignment of the microplate carrier with the optical path A reading greater than 0 015 OD represents an out of alignment condition Wells A01 A12 H01 and H12 are the only valid alignment holes for the reader on the PN 7260522 Test Plate If the reader fails this test review the following possible pro
33. then increase the gain to 1250V and test for PMT signal gt 1 If PMT signal is still lt 1 the PMT is not connected or is defective Probable Causes e The PMT is not connected e The PMT analog PCB PMT base or the PMT is defective PMT test signal too high at the 750 volt bias See 750V measurement on self test Measured value gt 3000 to fail This error indicates that the PMT dark current is higher than the specified limit Probable Causes e The PMT has been saturated due to ambient light leakage e The PMT has been damaged e The PMT base is defective or the coaxial cable from the base has lost the ground connection for the shield Synergy HT Operator s Manual 192 Appendix C Error Codes Code 2203 2300 2400 2401 Description and Probable Causes PMT test ratio test failed This error code indicates old basecode Recommend upgrading The 750 500 volt ratio is less than 2 This error indicates that the PMT dark current is higher than specified limit Probable Causes e The PMT has been saturated either due to ambient light leakage e The PMT has been damaged e PMT base is defective or the coaxial cable from the base has lost the ground connection for the shield Lamp control failure This error code indicates old basecode Recommend upgrading This error indicates a failure related to the lamp The lamp assembly is accessed by opening the hinged door on the front of the instrument The lamp may be out C
34. 45 13 4 Install the Fluorescence Lamp Assembly ceeeeeeaes 14 5 Select an Appropriate Location cc ccc ee eee eee eee eee ees 15 6 Connect the Power Supply cece eee eee eee eee teen eee teed 16 7 Unpack and Inspect the Dispense Module eeeee es 17 8 Install the Dispense Module ccceceeee eee eee teeta teen eae 20 9 Connect the Host Computer cece cece eee eee eee teeta 24 10 Install the Software on the Host Computer 008 24 11 Turn on the Reader soc sccvnscn ce esece see heise odhdveec es peepee eens 25 12 Establish Communication ce ceee cece cence eee eee teen ees 26 T3 R n a Systemi TOSE a saccagiecatveacesd pads ea e in e aa 28 14 Test the Injector System cc ce cece cece eee eee teen eee ees 29 Operational Performance Quallification cccccceeeeeeeee renee ennes 31 Repackaging and Shipping Instructions cee eeeeee eee eee eee 32 8 Chapter 2 Installation Product Registration If you have not already done so please register your product s with BioTek to ensure that you receive important information and updates about the product s you have purchased Register online through BioTek s Customer Resource Center at www biotek com or by contacting BioTek Customer Care Once registered you can log into the Customer Resource Center and e Register your product warranty
35. Controller PCB e Carrier front support screws are not adjusted or are worn causing the carrier to no longer be level The support pin is no longer inserting properly into the roller bearings e X axis PCB is not adequately adjusted to the right and will not allow the flag to enter the opto sensor enough to trip the sensor Loosen the two screws and slide the PCB to the right and retighten Run the carrier autocal for both absorbance and fluorescence e Carrier is not able to move into read chamber An object may be obstructing the carrier s path e See Field Change Notice L0030 BioTek Instruments Inc Code Error Codes 169 Description and Probable Causes 0201 Y axis motor did not find the home opto sensor transition This error indicates that a motor was not able to move to its home position as registered by feedback from an optical sensor Note The mid or XY sensor may cause an 0201 error See 2A01 for mid sensor see 2400 for XY sensor Probable Causes Y axis rails are where the bearings are dirty and worn and causing too much friction Defective or broken optical sensor If the Synergy is run with the wrong 24 Volt supply 48W instead of 100W a user will experience Y axis errors error code 0201 when reading a plate using incubation This is because the supply is not large enough to handle running the incubator and motors simultaneously Defective Motor Controller PCB or cable Carrier not able to mov
36. General Edited and reformatted text according to new template Added photographs for clarification as needed Chapter 1 Updated Optional Accessories list Chapter 2 Updated text and graphics to describe illustrate Removal of the front injector Redesign of the priming plate to limit splatter Improved system design to reduce need for periodic maintenance Elimination of the right front tip priming trough and redesign of the left rear trough Improved bottle holder setup Chapter 3 Updated diagram showing removal of Dispense Module from inner shipping box Updated procedure for setting up Dispense Module on the Injector Model Chapter 4 Removed instructions for setting injector position Chapter 6 Added preventive maintenance procedure for periodic cleaning of the top bottom fluorescence optical probes and the absorbance read channel optical path Appendix A Updated drawing of tip priming trough and priming plate 8 2005 Updated Warranty information Moved Specifications from Chapter 1 to Appendix D Corrected operating temperature 18 40 C and injector accuracy 1 ul at 5 50 yl to match published specifications Removed Chapter 2 Instrument Description and distributed the information and photos among the remaining chapters Reorganized the flow of the Installation chapter to better represent actual practice Added test to verify the injector system setup Changed the former Getting
37. KOG ieee vies va taea Eense davacitieaad evade iia vain aed neva 105 Test Procedure Gem srira e vias oe eee eee Sarees le ice eee deer 106 WTeSt Procedure KECA hanenn ia eaaa tea a a aE OANE AEEA EO EANA 108 RESUItS ANALYSIS ids ci a a a E AE AEREA Da eE EAE 109 Creating Test Protocols Using Gen5 ssssssssssrrrrsserrrrnsenrrrrnnenrnn 110 Creating Test Protocols USING KC4 ssssesssrrsserrrrrssurrnrnnenrrrnnnenren 115 Dispense Accuracy amp Precision Test Worksheet cccceceeeeeeeeeeeeeeaeees 121 Chapter 5 Preventive Maintenance ssssssusssnnnnnnunnnnnnnnnnnnnnnnnnnnnn 123 Recommended Maintenance Schedule cceeeeeeeeeee ee eeeeeeeeeeneees 124 OVEFVIEW EP ce dd ee A cestode ae E A TT 124 Dispense MOdUle t a A a aE AAE AEEA ENAERE LAD 124 Scheduler srirsse iii atria ETARE E E TET e E aA 126 Warnings amp PreCaUtiOns ccceccee cece een im aAa a eens 126 Cleaning Exposed Surfaces cccceeeee eee eee e ee teen cena seen eee ea neta ees 127 Inspecting Cleaning Excitation and Emission Filters eeeeees 128 Flushing Purging the Fluid Path ccceeeeeeee eee e eee eee teense eae eaees 129 Running a Dispense Protocol Optional cccceceeeee sees eeeeeeeeeeeeaeenees 130 Emptying Cleaning the Tip Priming Trough ccseceeeeeeeeeeeeeeeeeees 132 Cleaning the Priming Plates ici icc ccovedittv vein dita nonee dena acno ities eet 132 Cleaning the Internal Components
38. OD Mean x 0 010 0 005 For each well compare the standard deviation calculated in step 6 with the allowed deviation calculated in step 7 The standard deviation should be less than the allowed deviation Example Five readings in well A1 of 0 802 0 802 0 799 0 798 and 0 801 will result in a mean of 0 8004 and a standard deviation of 0 0018 The mean multiplied by 1 0 0 8004 0 010 equals 0 008 and when added to the 0 005 0 008 0 005 equals 0 013 which is the allowed deviation for well A1 Since the standard deviation for well A1 is less than 0 013 the well meets the test criteria e Repeatability Specification 1 0 0 005 OD from 0 000 to 2 000 OD 3 0 0 005 OD from 2 000 OD to 3 000 OD Synergy HT Operator s Manual 88 Chapter 4 Instrument Qualification 10 Data Reduction for Linearity For each of the three Test Solutions calculate the mean absorbance for the wells containing that solution mean of wells A1 to H2 A3 to H4 and A5 to H6 Perform a regression analysis on the data to determine if there is adequate linearity For example using Microsoft Excel gt In a spreadsheet enter the three mean values in ascending order and label the column as the Y values Enter 0 50 0 75 and 1 00 and label the column as the X values Select Tools Data Analysis Regression to open the Regression dialog Set Input Y Range to reference the above mentioned Y values and se
39. Procedure for Models with Injectors If you are unable to prime the Synergy HT due to a system failure decontaminate the instrument and the Dispense Module as follows 1 Turn to Chapter 5 Preventive Maintenance and perform the following procedures to remove the shroud and remove clean the internal tubes and injector heads When finished leave the shroud off the reader and proceed to step 2 below e Required Materials e Removing the Reader s Shroud e Removing the Internal Tubes and Injector Heads e Cleaning the Internal Tubes and Injector Heads Prepare an aqueous solution of 0 50 sodium hypochlorite bleach As an alternative 70 isopropyl alcohol may be used if the effects of bleach are a concern Be sure to check the percent NaClO of the bleach you are using this information is printed on the side of the bottle Commercial bleach is typically 10 0 NaClO if this is the case prepare a 1 20 dilution Household bleach is typically 5 0 NaClO if this is the case prepare a 1 10 dilution Slide the microplate carrier out of the instrument Moisten a cloth with the bleach solution or alcohol Do not soak the cloth Use the cloth to wipe e All surfaces of the shroud e All surfaces of the plate carrier e The instrument s rear panel e The exposed surfaces of the dispense module including the syringe valves Continued on the next page Synergy HT Operator s Manual 162 Appendix A Decontamination 6
40. Remove the external tubing and the syringes from the dispense module and soak them in the bleach or alcohol solution Wait for 20 minutes To remove the syringes Pull down the syringe bracket until it stops Remove the metal thumbscrew from underneath the bracket Unscrew the top of the syringe from the bottom of the syringe drive Gently remove the syringe and store it in its original packaging see Chapter 2 Installation 7 Moisten a cloth with DI or distilled water and wipe all surfaces that have been cleaned with the bleach solution or alcohol 8 Rinse all tubing and the syringes with DI water 9 Usea clean dry cloth to dry all wet surfaces on the instrument and the Dispense module 10 Reassemble the instrument and dispense module as necessary 11 Discard the used gloves and cloths using a Biohazard trash bag and an approved Biohazard container BioTek Instruments Inc Appendix B Computer Control The Synergy HT is completely computer controlled it is not equipped with a keypad or its own user interface BioTek s Gen5 and KC4 software packages support all Synergy HT models and are used for both instrument control and data reduction For information on serial protocol commands and or system integration please contact your BioTek sales representative or BioTek Instruments Inc 164 Appendix B Computer Control BioTek Instruments Inc Appendix C Error Codes This section lists
41. Robotic Interface Figure 40 shows the location of the microplate carrier in reference to the exterior surfaces of the Synergy HT and the mounting holes on the bottom The illustration should facilitate system setup with a robotic instrument such as the BioStack Microplate Stacker 216 Appendix E Instrument Dimensions for Robotic Interface Instrument Dimensions for Robotic Interface If you have purchased the BioStack to operate with the Synergy HT special alignment hardware is included in the BioStack alignment kit for correct positioning of the Synergy HT with the BioStack Refer to the Installation chapter in the BioStack Operator s Manual for instructions BioTek Instruments Inc Instrument Dimensions for Robotic Interface 217 m 000 A1 IES MEIS i 5 070 3 490 000 _ 3 400 17 132 a E 000 10 612 500 000 555 15 345 TOP VIEW SIDE VIEW BOTTOM VIEW The two arrows point to special mounting holes for alignment caps for operation with the BioStack Figure 40 Instrument dimensions mounting holes and carrier position Synergy HT Operator s Manual 218 Appendix E Instrument Dimensions for Robotic Interface
42. Scientific 1 L Sodium Borate Mfr 159532 or equivalent OR gt Phosphate Buffered Saline PBS pH 7 2 7 6 e g Sigma tablets Mfr P4417 or equivalent and pH meter or pH indicator strips with pH range 4 to 10 Sodium Fluorescein Powder 1 mg vial BioTek PN 98155 Bottom optics A clean Hellma Quartz 96 well titration plate Mfr 730 009 QG or equivalent such as the 96 well glass bottom Greiner SensoPlate Mfr 655892 Top optics A new clean 96 well solid black microplate such as Corning Costar Mfr 3915 The Hellma Quartz plate described above can also be used Excitation filter 485 20 nm and Emission filter 528 20 nm installed Deionized or distilled water Various beakers graduated cylinders and pipettes pH meter or pH indicator strips with pH range 4 to 10 95 Ethanol for cleaning the bottoms of the plates Aluminum foil Optional but recommended 0 45 micron filter Optional Black polyethylene bag s to temporarily store plate s Gen5 users Synergy HT FI_B prt and Synergy HT FI_T prt protocols described on page 95 KC4 users Synergy HT FI_B prt and Synergy HT FI_T prt protocols described on page 96 BioTek Instruments Inc Fluorescence Tests 91 Test Solutions Filter the solutions to remove particulates that could cause erroneous readings Do not allow dust to settle on the surface of the solution use plate covers or seals when not reading the plate
43. Started with KC4 chapter to a broader Getting Started chapter that includes information on the key instrument components Added new topic for configuring the system for luminescence reads Renamed the Performance Verification Qualification Tests chapter to Instrument Qualification Replaced former Dispense Precision amp Accuracy Tests with new tests that use a single green dye solution and a single microplate Restructured the Preventive Maintenance chapter to better represent actual practice Added a recommended maintenance table for models without injectors Added new photos to help with identification of the various components Updated the Error Codes appendix with recent information Additional minor corrections and improvements throughout Synergy HT Operator s Manual Xx Preface Rev Date Changes 5 06 Redesigned the front cover Removed unnecessary Warranty information a Warranty card ships with every instrument Added warning to shut down instrument and wait for the fluorescence lamp to cool down before replacing it Added the PN for a replacement fluorescence lamp 7080500 For models with injectors Simplified the installation and setup steps for the Dispense Module Added recommendation to set a tip prime volume equal to the per well dispense volume for volumes lt 20 ul Updated Absorbance Plate Test instructions related to Peak Wavelength to support the mo
44. and IVD to Safety Symbols Chapter 1 Introduction In the product introduction section added note that Synergy HT basecode software version 2 24 or greater is required for use with Gen5 Under Package Contents added notice that part numbers are subject to change over time and updated part numbers for the priming plate and tip priming trough Chapter 2 Installation Added section Product Registration Chapter 4 Instrument Qualification Modified sample System Test Report and Absorbance Test Plate Results to reflect more current date and minimum basecode for Synergy HT to work with Gen5 Absorbance Liquid Tests section Liquid Test 3 removed instructions for creating the rarely used Buffer Solution A Fluorescence Liquid Tests section Added option to use Sodium Borate instead of PBS with sodium fluorescein BioTek Instruments Revision History xi Rev Date Changes I Reconfigured the SF test solutions dilutions and pipette maps for efficiency and consistency with other BioTek products Added option to use Methylumbelliferone to test the top optics Dispense Module Tests section Corrected the formula for Accuracy Error Chapter 5 Preventive Maintenance Removed unnecessary Clean Supply Bottle section Modified the Running a Dispense Protocol procedure to include running the experiment and inspecting the plate Added a missing word to the Cleaning the Optical Probes s
45. and Validation Create a report or export template via the Report Builder File Export Builder or Power Export Builder options Select File Save As and give the file an identifying name The instructions below briefly describe how to create a simple Experiment and then read a plate in Gen5 See Gen5 s Help system for complete instructions To create an Experiment and read a plate using Gen5 1 2 3 Select File New Experiment Select the desired protocol and click OK Highlight a plate in the menu tree and select Plate Read The Plate Reading dialog will appear Click READ The door will open and the carrier will extend if it is not already extended Place the plate on the carrier and click OK to begin the read When the read is complete measurement values will appear in Gen5 To view them select the desired data set e g 528 20 645 40 from the Data drop down list Select File Save As and give the file an identifying name Synergy HT Operator s Manual 50 Chapter 3 Getting Started Controlling the Dispense Module This section applies to Synergy HT modds with injectors only Gen5 is used to perform several dispense module specific functions including initializing priming and purging Gen5 also contains certain configuration items that must be set before using the dispense module Read the following sections to become familiar with these functions and configuration items I
46. and describes the possible error codes that may appear on the controlling PC Error CodeS aswaa dua a aed e a tata alann mera een eires eure 166 Fatal EGhOns seuss awry ce dra awa eit easly Resumes ois Simeon waa 167 Non Fatal Err OFS oerrinne ecnruas take EE 168 Home Sensor Initial Find Errors 0100 0307 naasse 168 Home Sensor Verification Errors 0400 0409 4 172 Saturation Errors 0500 0616 ssssssssssssssssrrrrrrrrrrrsssss 174 Absorbance Reader Noise Errors 0700 0A16 access 180 Internal Self Test Errors OD00 2918 cccccceeseeeeeeees 184 Other Errors 2A00 4000 ssessssssrssssesssserrsrsrerrssesesss 196 Status String FORMAL sesmar vedisin P T a a Daya 202 166 Appendix C Error Codes Error Codes When an error occurs during operation with the Synergy HT an error code will appear on the controlling PC In the sample Gen5 and KC4 screens below the error codes are 0201 and 2BOA respectively fz x IN ExecuteReadParameters AN Reader Error 2B04 Reader Error 0201 syringe error Motor position sensor error Error successfully cleared Error codes are represented as four character identifiers The first character will be 0 1 2 3 4 or A For some error codes the fourth character gives specific information related to the type of error e 0 1 2 3 or 4 indicates a non critical error and the instrument will remain able to respond to commands See Non Fatal Errors s
47. and from the Synergy HT s microplate carrier Contact BioTek or visit our website to learn more Synergy HT Operator s Manual 6 Chapter 1 Introduction Product Support amp Service Technical Assistance Center TAC If your instrument s or software fails to function properly if you have questions about how to use or maintain our products or if you need to send an instrument to BioTek for service or repair please contact our Technical Assistance Center BioTek s TAC is open from 8 30 AM to 5 30 PM EST Monday through Friday excluding standard U S holidays You can send a fax or an e mail any time You can also request technical assistance via our website www biotek com Phone 800 242 4685 or Fax 802 654 0638 E Mail tac biotek com 802 655 4740 Please be prepared to provide the following information e Your name and company information along with a daytime phone or fax number and or an e mail address e The product name model and serial number e The software part number available at Help gt About Gen5 or Help gt About KC4 and basecode version available via Gen5 or KC4 for the Synergy HT by selecting System gt Reader Control e For troubleshooting assistance or instruments needing repair the specific steps that produce your problem and any error codes displayed in Gen5 KC4 see also Appendix C Error Codes Returning Instruments for Service Repair If you need to return
48. as follows zone 1 1 2 2 3 4 4 8 see table below for more information Error code Zone Zone Location 1511 Zone 1 Top Right 2 Top Left nie acu 3 Bottom Right 1513 Zone 1 and 2 4 Bottom Left 1514 Zone 3 When facing the front of the unit 1515 Zone 1 and 3 1516 Zone 2 and 3 1517 Zone 1 2 and 3 1518 Zone 4 1519 Zone 1 and 4 151A Zone 2 and 4 151B Zone 1 2 and 4 151C Zone 3 and 4 151D Zone 1 3 and 4 151E Zone 2 3 and 4 151F Zone 1 2 3 and 4 Probable Causes e Thermistor is defective for that zone e Motor power supply PCB is defective Synergy HT Operator s Manual 190 Code 1521 152F 1700 1800 Appendix C Error Codes Description and Probable Causes Incubator A to D failed This error indicates that one or more of incubator zones A to D are defective After turning on the incubator wait at least 10 minutes for it to stabilize Zone encoding is as follows zone 1 1 2 2 3 4 4 8 see table below for more information Error code Zone Zone Location 1521 Zone 1 Top Righi 2 Top Left na acu 3 Bottom Right 1523 Zone 1 and 2 4 Bottom Left 1524 Zone 3 When facing the front of the unit 1525 Zone 1 and 3 1526 Zone 2 and 3 1527 Zone 1 2 and 3 1528 Zone 4 1529 Zone 1 and 4 152A Zone 2 and 4 152B Zone 1 2 and 4 152C Zone 3 and 4
49. be unevenly distributed clean the internal dispense tubes and injector heads as described in Cleaning Internal Components on page 133 Synergy HT Operator s Manual 132 Chapter 5 Preventive Maintenance Emptying Cleaning the Tip Priming Trough Applies only to Synergy HT modds with injectors The tip priming trough is asmall removable priming cup located in the left rear of the microplate carrier used for performing the Tip Prime The trough holds about 1 5 mL of liquid and must be periodically emptied and cleaned by the user To empty clean the tip prime trough 1 Extend the microplate carrier and carefully remove the tip priming trough from its pocket in the left rear of the carrier 2 Wash the trough in hot soapy water Use a small brush to clean in the corners 3 Rinse the trough thoroughly and allow it to dry completely 4 Replace the trough in the microplate carrier When starting aGen5 Experiment that includes dispensing Gen5 will prompt you to empty thetip prime trough Follow the instructions provided When starting a KC4 Read amp Dispense protocol KC4 may prompt you to empty the tip priming trough In this case KC4 will automatically open the System Reader Control Dispenser dialog Empty the trough and then click the Dump Tip Prime Trough s button The Tip Prime Trough value will reset to 1500 ul remaining Cleaning the Priming Plate Applies only to Synergy HT modds with injectors
50. below Each filter and plug is held securely in place with a C clip filter retainer gt Note Each filter has its wavelength and bandpass values printed on its side with an arrow to indicate the proper direction of light through the filter Direction of light Supporting metal bracket Sx EXCITATION gee Filter wheel Thumbscrew ve Direction of light VUTEC EMISSION wae Filter wheel Note the difference in filter orientation between the Excitation and Emission filter wheels Figure 21 Profiles of the Excitation and Emission filter wheels showing proper filter orientation BioTek Instruments Inc Key Components 39 Important The Synergy HT is shipped with a set of Excitation and Emission filters installed and the Synergy s onboard software is pre configured with the filter values and their locations If you change the contents of a filter wheel you must update Gen5 s or KC4 s filter table and then download the information to the reader The Synergy does not automatically detect which filters are installed See page 46 for information on updating Gen5 s filter table See page 52 for information on updating KC4 s filter table Removing the Filter Wheels The filter wheels can be removed if their contents need to be changed It is important to note that The Excitation and Emission filter wheels are not interchangeable and are labeled as follows EX Excitation EM Em
51. bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and properly fastened e Order sorting filter wheel is jammed not aligning the filter wheel to block the light path or the filter is degraded and is not passing enough light energy 0901 0906 The absorbance reference channel Dark current value failed See Dark on the system test See criteria in text below The last number is the lambda table position number This error can indicate one of the following scenarios e The reference channel failed lt 100 during the optic test with the flash on e The reference channel failed lt 100 during a read or blank read not in sweep mode with the flash off e The reference channel failed lt 100 or the Dark value has changed more than 10 from the last self test data during a read or blank read with the flash on Probable Causes e Absorbance analog PCB or reference channel analog PCB is defective e Cable between reference channel and analog PCB is defective or disconnected e Reference channel photodetector is defective or the optic spray is damaged e A faulty analog PCB or faulty internal grounding may cause internal electronic noise e There may be an ambient light leak Make sure the plate carrier door and the front hinged BioTek Instruments Inc Error Codes 183 Code Description and Probable Causes door are properly closed e Electrical noise may be penetrating the measurem
52. chamber and the probe needs to move down but the door is locked the carrier cannot move out of the way of the top probe assembly BioTek Instruments Inc Error Codes 201 Code Description and Probable Causes 4000 PMT Overload at well location This error indicates that at the well specified by the last 3 digits of the error code the PMT was saturated Probable Causes e Chemistry is too bright for the sensitivity selected Well location error code for a 96 well plate 1 2 3 4 5 6 7 8 9 10 11 12 A 4001 4002 4003 4004 4005 4006 4007 4008 4009 400A 400B 400C B 400D 400E 400F 4010 4011 4012 4013 4014 4015 4016 4017 4018 c 4019 401A 401B 401C 401D 401E 401F 4020 40021 40022 4023 4024 D 4025 4026 4027 4028 4029 402A 402B 402C 402D 402E 402F 4030 E 4031 4032 4033 4034 4035 4036 4037 4038 4039 403A 403B 403C F 403D 403E 403F 4040 4041 4042 4043 4044 4045 4046 4047 4048 G 4049 404A 404B 404C 404D 404E 404F 4050 4051 4052 4053 4054 H 4055 4056 4057 4058 4059 405A 405B 405C 405D 405E 405F 4060 Well location error code for 384 well plate Add 40 to the code in the box If code in the box is gt FF add 4 to the code 1 2 3 4 5 6 7 8 9J 1
53. changes proportionally with changes in concentration Proving that the system is linear allows the Sensitivity Test to be run on two points instead of using serial dilutions Important The tests presented in this section require specific microplates solutions and EX EM filters Your laboratory may require a deviation from some of these tests For example you may wish to use a different fluorescing solution and or microplate If deviation from the tests as presented in this section is required the following steps should be taken the first time each test is run e g during the Initial OQ 1 Perform the tests exactly as described on the following pages Rerun the tests using your particular solutions filters microplates etc If results are comparable then the results from these tests will be your baseline for future tests Be sure to document your new test procedure s and save all test results Synergy HT Operator s Manual 90 Chapter 4 Instrument Qualification Required Materials Microplates should be perfectly clean and free of dust or bottom scratches Use new microplates from sealed packages Manufacturer part numbers are subject to change over time Methylumbelliferone can be used as an alternative or supplemental method for performing these tests for the top probe See the instructions starting on page 98 Buffer gt NIST traceable Sodium Borate Reference Standard pH 9 18 e g Fisher
54. e Defective Analog PCB e Defective Flash Lamp and or Flash Lamp Power Supply Inconsistent flashes high probability e Defective motor Power PCB e Defective monochromator low probability e Order sorting filter wheel motor failed due to the bearings The filter wheel does not move to the open hold allowing the monochromator to find home 0306 Probe height Z axis did not find home This error code indicates old basecode Recommend upgrading Probable Causes e The optical flag is not adjusted properly e The Z axis lead screw is no longer glued to the motor shaft e The upper fiber bundle is hitting the top of the shroud due to not being properly tied down 0307 Probe changer motor did not find home This error code indicates old basecode Recommend upgrading Probable Causes e An obstruction is preventing the motor from moving e The motor and or motor drive circuit is defective e The sensor and or sensor circuit defective Home Sensor Verification Errors 0400 0409 These errors occur when the optical sensor is not found again within a range of where it was last found The causes can range from loose transmission components to marginal adjustments The last digit of the error identifies the axis in question Code 0400 Description and Probable Causes Carrier X axis failed positional verify X axis motor failed to get to the same position when moved a known number of steps from the home position and back Probable Causes
55. e The optical trigger flag has moved or is loose e Dirty rail nylon bushings or bearings e Carrier support pin is out of adjustment e Reference Field Change Notice L0030 BioTek Instruments Inc Code 0401 0402 0403 0404 0405 0406 Error Codes 173 Description and Probable Causes Carrier Y axis failed positional verify Y axis motor failed to get to the same position when moved a known number of steps from the home position and back Probable Causes e The optical trigger flag has moved or is loose e Foreign object in the path of the carrier e Dirty rail and bearings EX filter wheel failed positional verify Probable Causes e The filter cartridge was removed and then reseated when the instrument was powered up Running the system test will clear this error e Filter wheel is binding against the motor gear e Motors failed due to the bearings EM Filter Wheel failed positional verify Probable Causes e The filter cartridge was removed and then reseated when the instrument was powered up Running the system test will clear this error e Filter wheel is binding against the motor gear e Motors failed due to the bearings Monochromator Filter Wheel failed positional verify order sorting filters Probable Causes e The optical trigger flag has moved or is loose e Filter wheel is binding against the motor gear e Motors failed due to the bearings Monochromator failed to find the zero order position White
56. interval out of range DISPENSER NOT ATTACHED n27 dispenser module not attached DISPENSER NOT INITIALIZED 329 dispenser has not been initialized DISPENSER NOT PRIMED 2A dispenser has not been primed INVALID DISPENSER OPERATION 32B invalid dispenser operation type requested DISP START LATE as Ohad dispense event missed its scheduled start time NUM SHAKE EVENTS voor number of shake events defined invalid SHAKE START TIME 3 shake start time out of range READ EVENT EXPECTED M32 read event must follow shake event with repeat SHAKE TIME 1337 shake time out of range Synergy HT Operator s Manual 206 Appendix C Error Codes Dispense Read Runtime Definition Error Test Codes Continued SHAKE SPEED 34 shake speed out of range INVALID DEF FORMAT 40 definition format obsolete INVALID _RETURN_TIME AL event start completion time over returnable limit Well Number Codes lt lowest three digits in returned error code gt The row and column of the offending well can be extracted from the well code by applying the following formula examples use 8 x 12 96 well plate I Convert the ASCII hex string to a decimal equivalent Ex 057 indicates 57 hex yielding a well code of 87 decimal 2 Row well code columns in plate rounded up to a whole number Ex 87 12 7 25 indicating row 8 or H oe Column well code row 1 columns in plate Ex 87 8 1 12 column 3 Note If this code
57. mode 96 and 384 well plates 0 000 to 1 000 OD 2 0 0 010 OD Sweep mode 96 and 384 well plates For the above performance the Gain on the Optics Test should be below 10 0 Optics range 200 to 999 nm accuracy 2nm repeatability 0 2 nm bandpass 2 4 nm Detector Photodiodes 2 Synergy HT Operator s Manual 210 Appendix D Specifications Read Modes The Synergy HT supports the following read modes e Normal mode is the slowest of the three available modes After positioning the well over the probe the instrument waits 100 milliseconds before taking the measurement 8 flash data collection Note The 100 ms delay is to allow for more complete fluid settling If the OD is gt 2 000 the reader takes 64 more measurements 64 flash data collection e Rapid mode is faster than Normal mode because the instrument does not wait before taking the measurement and performs an 8 flash data collection even for ODs gt 2 000 e Sweep mode is the fastest of the three available modes The plate carrier sweeps each row past the optics channel without stopping collecting data with a single flash at each well as it goes by Read Timing The following read times are based on a single or dual wavelength measurement Actual reading times can vary depending upon the wavelength read mode and other reading parameters selected In Normal read mode see above the optical density of the solution affects ti
58. module is properly installed and there are no leaks e The IQ procedure should be performed initially before the reader is used for the first time e The successful completion of the IQ procedure verifies that the instrument is installed correctly The Operational Qualification procedure should be performed immediately following the successful IQ see next page BioTek Instruments Inc 1Q 0Q PQ 63 Operational Qualification confirms that the equipment operates according to specification initially and over time The recommended OQ procedure consists of performing the System Test Absorbance Plate Test a series of Liquid Tests and if the external dispense module is used the Dispense Accuracy and Precision Tests Your facility s operating policies may also require that you perform an actual assay prior to accepting the reader for routine use If this is the case you should not use the data obtained from the first assay run on the reader until you have confirmed that the package insert criteria have been met The OQ procedure should be performed initially before first use and then routinely the recommended interval is annually It should also be performed after any major repair or upgrade to the hardware or software Although out of tolerance failures will be detected by the OQ tests results should be compared with those from the routine Performance Qualification tests and previous OQ tests to monitor for trends The
59. monochromator mirror grating is damaged e Absorbance analog PCB intermittently failed e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Missed flashes or an erratic flash lamp e Voltage to the lamp has increased due to failure of the lamp power supply or the motor Synergy HT Operator s Manual 176 Appendix C Error Codes Code 0502 0503 Description and Probable Causes power supply PCB sent the wrong voltage request to the lamp power supply Scenario 4 Reference channel e The monochromator mirror grating is damaged e Absorbance analog PCB intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to lamp has increased due to failure of the lamp power supply or motor power supply PCB sent the wrong voltage request to the lamp power supply Light beam saturated too much light Air measurement channel reading reached 65535 or PMT Relative Fluorescing Units RFU reached FFFF 99999 This error can indicate one of the following scenarios e 1 Prior to a fluorescence read or during a system test the PMT is tested for operation the gain is set at 102 Is the PMT operating properly e 2 Filter 2 in absorbance mode has satur
60. off the instrument and disconnect the fluid line power cable and PC cable Warning The fluorescence lamp assembly is hot when the instrument is powered on If the instrument is on turn it off and allow the lamp to cool down before attempting to replace it BioTek Instruments Inc Cleaning Exposed Surfaces 127 Cleaning Exposed Surfaces Important Turn off and unplug the instrument for all cleaning operations Important Do not immerse the instrument spray it with liquid or usea wet cloth Do not allow the cleaning solution to run into the interior of the instrument If this happens contact BioTek s Service Department Exposed surfaces may be cleaned not decontaminated with a cloth moistened not soaked with water or water and a mild detergent You ll need e Deionized or distilled water e Clean lint free cotton cloths e Mild detergent optional To clean the exposed surfaces 1 Turn off and unplug the instrument N Moisten a clean cotton cloth with water or with water and mild detergent Do not soak the cloth Wipe the plate carrier and all exposed surfaces of the instrument Wipe all exposed surfaces of the dispense module if used 3 4 5 If detergent was used wipe all surfaces with a cloth moistened with water 6 Useaclean dry cloth to dry all wet surfaces 7 Reassemble the instrument as necessary Models with injectors If the Tip Priming Trough overflows wipe the c
61. or a halo effect This may indicate deterioration due to moisture exposure over along period of time e If you have any concerns about the quality of the filters contact your BioT ek representative 4 Clean the filters using lens cleaning tissue moistened with a small amount of isopropyl ethyl or methyl alcohol Ensure that the filters remain in their current locations 5 Replace the filter wheel brackets in their respective positions and replace the thumbscrews Close the hinged door BioTek Instruments Inc Flushing Purging the Fluid Path 129 Flushing Purging the Fluid Path Applies only to Synergy HT modds with injectors At the end of each day that the dispense module is in use flush the fluid path using Gen5 s or KC4 s priming utility Leave the fluid to soak overnight or over a weekend and then purge the fluid before using the instrument again Note This flushing and purging routineis also recommended before disconnecting the outlet tubes from the rear of the reader and before decontamination to remove any assay residue prior to applying isopropyl alcohol or sodium hypochlorite To flush the fluid path 1 Fill two supply bottles with deionized or distilled water Insert the supply inlet tubes into the bottles Place the priming plate on the carrier Gen5 Select System Reader Control Synergy Com lt gt KC4 Select System Reader Control Click the Dispenser tab and select Dispenser 1
62. priming plate from the carrier and empty its contents If the priming plate is empty the prime volume was too low BioTek Instruments Inc Gen5 Software 51 Purge Utility Gen5 provides a special utility to purge fluid from the dispense tubing and syringe by pumping the fluid in reverse back into the supply bottle To purge the dispense module a u oR WON In Gen5 select System Reader Control Synergy Com lt gt and click the Dispenser tab Select the Dispenser number 1 or 2 associated with the supply bottle Enter the desired purge Volume in uL Select a prime Rate in uL second Click Purge to start the process Syringe Maintenance Position Gen5 provides access to special syringe setup functions for maintenance and calibration purposes If a syringe needs to be installed or replaced it must first be moved to its Maintenance Position To do this using Gen5 1 In Gen5d select System Reader Control Synergy Com lt gt and click the Dispenser tab Select the appropriate Dispenser number 1 or 2 associated with the syringe Click Maintenance Thesyringe plunger will move to its furthest from home position The syringe can then be disconnected form the drive bracket and unscrewed from the valve See Install Dispense Module Components in Chapter 2 Installation for information on installing removing the syringes Important Do not change the syringe positions or calibrate t
63. range necessary configuration data missing failed configuration checksum test carrier calibration data missing lt motor gt not homed successfully incubator failure error code lt zone s gt computer control assay definition error interval too short for selected options malloc failed A D calib STBY transition not detected bandpass overlap in filterset invalid parameter value selected PMT test signal out of range lt test type gt lt motor gt failed verify at test sensor BioTek Instruments Inc Non Fatal Errors Continued FLASH MISS ERR XY LIMIT ERR MOTOR IN USE ERR VREF ERR PLATE JAM ERR SYRINGE ERR DISP CNFG DATA ERR DISPREAD ERR PROBE Z CAL ERR SPOOL TIMEOUT ERR ti X BLOCK ERR NETIC OV Pi RRUN ERR ATOD ERR DOOR LOCK ERR ti PMT WELL ERR 25007 26007 28007 29007 2A00 2B00 2c00 2D00 2E00 2F00 30007 31007 32007 3300 40007 Status String Format 203 lt motor gt went by flash location too soon physical limit exceeded for area scan request lt motor gt currently in use voltage reference failed lt test type gt lt motor gt didn t see middle sensor syringe error lt syringe test code gt dispenser data error lt disp cnfg test code gt dispense read error lt disp read param test gt probe Z calibration failure lt cal code gt no ACK received for data handshaking wrong bl
64. shaking the plate at Variable speed for four minutes This will allow any air bubbles in the solution to settle and the meniscus to stabilize Alternatively wait 20 minutes after pipetting the diluted test solution before reading the plate Using Gen5 or KC4 read the microplate five times at 405 nm using the Normal read mode single wavelength no blanking Save the data after each read Normal plate position Without delay rotate the microplate 180 degrees so that well A1 is in the H12 position Read the plate five more times saving the data after each read Turnaround plate position Print out the ten sets of raw data or export them to an Excel spreadsheet Data Reduction Calculate the mean value for each physical well location in columns 1 and 2 for the five plates read in the Normal position and then again for the five plates read in the Turnaround position This will result in 32 mean values Perform a mathematical comparison of the mean values for each microwell in its Normal and Turnaround positions that is compare A1 to H12 A2 to H11 B1 to G12 H2 to A11 To pass this test the differences in the compared mean values must be within the accuracy specification for the instrument Example If the mean value for well A1 in the Normal position is 1 902 with a specified accuracy of 1 0 0 010 OD then the expected range for the mean of the same well in its Turnaround H12 position is 1 8
65. standard deviation of 0 0026 The mean 1 951 multiplied by 1 0 1 951 x 0 010 0 0195 which when added to the 0 005 0 0195 0 005 0 0245 OD which is the allowable deviation Since the standard deviation is less than this value the reader meets the test criteria e Repeatability Specification 1 0 0 005 OD from 0 000 to 2 000 OD 3 0 0 005 OD from 2 000 OD to 3 000 OD Alignment Test 1 Using the plate prepared for the Linearity Test on the previous page conduct a Turnaround test by reading the plate five times with the A1 well in the H12 position Save the data after each read This test results in values for the four corner wells that can be used to determine alignment 2 Calculate the means of the wells A1 and H1 in the Normal plate position data from Linearity Test and in the Turnaround position from Step 1 above 3 Compare the mean reading for well A1 to its mean reading when in the H12 position Next compare the mean values for the H1 well to the same well in the A12 position The difference in the values for any two corresponding wells should be within the accuracy specification for the instrument Example If the mean of well A1 in the normal position is 1 902 where the specified accuracy is 1 0 0 010 OD then the expected range for the mean of the same well in the H12 position is 1 873 to 1 931 OD 1 902 x 1 0 0 019 0 010 0 029 which is added and subtracted from 1 902 for the range
66. successful return to Gen5 s main screen e If the communication attempt is not successful try the following gt Isthereader connected to the power supply and turned on gt Is the communication cable firmly attached to both the reader and the computer gt Did you select the correct Reader Typein step 4 gt Choose a different COM port gt If using the USB cable did you install the driver software See 9 Connect the Host Computer If you remain unable to get Gen5 and the reader to communicate with each other contact BioTek s Technical Assistance Center See page 6 BioTek Instruments Inc 12 Establish Communication 27 Using KC4 Perform these steps to set and test the communication parameters 1 2 3 4 Start KC4 Login if prompted The default System Administrator password is admin Close any warning messages that appear until KC4 s main screen appears Select System Readers Scroll through the list of Available Readers until you see your model Highlight click once on the model name e Synergy HT I SIAFR SIAFR Custom e Synergy HTTR I SIAFRT SIAFRT Custom e Synergy HTTR w Injectors SIAFRTD SIAFRTD Custom Click the Port button and then click Setup Set the Transmission Rate to 9600 and click OK Select the computer s COM port to which the reader is connected and then click OK e If using the USB cable the information can be found via the Windows Control Panel und
67. the injector tip due to evaporation since the last dispense All priming activities are controlled via Gen5 see page 50 or KC4 see page 56 Both types of primes require a fluid reservoir to be present on the microplate carrier e Thepriming plate is about the same size as a standard microplate and is placed on the microplate carrier for a Prime operation to prime the dispense system with fluid e Thetip priming trough is a small removable priming cup located in the left rear of the carrier and is used for performing the Tip Prime before dispensing The trough holds up to 1 5 ml of liquid and must be periodically emptied and cleaned by the user Priming plate Y A Tip priming trough Figure 25 Priming plate and tip priming trough Synergy HT Operator s Manual 46 Chapter 3 Getting Started Gen5 Software BioTek s Gen5 software supports all Synergy HT reader models Use Gen5 to control the reader and the dispense module perform data reduction and analysis on the measurement values print export results and more This section provides brief instructions for creating experiments and reading plates It also explains how to use Gen5 to perform some functions that are specific to the dispense module Viewing Updating the Filter and Wavelengths Tables The Synergy HT ships with a set of Excitation and Emission filters installed and the reader s onboard software is pre configured with the filter v
68. the instructions provided in Gen5 s Getting Started Guide or KC4 s User s Guide to install the software BioTek Instruments Inc 11 Turn on the Reader 25 11 Turn on the Reader Locate the power switch on the front panel and turn on the Synergy HT The reader will automatically initiate a System Test and eject the microplate carrier Figure 16 Carrier eject button top and power ON OFF switch Synergy HT Operator s Manual 26 Chapter 2 Installation 12 Establish Communication Important If you are using the USB cable refer to the instructions that shipped with the USB Virtual COM Driver Software CD to install the necessary drivers and identify the Com Port number Using Gen5 Perform these steps to set and test the communication parameters 1 2 3 4 5 6 Start Gen5 Login if prompted The default System Administrator password is admin When the Welcome to Gen5 screen appears select System Menu Select System Reader Configuration and click Add Set the Reader Type to Synergy Set the Com Port to the computer s COM port to which the reader is connected e If using the USB cable the information can be found via the Windows Control Panel under Ports in the Hardware Device M anager area of System Properties e g USB Serial Port COM 5 Click the Test Comm button Gen5 will attempt to communicate with the reader e If the communication attempt is
69. the lines at the beginning of each day ensures optimal performance of the dispense system See Chapter 5 Preventive Maintenance for more information e For models with injectors When dispensing volumes less than or equal to 20 uL well we recommend specifying a tip prime volume that is equal to the dispense volume For dispense volumes greater than 20 uL well we recommend a tip prime volume of 20 uL BioTek Instruments Inc Chapter 4 Instrument Qualification This chapter contains procedures for qualifying the initial and ongoing performance of the Synergy HT and the external dispense module if used OVERVIEW an aar earra dotveitee tees tendo leben cts s deeeketied Gets 62 TOF OO PO korrier ennan e deeutinnrndeekdsponetetene coolers a aa 62 Recommended Qualification Schedule cccc cece eeeeeee ee eeeeee ees 64 System TeSt esoe ion ETE chunterk ee der I Aedes Ea 65 Absorbance Plate TeSt sites amenna oe i tee eee 70 Absorbance Liquid Tests wisiscicccsveieyd diwervetews canes es veaweiw needa 80 Fluorescence TeEStS scien Sobor scares dee EEE EEEE nates 89 Dispense Module TeSts cccccccccseesseeesseeeeeeeseeeeseetesantesaeneas 102 Dispense Accuracy amp Precision Tests Worksheet cccceeeeeeaes 121 62 Chapter 4 Instrument Qualification Overview Every Synergy HT reader and external dispense module is fully tested at BioTek prior to shipment and they should operate properly upon initial set
70. the tip priming trough in the left rear pocket of the carrier Place the priming plate on the carrier Priming plate Tip priming trough Figure 17 Installing the tip priming trough and priming plate on the microplate carrier 2 Fill thetwo reagent bottles with distilled or deionized water Place the bottles in their holders and place the holders directly in front of the syringes Insert the inlet tubes into the bottles The dispense module s setup should resemble the photo in Figure 18 Make any final adjustments if necessary Synergy HT Operator s Manual 30 Chapter 2 Installation Figure 18 The fully assembled dispense module 3 Gen5 Select System Reader Control Synergy Com lt gt KC4 Select System Reader Control 4 Click the Dispenser tab 5 With Dispenser set to 1 set the Volume to 5000 ul and click Prime The syringe should move down and up repeatedly drawing fluid from the bottle The fluid should pump through the tubing and dispense into the priming plate Examine the fittings no leaks should be detected If leaks are detected tighten all fittings and repeat the prime If leaks are still detected contact BioTek s Technical Assistance Center 6 When the prime finishes set Volume to 2000 wl and click Purge to clear the fluid lines 7 Set Dispenser to 2 and repeat steps 5 and 6 8 When finished remove and empty the priming plate 9 Close the software The in
71. wells C2 F2 and dispense it into wells C3 F3 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from wells C3 F3 and dispense it into wells C4 F4 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from wells C4 F4 and dispense it into wells C5 F5 Mix the wells using the pipette Aspirate 150 uL from wells C5 F5 and discard 1 2 3 4 5 6 7 8 9 10 11 12 A B 100 50 25 12 5 6 25 c nM nM nM nM nM Buu BYT Bur 100 50 25 12 5 6 25 D nM nM nM nM nM EE BEE BUF 100 50 25 12 5 6 25 E nM nM nM nM nM Uh Bue BUF 100 50 25 12 5 6 25 F nM nM nM nM nM BUF BUF BUF G H BioTek Instruments Inc Fluorescence Tests 101 Protocol Reading Parameters Your tests may require modifications to some of the parameters below such as Plate Type or Sensitivity see Troubleshooting on page 93 The Plate Type setting should match the plate you are actually using The following tables contains the recommended settings for the Synergy HT FI_MUB prt protocol GEN5 USERS Protocol Name Synergy HT FI_MUB prt Parameter Setting Detection Method Delay After Plate Movement Measurements Per Data Point Top Probe Vertical Offset KC4 USERS Protocol Name Synergy HT FI_MUB prt Plate Type Costar 96 black opaque 3915 Top Probe Vertical Offset Synergy HT Operator s Manual 102
72. would indicate an ABS measurement channel error A 0501 error would represent an absorbance reference channel If the fluorometric measurement system is in use filter wheels homing a 0500 error could indicate a missing filter In this case a 0501 would indicate a missing filter in position 1 of the wheel which was last homing prior to the error condition being detected BioTek Instruments Inc Error Codes 175 Code Description and Probable Causes 0500 Light beam saturated too much light Air reading reached 65535 or PMT Relative Fluorescing Units RFU reached FFFF 99999 This error can indicate one of the following scenarios e 1 During an absorbance filter calibration or wavelength scan the reference channel is saturated when storing to memory e 2 During a fluorescence dispensing read after dispensing the read times out due to the sample saturating the PMT e 3 During a spectral scan read the reference channel reached saturation at one or more of the selected wavelengths e 4 During fluorescence autocalibration the light level did not reduce to more than half the saturation light level when the carrier was moved away from center Probable Causes Scenario 2 fluorescence dispensing read e Incorrect chemistry was dispensed into the well Scenarios 1 and 3 spectral scan e Monochromator has a defect in the mirror gradients e Absorbance analog PCB intermittently failed e Order sorting filter w
73. you 1 2 3 4 5 6 7 In the protocol open the Plate Layout dialog Set the Type set to Assay Control Click the Browse button associated with the ID field Assign the first three controls as Disp_80 Disp_20 and Disp_5 Click OK Set ID to Disp_80 and highlight wells A1 to H4 Set ID to Disp_20 and highlight wells A5 to H8 Set ID to Disp_5 and highlight wells A9 to H12 Click OK to save the changes and close the dialog Tip After running the experiment view the Statistics for each Delta OD Data Set to view the calculations Add Data Reduction Steps Each Read step is performed using two wavelengths so you ll need to create two data reduction steps to calculate the Delta OD values 1 oe oy A ty In the protocol open the Data Reduction dialog and click the Transformation button Click the Select Multiple Data Sets button and then click the DS2 radio button see below Set the Data In for DS1 to the 80 uL Read step at 405 nm Set the Data In for DS2 to the 80 uL Read step at 750 nm Click OK to return to the Transformation dialog In the New Data Set Name field type an identifying name such as Delta OD 80 ul_Disp 1 see sample screen on the next page Clear Use single formula for all wells In the Current Formula field type DS1 DS2 and then highlight wells A1 to H4 to assign the formula Synergy HT Operator s Manual 114 Chapter 4 Instrument Qualification Transform
74. 0 11 12 13 14 15 16 17 18 19 20 21 22 23 24 A 01 02 03 04 05 06 07 08 09 OA 0B OC OD OE OF 10 11 12 13 14 15 16 17 18 B 19 1A 1B 1C 1D 1E 1F 20 21 22 23 24 25 26 27 28 29 2A 2B 2C 2D 2E 2F 30 C 31 32 33 34 35 36 37 38 39 3A 3B 3C 3D 3E 3F 40 41 42 43 44 45 46 47 48 D 49 4A 4B 4C 4D 4E 4F 50 51 52 53 54 55 56 57 58 59 SA 5B 5C 5D 5E SF 60 E 61 62 63 64 65 66 67 68 69 6A 6B 6C 6D 6E 6F 70 71 72 73 74 75 76 77 78 F 79 7A 7B 7C 7D 7E 7F 80 81 82 83 84 85 86 87 88 89 8A 8B 8C 8D 8E 8F 90 G 91 92 93 94 95 96 97 98 99 9A 9B 9C 9D 9E OF AO A1 A2 A3 A4 A5 AG A7 AB H A9 AA AB AC AD AE AF BO B1 B2 B3 B4 BS B6 B7 BB BO BA BB BC BD BE BF CO C1 C2 C3 C4 C5 C6 C7 C8 cC9 CA CB CC CD CE CF DO D1 D2 D3 D4 D5 D6 D7 D8 J D9 DA DB DC DD DE DF E0 E1 E2 E3 E4 E5 E6 E7 E8 E9 EA EB EC ED EE EF FO K F1 F2 F3 F4 F5 F6 F7 F8 F9 FA FB FC FD FE FF 100 101 102 103 104 105 106 107 108 1 109 10A 10B 10C 10D 10E 10F 110 111 112 113 114 115 116 117 118 119 11A 11B 11C 11D 1
75. 0 failed position verify also known as the first syringe drive Probable Causes The optical trigger flag has moved is loose or has intermittently failed For certain older dispensers opto cable 7330506 may need to be replaced with 7120734 The linear way and bearings are dirty or lack sufficient grease The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft The assembly is not able to move because a foreign object is preventing the movement 0409 Syringe 1 failed position verify also known as the second syringe drive Probable Causes The optical trigger flag has moved is loose or has intermittently failed For certain older dispensers opto cable 7330506 may need to be replaced with 7120734 The linear way and bearings are dirty or lack sufficient grease The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft The assembly is not able to move because a foreign object is preventing the movement Saturation Errors 0500 0616 These errors occur in several places when the measurement channel in use is found to be in an unexpected saturation condition The error decoding is complex and requires some orientation in terms of where the failure occurs The correct identification will depend on listening to the unit function prior to the reported error For example if the monochromator motor were in motion during a self check a 0500 error
76. 10 H 40 6 cm x 38 cm x 25 4 cm Weight 38 Ib 17 kg Environment Operational temperature 18 to 40 C Humidity 10 to 85 relative humidity non condensing Power Consumption 100 VA max 130 VA max with injectors BioTek Instruments Inc Specifications 209 Absorbance Accuracy Linearity Repeatability All qualifications were conducted using 96 384 well flat bottom microplates Measurement Range 0 000 to 4 000 OD All qualifications were conducted with 96 and 384 well flat bottom plates Accuracy 0 000 to 2 000 OD 1 0 0 010 OD Normal and Rapid modes 96 well plates 0 000 to 2 000 OD 2 0 0 010 OD Normal and Rapid modes 384 well plates 2 000 to 2 500 OD 3 0 0 010 OD Normal and Rapid modes 96 and 384 well plates 2 500 to 3 000 OD 3 0 0 010 OD Normal mode 96 well plates 0 000 to 1 000 OD 1 0 0 010 OD Sweep mode 96 and 384 well plates Linearity 0 000 to 2 000 OD 1 0 Normal and Rapid modes 96 well plates 0 000 to 2 000 OD 2 0 Normal and Rapid modes 384 well plates 2 000 to 2 500 OD 3 0 Normal and Rapid modes 96 and 384 well plates 2 500 to 3 000 OD 3 0 Normal mode 96 well plates 0 000 to 1 000 OD 1 0 Sweep mode 96 and 384 well plates Repeatability 0 000 to 2 000 OD 1 0 0 005 OD Normal and Rapid modes 96 and 384 well plates 2 000 to 2 500 OD 3 0 0 005 OD Normal and Rapid modes 96 and 384 well plates 2 500 to 3 000 OD 3 0 0 005 OD Normal
77. 1E 11F 120 M 121 122 123 124 125 126 127 128 129 12A 12B 12C 12D 12E 12F 130 131 132 133 134 135 136 137 138 N 139 13A 13B 13C 13D 13E 13F 140 141 142 143 144 145 146 147 148 149 14A 14B 14C 14D 14E 14F 150 O 151 152 153 154 155 156 157 158 159 15A 15B 15C 15D 15E 15F 160 161 162 163 164 165 166 167 168 P 169 16A 16B 16C 16D 16E 16F 170 171 172 173 174 175 176 177 178 179 17A 17B 17C 17D 17E 17F 180 Synergy HT Operator s Manual 202 Appendix C Error Codes Status String Format Following execution of each command the Synergy sends a status string back to the computer This string consists of 5 successive ASCII characters a four byte string representing a hexadecimal status code and then ETX Fatal errors indicate a hardware failure require recycling of instrument power Fatal Errors TCB NOT AVAIL E EAD NOT AVAIL OT AVAIL ERR HECKSUM ERR OWER ERR FLASH TIMEOUT FLASH ERR FLASH TIMEOUT EAP CORRUPTED TQAUVUNMNAZAD Non Fatal Errors ABORT ERR NO SENSOR ERR NO BEAM ERR ERR RR RR MOTOR VERIFY ERR SATURATION ERR FILTER GAIN ERR NOISE TEST ERR OFFSET TEST ERR DARK RANGE ERR AIR RANGE ERR WAVECAL DATA ER WAVE NOT FOUND FILTER SIGNAL E CNFG D
78. 24 Chapter 5 Preventive Maintenance Recommended Maintenance Schedule Overview A general Preventive Maintenance PM regimen for all Synergy HT models includes periodically cleaning all exposed surfaces and inspecting cleaning the Excitation and Emission filters For models with the external dispense module additional tasks include flushing purging the fluid path and cleaning thetip prime trough priming plate supply bottles internal dispense tubing and injector heads Dispense Module To keep the dispense module and injectors in top condition flush and purge the fluid lines with deionized DI water every day or upon completion of an assay run whichever is more frequent Some reagents may crystallize or harden after use clogging the fluid passageways Flushing the tubing at the end of each day letting the DI water soak them overnight and then purging the lines at the beginning of each day ensures optimal performance of the dispense system Perform a visual inspection of the dispensing accuracy before conducting an assay that requires dispense to verify instrument performance It is important to keep the dispensing lines scrupulously clean at all times Take special care when using molecules active at very low concentrations e g enzymes inhibitors Remove any residual reagent in the dispensing lines using a suitable cleaning solution review the reagent s package insert for specific recommendations A daily cleaning re
79. 6 Appendix C Error Codes Code OF10 OF10 OF60 OF11 OF16 1000 1100 Description and Probable Causes Measurement channel correction outside limits See criteria in text below Non Time Resolved This error indicates that during a spectral scan the flash on value minus flash off is lt 8000 normal mode lt 8000 calibrated resets 8 for sweep mode Probable Causes e Lamp alignment or lamp power supply failure e Defective reference channel analog PCB absorbance channel analog PCB or the cable in between e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path and white light is passing through or the filter is degraded and is not passing enough light energy or is blocking the light e Damaged optic spray e See the next code for more possibilities Time Resolved function Reference channel correction outside limits Fail if Reference signal lt 500 and gain gt 248 Note The order of lt Filter gt lt Channel gt is not consistent with other error codes This error indicates that during a Time Resolve read delta air dark was lt 500 and the gain could not be incremented past 248 Probable Causes e Lamp alignment or lamp power supply failure e Absorbance channel analog PCB e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path and white light is passing through or the filter is degraded and is not passin
80. 73 to 1 931 OD 1 902 x 0 010 0 010 0 029 1 902 0 029 1 873 1 902 0 029 1 931 BioTek Instruments Inc Absorbance Liquid Tests 83 e Accuracy Specification The following accuracy specifications are applied using Normal mode and a 96 well microplate 1 0 0 010 OD from 0 000 to 2 000 OD 3 0 0 010 OD from 2 000 OD to 3 000 OD Liquid Test 2 The recommended method for testing the instrument s alignment repeatability and accuracy is to use the Absorbance Test Plate see page 70 If the Test Plate is not available however Liquid Test 2 can be used for these tests Required Materials e Anew 96 well flat bottom microplate such as Corning Costar 3590 e Ten test tubes numbered consecutively set up in a rack e Calibrated hand pipette Class A volumetric pipette recommended e Stock solution A or B see page 81 e A0 05 solution of deionized water and Tween 20 Prepare the Dilutions Refer to the table below and e Create a percentage dilution series beginning with 100 of the original concentrated stock solution A or B in the first tube 90 of the original solution in the second tube 80 in the third tube all the way to 10 in the tenth tube e Dilute using the 0 05 solution of deionized water and Tween 20 Table 2 Test Tube Dilutions for Liquid Test Tube Number Volume of Original Concentrated Solution mL Volume of 0 05 Tween Solution mL Absorbance expected if origin
81. 80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_5 Disp_5 Disp_5 DS1 D 2 D 1 DS2 D 1 D 2 DS1 Ds2 Drag 9 Click OK to add the transformation to the Data Reduction list 10 Create another Transformation similar to the above with these characteristics DS1 set to the 20 and 5 uL Read step at 630 nm DS2 set to the 20 and 5 uL Read step at 750 nm e New Data Set Name resembling Delta OD 20 and 5 ul_Disp lt gt e Formula DS1 DS2 applied to wells A5 to H12 11 When you re finished the Data Reduction Steps list will show two Delta OD transformations ig Data Reduction Data Reduction Steps Description Data Out G transformation Delta OD 80 ul_Disp 1 transformation Delta OD 20 and 5 ul_Disp 1 Add Step Transformation WellAnalvsis 12 Click OK to close the Data Reduction dialog e K Select File Save As and give the protocol an identifying file name such as Synergy HT Dispenser 1 prt BioTek Instruments Inc Dispense Module Tests 115 Creating the Test Protocols Using KC4 To perform the Dispense Accuracy and Precision tests you ll need to create a total of six Dispense protocols in KC4 three per dispenser If you will be using one of BioTek s absorbance readers including the Synergy HT you ll need to create two additional protocols for reading the plate after the
82. ATA ERR CNFG CHECKSUM E AUTOCAL DATA ER MOTOR NOT HOMED INCUBATOR FAILU SC ASSAY DEF ER KIN INTERVAL ER MALLOC ERR ATOD INIT ERR OVERLAP ERR ERR RE INVALID PARAM ERR PMT ERR SENSOR POS ERR A100 A200 A300 A400 A500 A600 A700 A800 A900 01007 02007 03007 04007 05007 06007 07007 08007 09007 OA00 OD00 OE00 OF00 1000 1100 1200 1300 500 1600 1700 1900 1000 1F00 2100 22007 24007 task control block not available read function already invoked lt device gt not available failed code checksum test on powerup power dropped below safe level data flash write timed out read didn t match write test lt chip gt code flash write timed out memory allocation heap corrupted read function aborted lt motor gt didn t find opto sensor transition lt motor gt didn t find saturation transition lt motor gt failed positional verify A D signal saturated lt filter gt lt channel gt lt filter gt gain out of range reader channel failed noise test reader channel failed offset test read time channel lt filter gt dark out of range read time channel lt filter gt air blank out wavelength calibration data missing wavelength not found in table lt read filter gt channel lt filter gt signal out of
83. Dispense module communication cable PN 75107 Dispense module front cover PN 7082137 Supply bottles 2 30 mL PN 7122609 and holder assemblies 2 PN 7090564 Injector tip cleaning stylus PN 2872304 VV VV VV V WV BioTek Instruments Inc Optional Accessories 5 Optional Accessories 2 Part numbers are subject to change over time Please contact BioTek Customer Care with any questions e 7 filter Absorbance Test Plate PN 7260522 for absorbance measurement testing e Fluorescence Test Plate PN 7092092 for fluorescence measurement testing e Product Qualification IQ OQ PQ package PN 7090521 e PCR Tube Adapter Plates PNs 6002072 and 6002076 e Terasaki Adapter Plate PN 7330531 e BioCell Quartz Vessel PN 7272051 and Adapter Plate PN 7270512 e Additional Fluorescence Filters contact BioTek for part numbers and availability e Absorbance Liquid Test Solutions gt BioTek Wetting Agent Solution PN 7773002 gt BioTek QC Check Solution No 1 PN 7120779 25 ml or PN 7120782 125 ml e BioTek Sodium Fluorescein Powder PN 98155 e Models with injectors Dispense Module Liquid Test Solutions gt BioTek Green Test Dye Solution PN 7773003 gt BioTek Blue Test Dye Solution PN 7773001 gt BioTek QC Yellow Solution PN 7120782 The Synergy HT is compatible with BioTek s BioStack Microplate Stacker The BioStack rapidly and systematically transfers microplates up to 30 per stack to
84. Getting Started Key Components Power Switch Carrier Eject Button Microplate Carrier Microplate carrier Well Al Carrier eject button Power switch Synergy HT Figure 19 Power switch carrier eject button microplate carrier e The power switch is labeled I O indicating on and off respectively An LED on the switch indicates that the power is on e The microplate carrier eject button can be used to move the microplate carrier into or out of the measurement chamber and also to stop the instrument from beeping when it encounters an error e The microplate carrier supports microplates and adapter plates as described in Appendix D Specifications The plate is positioned so that well A1 isin the left rear corner of the carrier A spring clip holds the plate securely in place The microplate loading door helps to ensure a light impermeable measurement chamber When a plate read is initiated the carrier slides into the measurement chamber and then moves in the X and Y axes to align each microwell with the top or bottom fluorescence probe or bottom absorbance probe as specified in the Gen5 procedure or KC4 protocol When the read is complete the plate carrier slides to its full out position 7 For fluorescence and luminescence reading modes the height of the top optical probe can be adjusted Use the Top Probe Vertical Offset option to define how far the top probe shall be offset from t
85. HT Operator s Manual 144 Chapter 5 Preventive Maintenance 3 Lift the ground wire and move it off to the side 4 Locate the two black thumbscrews that hold the incubator housing in place Remove both of them and set them aside BioTek Instruments Inc Cleaning the Internal Components 145 5 Turn the top probe screw counterclockwise to lower the probe hanger all the way to the bottom Rotate the screw not the ring around it Ky Rotate J counterclockwise 4 vin Note When replacing the incubator housing the two forks on its right side should wrap around the holding screws The forks should not slide under the fixed foam housing Synergy HT Operator s Manual 146 Chapter 5 Preventive Maintenance 7 Usea 1 8 Allen wrench to remove the top optical probe s holding screw 8 Gently pull the optical probe up and out of its socket to expose it for cleaning Soak the probe in alcohol for one minute maximum Wipe with lens cleaning tissue and set aside Clean with BioTek Instruments Inc Cleaning the Internal Components 147 9 Usea3 32 Allen wrench to remove the two shoulder screws securing the top probe hanger Remove the screws and set them aside v Shoulder screws a 10 Drop the top probe hanger down and slide to the left to remove it Turn the hanger upside down to clean the absorbance lens see instructions on the next page Do n
86. Multi Mode Microplate Reader synergy HT Operator s Manual ssBiolek Synergy HT Multi Mode Microplate Reader Operator s Guide November 2008 2008 Part Number 7091000 Revision I BioTek Instruments Inc ii Preface Notices BioTek Instruments Inc Highland Park P O Box 998 Winooski Vermont 05404 0998 USA All Rights Reserved 2008 BioTek Instruments Incorporated No part of this publication may be reproduced transcribed or transmitted in any form or by any means electronic or mechanical including photocopying and recording for any purpose other than the purchaser s use without written permission of BioTek Instruments Inc Trademarks BioTek is a registered trademark and Synergy HT Gen5 KC4 and BioStack are trademarks of BioTek Instruments Inc All other trademarks are the property of their respective holders Restrictions and Liabilities Information in this document is subject to change and does not represent a commitment by BioTek Instruments Inc Changes made to the information in this document will be incorporated in new editions of the publication No responsibility is assumed by BioTek for the use or reliability of software or equipment that is not supplied by BioTek or its affiliated dealers BioTek Instruments Contents iii Contents NOUCES Aw eat Ata a ManhAviniaa iia Ate aie ama ii Contact Information seisilesccesdcedvieetedele orreisce
87. Read columns 1 4 at 405 750 nm calculate the Delta OD e Read columns 5 12 at 630 750 nm calculate the Delta OD The detailed procedure is described on the next page To add a step to the procedure click the appropriate button on the left side of the Procedure dialog see Figure 33 on page 110 and define the required parameters The comments suggested for use with the Plate Out In steps are optional but they may be useful for the person running the test When the Plate Out In step is executed Gen5 displays its comment in a message box as demonstrated below F Protocol Plate 1 2 Audit Trail Matrix Statistics Data Delta OD 80 ul Disp 1 5 elt 2 4 5 Weigh the plate 80 ul test RECORD the weight TARE the balance Place the plate back on the carrier Click OK to continue Figure 34 Sample comment associated with a Plate Out In step presented to the operator at the start of the experiment Synergy HT Operator s Manual 112 Chapter 4 Instrument Qualification Gen5 Procedure Steps Dispense Dispenser lt select 1 or 2 depending on the protocol gt Dispense to wells A1 H4 Plate Out In Dispense 80 uL at rate 275 uL sec Plate Out In Suggested comment Weigh the plate 80 ul test RECORD the weight TARE the balance Place the plate back on the carrier Click OK to continue Dispenser lt select 1 or 2 depending on the protoco
88. Set the Volume to 5000 uL Keep the default prime Rate Click Prime to start the process When the process is complete carefully remove the priming plate from the carrier and empty it Repeat the process for Dispenser 2 Leave the water in the system overnight or until the instrument will be used again Purge the fluid from the system see below and then prime with the dispense reagent before running an assay To purge the fluid from the system 1 2 Oy i a ee Place the inlet tubes in empty supply bottles or a beaker Gen5 Select System Reader Control Synergy Com lt gt KC4 Select System Reader Control Click the Dispenser tab and select Dispenser 1 Set the Volume to 2000 UL Click Purge to start the process When the purge is complete repeat the process for Dispenser 2 After purging the system you may wish to run a quick Dispense protocol to visually verify the dispense accuracy See the next page for instructions for creating the protocol Synergy HT Operator s Manual 130 Chapter 5 Preventive Maintenance Running a Dispense Protocol Optional Applies only to Synergy HT modds with injectors After flushing purging the system page 129 and before running an assay that requires dispense take a moment to visually inspect the dispensing accuracy Usea DI H20 Tween solution to check for dispense accuracy following maintenance e g add 1 ml Tween 20 to 1000 ml of deionized water
89. a wet cloth Do not allow the cleaning solution to run into the interior of the instrument If this happens contact the BioTek Service Department Important Turn off and unplug the instrument for all decontamination and cleaning operations Turn off and unplug the instrument Prepare an aqueous solution of 0 50 sodium hypochlorite bleach As an alternative 70 isopropyl alcohol may be used if the effects of bleach are a concern Be sure to check the percent NaClO of the bleach you are using this information is printed on the side of the bottle Commercial bleach is typically 10 0 NaClO if this is the case prepare a 1 20 dilution Household bleach is typically 5 0 NaClO if this is the case prepare a 1 10 dilution eR a Moisten a cloth with the bleach solution or alcohol Do not soak the cloth Manually open the plate carrier door slide out the plate carrier Wipe the plate carrier and all exposed surfaces of the instrument Wait 20 minutes Moisten a cloth with deionized DI or distilled water and wipe all surfaces of the instrument that have been cleaned with the bleach solution or alcohol Use a clean dry cloth to dry all wet surfaces Reassemble the instrument as necessary Discard the used gloves and cloths using a Biohazard trash bag and an approved Biohazard container BioTek Instruments Inc Routine Procedure for Models With Injectors 157 Routine Procedure for Models With Injectors
90. absorbance analog PCB may be floating or disconnected e There may be an ambient light leak Ensure the plate carrier door and the front hinged door are properly closed e Analog PCB failure the photodetector is noisy e A faulty analog PCB or faulty internal grounding may cause internal electronic noise 0710 Absorbance measurement channel noise test at max gain See noise Delta on self test fail if lt 20 This error indicates significant variations in background electronic noise were detected when blocking the light and increasing the gain to maximum Probable Causes e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and properly fastened e The coaxial cable ground between the reference channel and the absorbance analog PCB may be floating or not connected e There may be an ambient light leak Ensure the plate carrier door and the front hinged door are properly closed e Analog PCB failure the photodetector is noisy e Faulty analog PCB or faulty internal grounding may cause internal electronic noise BioTek Instruments Inc Error Codes 181 Code Description and Probable Causes 0800 PMT failed Noise Offset where the initial offset value was not between 700 and 2450 see the offset voltage on the system test This error indicates that during the system test the background electronic signal that was
91. according to the guidelines on its packaging Prepare a PBS solution from the Sigma tablets In a beaker mix 50 mL of the PBS solution with 10 mg of the B NADH powder and mix thoroughly This is the Sigma PBS Solution BioTek Instruments Inc Absorbance Liquid Tests 87 Procedure In this procedure the prepared stock buffer solution B is referred to as the 100 Test Solution 1 Carefully prepare a 75 Test Solution by diluting 15 mL of the 100 Test Solution e If using the Sigma PBS Solution use 5 mL as the diluent Carefully prepare a 50 Test Solution by diluting 10 mL of the 100 Test Solution e If using the Sigma PBS Solution use 10 mL as the diluent Carefully pipette the three solutions into a new 96 well microplate e 150 uL of the 100 Test Solution into all wells of columns 1 and 2 e 150 uL of the 75 Test Solution into all wells of columns 3 and 4 e 150 uL of the 50 Test Solution into all wells of column 5 and 6 Using Gen5 or KC4 read the microplate five times using Normal mode single wavelength at 340 nm no blanking Save the data after each read Print out the five sets of raw data or export them to an Excel spreadsheet Data Reduction for Repeatability For each well calculate the mean and standard deviation of the five readings For each mean calculated in step 6 calculate the allowed deviation using the repeatability specification for a 96 well plate of 1 0 0 005
92. act the second wavelength from the first The name of the resulting data set is Delta OD Select the appropriate Plate Type or set it to 96 WELL PLATE Set First Well and Last Well as follows First Last Well A1 H4 A5 H1i2 If the reader supports shaking set e Shake Intensity to 4 Variable e Duration to 15 seconds Leave all other parameters set to their defaults Click OK to return to the Main Menu Select Protocol Plate Layout Assign wells to the plate as shown on page 119 Click OK to return to the main menu Select Protocol Save As and give the protocol an identifying name that includes the dispense volume such as Delta 80 and Delta 20 and 5 Synergy HT Operator s Manual 118 Chapter 4 Instrument Qualification Sample KC4 Reading Reading parameters Parameters Dialog for the 80 uL Read Protocol using the Synergy HTTR w Inj rs A jecto scales l Seerning Po Time Hesolyed 9 C abs ats es mA Noms i mo Sample KC4 Reading Parameters Dialog for the 20 amp 5 pL Read Protocol using the ELx808 BioTek Instruments Inc Dispense Module Tests 119 Sample KC4 Plate Layout for the 80 pL Read Protocol Synergy HT Operator s Manual 120 Chapter 4 Instrument Qualification BioTek Instruments Inc 1n eu Is Ag p sosddy PaMalAdy
93. ag has moved or is loose A heavy object is on top of the reader causing the case to interfere with the z axis travel The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft Reference Field Change Notice L0031 Defective or broken optical sensor Fiber optic cable is not tied to the upper probe assembly and has moved into the top probe s path not allowing the top probe to reach the optical sensor Defective motor Motor Controller PCB or cable Upper absorbance lens is loose and has rotated 90 degrees thus jamming the lens into the photodiode Damage to the photodiode can occur 0207 Probe Changer did not home Probable Causes Defective optical sensor Defective motor Motor Controller PCB or cable The probe changer assembly is not able to move because a foreign object is preventing the movement 0208 Syringe 0 did not home also known as the first syringe drive Probable Causes Linear way is dirty or lack of lubrication is causing the bearings to jam The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft Defective optical sensor Defective motor Motor Controller PCB or cable Cable between the dispenser and the Synergy is defective too long or there is a lost connection For certain older dispensers opto cable 7330506 may need to be replaced with 7120734 0209 Syringe 1 did not home also known as the second
94. ages Gen5 and KC4 provide the user with instrument control e The Synergy HT can operate with standard robotic systems such as BioTek s BioStack Microplate Stacker e The intended use of this instrument is dependent on the instrument s labeling If there is an IVD label then the instrument may be used for clinical research and development or other non clinical purposes If there is no such label then the instrument may only be used for research and development or for other non clinical purposes Quality Control It is considered good laboratory practice to run laboratory samples according to instructions and specific recommendations included in the assay package insert for the test to be conducted Failure to conduct Quality Control checks could result in erroneous test data Warranty amp Product Registration Please take a moment to review the Warranty information that shipped with your product Please also register your product with BioTek to ensure that you receive important information and updates about the product s you have purchased You can register online through the Customer Care Center at www biotek com or by calling 888 451 5171 or 802 655 4740 Synergy HT Operator s Manual xiv Preface Warnings A Operate the instrument on a flat surface away from excessive humidity Bright sunlight or strong incandescent light can reduce the linear performance range of the instrument Measuremen
95. ak Ensure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and properly fastened If noise Min is lt 10 e The photodetector is not connected or is defective giving a noise reading of zero e Absorbance channel analog PCB is defective 0810 Absorbance measurement channel failed offset range See noise Max lt 20000 and noise Min gt 10 on the system test This error indicates that during the system test the background electronic signal that was detected is outside of acceptable limits at maximum gain when blocking the light Probable Causes If noise Max is gt 20000 e The photodetector is too noisy and is defective e Absorbance channel analog PCB is defective e A faulty analog PCB or faulty internal grounding may cause internal electronic noise e There may be an ambient light leak Make sure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The bottom and top Synergy HT Operator s Manual 182 Appendix C Error Codes Code Description and Probable Causes shrouds are part of the electrical shielding Check to see if the shrouds are installed and are properly fastened If noise Min is lt 10 e The photodetector is not connected or t
96. al solution is 2 0 at 200 uL Note The choice of dilutions and the absorbance of the original solution can be varied Use Table 2 as a model for calculating the expected absorbances of a series of dilutions given a different absorbance of the original solution Synergy HT Operator s Manual 84 Chapter 4 Instrument Qualification Prepare the Plate e Pipette 200 uL of the concentrated solution from Tube 1 into each well of the first column A1 to H1 of a new flat bottom microplate e Pipette 200 uL from each of the remaining tubes into the wells of the corresponding column of the microplate Tube 2 into wells A2 to H2 Tube 3 into wells A3 to H3 and so on Linearity amp Repeatability Tests 1 Using Gen5 or KC4 read the microplate prepared above five times using Normal mode dual wavelength at 450 630 nm Save the data after each read gt Tip When you create a dual wavelength protocol in KC4 KC4 automatically generates a multi plate transformation for calculating the delta OD The name of the resulting data set is Delta OD Do not discard the plate you will use it for the Alignment test Print out the five sets of Delta OD data or export them to an Excel spreadsheet Data Reduction for Linearity Calculate the mean absorbance for each well and average the means for each concentration Perform a regression analysis on the data to determine if there is adequate linearity For
97. alues and their locations When Gen5 establishes communication with the reader it asks for this information and then stores it in a filter table on the computer To view this table in Gen5 select System Reader Configuration highlight the Synergy reader and click View Modify Click Setup and then click the Fluorescence Luminescence tab Reader Setup Synergy Com6 x Probe Size Absorbance Fluorescence Luminescence Dispenser 1 Dispenser 2 485 7 20 2 Hole x 0 Plug 7 20 3 590 7 30 530 25 4 645 x1 I 40 Excitation p Emission Get Values Center Bandwidth Filter Center Bandwidth 360 7 40 1 460 7 40 Send Values Filter wheel position numbers 1 to 4 mE Regarding the Absorbance Wavelengths table The Synergy HT performs absorbance reads in the range of 200 to 999 nm Click the Absorbance tab to specify and calibrate 6 wavelengths to be made available as default selections within a protocol s Reading Parameters dialog To change the settings and download them to the instrument 1 Enter filter values in the Center fields or use the drop down boxes to select PLUG or HOLE 2 For each Center wavelength value enter its accompanying Bandwidth The Bandwidth is printed on the side of each filter 3 When finished click Send Values to download the information to the reader Clicking Get Values uploads information from
98. an instrument to BioTek for service or repair please contact the TAC for a Return Materials Authorization RMA number before shipping the instrument Repackage the instrument properly see Chapter 2 Installation write the RMA number on the shipping box and ship to this address BioTek Instruments Inc ATTN RMA xxxxx 100 Tigan Street Highland Park Winooski Vermont 05404 USA Contacting BioTek for Applications Support BioTek s fully equipped Application Laboratory provides our on staff scientists with the means to assist you with the integration of our instrumentation and software with your unique scientific applications If you are having difficulty with optimizing fluorescence sensitivity or integrating a unique data reduction transformation or you are just looking for a recommendation on an appropriate fluorophore contact us Phone 888 451 5171 E Mail applications biotek com BioTek Instruments Inc Chapter 2 Installation This chapter includes instructions for unpacking and setting up the Synergy HT and if applicable the external dispense module Instructions are also included for repackaging the reader and dispense module for shipment Product Registration 2 2ssycaeeter a trare teen eee ey de ae gee clas 8 1 Unpack and Inspect the Reader ereere ereenn 9 2 Remove the Shipping Panel cceee eee eeee eee teen eeeeeeee 11 3 Remove the Microplate Carrier Shipping Screw
99. arrier and the surface beneath the carrier with a dry cotton cloth If overflow is significant you may have to remove the shroud of the instrument to better access the surface beneath the carrier See page 135 for instructions on removing the shroud See page 149 for instructions for cleaning the surface beneath the carrier Synergy HT Operator s Manual 128 Chapter 5 Preventive Maintenance Inspecting Cleaning Excitation and Emission Filters Laboratory air is used to cool the lamp and the filters can become dusty as a result The filters should be inspected and cleaned at least every three months You ll need e Isopropyl ethyl or methyl alcohol e Lens cleaning tissue 2 Do not touch the filters with your bare fingers To inspect and clean the Excitation and Emission filters 1 Turn off and unplug the instrument 2 Pull down the hinged door on the front of the instrument Observe the two thumbscrews within the compartment The left thumbscrew holds the excitation EX filter wheel in place the right secures the emission EM filter wheel Remove each thumbscrew and slide the filter wheel s supporting metal bracket straight out of the compartment Chapter 3 Getting Started contains illustrations for identifying the filter wheels and their unique characteristics This chapter also contains instructions for replacing filters if necessary 3 Inspect the glass filters for speckled surfaces
100. as missed flashes or there is an erratic flash during the blank read e Dirty optics or spilled substance on the optics e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the filter is degraded and is not passing enough light energy or is blocking the light Internal Self Test Errors ODO0O 2918 Code 0D00 0D01 0D06 Description and Probable Causes Wavelength calibration data missing during a spectral scan See criteria in text below This error indicates that wavelength data is missing prior to a spectral scan meaning the wavelength has not been calibrated This includes the Gain test data Look for the resets to be lt 1 or gt 8 to fail Probable Causes e Memory is corrupt e The wavelength was not calibrated prior to the read e The gain test skipped or failed for this wavelength Wavelength calibration data missing during an absorbance read See criteria in text below This error indicates that wavelength data is missing prior to an absorbance read meaning the wavelength has not been calibrated This includes self test and Gain test Look for the resets to be lt 1 or gt 8 to fail The last number is the lambda table position number Probable Causes e The wavelength was not calibrated prior to the read e The gain test skipped or failed for this wavelength BioTek Instruments Inc Error Codes 185 Code Description and Probable Causes OEO1 OE06 W
101. ated the reference channel Probable Causes Scenario 1 Fluorescence or System test e The PMT is defective e The connector from the PMT base to the analog board was not built correctly The ground shield is not grounding properly e The PMT base is defective e The PMT analog PCB is defective Scenario 2 Reference channel e The monochromator mirror grating is damaged e Absorbance analog PCB has intermittently failed e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Missed flashes or an erratic flash lamp e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Light beam saturated too much light Air reading reached 65535 This error indicates that filter 3 in absorbance mode has saturated the reference channel Probable Causes e The monochromator mirror grating is damaged e Absorbance analog PCB intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply BioTek Instruments Inc Er
102. ation x Data In e D51 Current Plate 80 ul Read_Disp 1 405 D52 Current Plate 80 ul Read_Disp 1 750 New Data Set Name Delta OD 80 ul_Disp 1 Mode Current Formula com Difference Between Rows I Use single Formula for all wells Difference Between Columns Current Formula D51 DS2 Direction AB iz Write Formula and click in a cell to apply it OBA P21 5 6 Disp_20 Disp_20 7 Disp_20 8 Disp_20 1 Disp_5 1 2 3 4 Disp_80 Disp_80 Disp_0 Disp_80 DS1 DS2 D 1 DS2 D 1 D 2 DS1 Ds2 Disp_80 Disp_80 Disp_80 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_6 Disp_S Disp_5 DS1 D 2 D 1 DS2 DS1 D 2 D 1 Ds2 Disp_80 Disp_80 Disp_s0 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_5 Disp_5 Disp_S Disp_5 DS1 DS2 D 1 DS2 DS1 DS2 DS1 Ds2 Disp_80 Disp_80 Disp_80 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_5 Disp_S Disp_5 DS1 D 2 D 1 DS2 DS1 D 2 D 1 Ds2 Disp_80 Disp_80 Disp_80 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_5 Disp_S Disp_5 DS1 DS2 D 1 DS2 D 1 DS2 DS1 Ds2 Disp_80 Disp_80 Disp_80 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_S Disp_S Disp_5 DS1 D 2 D 1 DS2 D 1 D 2 D 1 Ds2 Disp_80 Disp_80 Disp_80 Disp_80 Disp_20 Disp_20 Disp_20 Disp_20 Disp_S Disp_5 Disp_S Disp_5 DS1 DS2 D 1 Ds2 Ds1 D 2 DS1 Ds2 Disp_80 Disp_80 Disp_80 Disp_
103. avelength combination e If you need to change the wavelength values click the Wavelength List button Click KC4 s Help button for assistance Enter the Peak Wavelength value from the Peak Wavelength Certificate e Note If the certificate specifies multiple peak wavelength values you will need to run an independent test using spectral scan reads KC4 supports the entry of only one Peak Wavelength value and it must be in the range of 580 to 590 nm See the special instructions on the next page Review all of the values you entered and then click OK to save the data The information you just entered will be available in KC4 each time the Absorbance Plate Test is performed Procedure KC4 To run the Absorbance Plate Test using KC4 1 ote ee ee Place the Test Plate in the carrier so the well A1 is in the left rear corner of the carrier as you are facing the carrier In KC4 select System Diagnostics Run Universal Plate Test Enter or select the instrument s Serial Number Select the desired Test Plate Check Peak Absorbance Search e Note If the Test Plate s Peak Wavelength Certificate specifies multiple peak wavelength values do not check Peak Absorbance Search Run an independent test using spectral scan reads instead see the special instructions on the next page BioTek Instruments Inc Absorbance Plate Test 75 Select the Wavelength s to be included in this test by clicking in the Test
104. avelength not found in table Absorbance Fluorescence Luminescence This error indicates that the specified wavelength is not detected in the instrument s filter table The last number is the filter set number in the assay protocol Probable Causes e Wavelength or bandpass was not entered correctly or was missing in filter table e Wavelength or bandpass was entered correctly in the PC software but was never sent to reader e Verify the lambda table and the fluorescence filter table have the wavelengths loaded into the instrument from the controlling PC software Compare the contents of the lambda table and Excitation and Emission filter wheels with the software s filter table OF00 Reference channel correction outside limits See criteria in text below This error can indicate one of the following scenarios e During a spectral scan the flash on value minus flash off is lt 500 normal mode or lt 500 calibrated resets 8 for sweep mode e During a spectral scan the blanking on air uses minimal flashes to test the light performance A ratio is used to determine the performance of the lamp The ratio used is Reference channel blank for wavelength flash on reference data corrected reference dark offset The ratio will fail if lt 0 5 or gt 2 0 Probable Causes e Lamp alignment or lamp power supply failure e Defective reference channel analog PCB or absorbance channel analog PCB or the cable in between e Order sort
105. bable Causes e A faulty analog PCB or faulty internal grounding may cause internal electronic noise e There may be an ambient light leak Make sure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Check to see if the shrouds are installed and are properly fastened e Order sorting filter wheel is jammed not aligning the filter wheel to block the light path or the filter is degraded and is not passing enough light energy Reference Channel Air Blank out of range See criteria in text below This error indicates the Air reading at the time of the plate read was lt 50 of the Air reading at the time of the optic test The last number is the lambda table position number Probable Causes e Flash lamp has missed flashes or there is an erratic flash during the blank read e Dirty optics or spilled substance on the optics e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the filter is degraded and is not passing enough light energy or is blocking the light Measurement Channel Air Blank out of range See criteria in text below This error indicates the Air reading at the time of the plate read was lt 50 of the Air reading at the time of the optic test The last number is the lambda table position number Probable Causes e Flash lamp h
106. bed in Cleaning Internal Components on page 133 To create the Dispense protocol in KC4 1 2 3 10 11 If a Data file is open close it now Data Close Select Protocol New and then Protocol Reading In the Reading parameters dialog e Check the Dispense box and set the Reading Type to Endpoint e Select a Plate Type of 96 WELL PLATE or select a custom format that exactly matches the plate being used for this test e Leaveall other parameters set to their default values Next to the Dispense check box click the Read amp Dispense button In the Read amp Dispense dialog set Read Mode to Plate Click the Dispense button In the Dispense Step dialog e Select Dispenser 1 e Set Priming to Before the process and Volume to 5 uL e Leave Start Time set to 00 00 00 e Set the Volume to 100 uL or an amount to match your assay protocol e Select a Rate adjust the rate to support the dispensing volume e Click OK to close the dialog and add the Dispense step to the list Click OK to close the dialog and return to the Reading parameters dialog Click OK to save the settings and return to the main menu Select Protocol Save As and give the protocol an identifying name such as Dispenser 1 Observation Repeat these steps to create a protocol for Dispenser 2 Run the two protocols on two different plates When finished visually assess the fluid level in the wells for accuracy If the well volume appears to
107. bing and Injector Heads seceeeee eee eeee 159 Clean the Tip Prime Trough and Priming Plate seeeeeee eee 160 Alternate Procedure for Models with Injectors sseeeeeeeeeeeeeee ees 161 Appendix B Computer Control ccccsscceeeeeeneeceeeeeseeeeensesseseesseseessenees 163 Appendix C Error COde S cccccccscseecceenescesseeseeseeeeseeaeeaneaseasensensensenees 165 Appendix D Specifications cccsccsceccencenceeeeeeeeeeeneceeseeneeeeseeseensenees 207 Appendix E Instrument Dimensions for Robotic Interface 215 BioTek Instruments Contact Information vii Contact Information 2 For more detailed information on contacting BioTek for product support and service turn to page 6 BioTek Instruments Inc Highland Park P O Box 998 Winooski Vermont 05404 0998 USA Customer Service and Sales Internet www biotek com Phone 888 451 5171 toll free in the U S 802 655 4740 outside the U S Fax 802 655 7941 E Mail customercare biotek com Service TAC Phone 800 242 4685 toll free in the U S 802 655 4740 outside the U S Fax 802 654 0638 E Mail tac biotek com European Coordination Center Authorized European Representative BioTek Instruments GmbH Kocherwaldstrasse 34 D 74177 Bad Friedrichshall Germany Internet www biotek de Phone 49 0 7136 9680 Fax 49 0 7136 968 111 E Mail info biotek de Synergy HT Operator s Manual
108. blems and solutions gt Ensure that the Test Plate is correctly seated in the microplate carrier gt Check the four alignment holes A01 A12 H01 H12 to ensure they are clear of debris gt Check the microplate carrier to ensure it is clear of debris Accuracy Accuracy is a measure of the optical density of Test Plate wells C01 D04 E02 F05 G03 and H06 as compared with known standard values contained in the Standards Certificate that accompanies each Test Plate If the reader fails this test review the following possible problems and solutions gt Check the neutral density filters on the Test Plate to ensure they are clean If necessary clean them with lens paper Do not remove the filters from the test plate and do not use alcohol or other cleaning agents BioTek Instruments Inc Absorbance Plate Test 79 gt Verify that the filter calibration values entered in Gen5 or KC4 are the same as those on the Test Plate s Standards Certificate gt Verify that the Test Plate is within its calibration certification period The calibration sticker is affixed directly to the plate If it is out of date contact BioTek to schedule a recertification e Repeatability Repeatability is a measure of the instrument s ability to read the same well with minimum variation between two reads with the well in the same location If the reader fails this test review the following possible problems and solutions g
109. brication e Defective or broken optical sensor e The syringe valve did not open e The glue between the lead screw and the motor has separated Previous syringe move produced an FMEA sensor clear error This error indicates that the previous move caused the syringe to lose steps Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Linear way is dirty or needs lubrication e Defective or broken optical sensor e The syringe valve did not open e Syringe was not installed correctly or was not cleaned causing stress on syringe movement by not allowing the syringe to move freely e The glue between the lead screw and the motor has separated Number of microsteps requested for move too large This error indicates that during a dispense or move home function the number of steps to move was gt the maximum syringe travel 944 full steps Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Program or user error Trough plate not in carrier for prime or purge This error indicates that prior to a prime or purge the priming plate was not in the carrier Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e The plate or trough being used is not opaque enough e There is no priming plate or trough in carrier Syringe calibration data not set This error indicates that configuration data for either syringe has not been entered or the data is corrupted This c
110. cceeeeeeeeeeeeees FEAUUIES erig voice cadena a AE A Seeder candela dade ERRELE Package Contentsixcscccccceneentesiaes AAA AARAA TAERA ENEE EERTE LE Optional Accessories priri aa L EER SE Product Support amp Service ssssssrrrsssrrrrrsrrrrrrsnrrrrrnnrrnrenrrrnns Technical Assistance Center T AC sssssssssrssrsssresrssrrrrrrrrress Returning Instruments for Service REP ir cccceeeeeeeeeeeeee ees Contacting BioTek for Applications SUPPOrt ccee cece eee eee eee 2 Chapter 1 Introduction Synergy HT Multi Mode Microplate Reader The Synergy HT is a single channel absorbance fluorescence and luminescence microplate reader It is computer controlled using BioTek s Gen5 or KC4 PC software for all operations including data reduction and analysis Note Synergy HT basecode software version 2 24 or greater is required for use with Gen5 The Synergy HT is robot accessible and compatible with BioTek s BioStack Microplate Stacker When making fluorescence determinations the Synergy HT uses a tungsten quartz halogen lamp with interference filters for wavelength specificity in conjunction with a photomultiplier PMT tube detector The Synergy HT has both top and bottom probes for fluorescence measurements The top probe can be adjusted vertically for the correct reading height via Gen5 s KC4 s Top Probe Vertical Offset reading parameter see Chapter 3 Getting Started Luminescence is measured by the lo
111. ce pursuant to Part 15 of the FCC Rules These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment Like all similar equipment this equipment generates uses and can radiate radio frequency energy and if not installed and used in accordance with the instruction manual may cause harmful interference to radio communications Operation of this equipment in a residential area is likely to cause interference in which case the user will be required to correct the interference at his own expense Canadian Department of Communications Class A This digital apparatus does not exceed Class A limits for radio emissions from digital apparatus set out in the Radio Interference Regulations of the Canadian Department of Communications Le present appareil numerique n met pas du bruits radioelectriques depassant les limites applicables aux appareils numerique de la Class A prescrites dans le Reglement sur le brouillage radioelectrique edicte par le ministere des Communications du Canada User Safety This device has been type tested by an independent laboratory and found to meet the requirements of the following Underwriters Laboratories UL 61010A 1 ist edition 2002 Electrical Equipment for Laboratory Use Part 1 General Requirements Canadian Standards Association CAN CSA C22 2 No 1010 1 1992 Safety requirements for electrical equipment for measurem
112. ck Gen5 s Help button for assistance 7 Select the number of Peak Wavelength tests to run 1 to 4 based on the number of peak wavelength values provided on the Peak Wavelength Certificate 8 Enter the Expected Peak value s from the 2 4 nm Spectral Bandpass column of the Peak Wavelength Certificate Select 1 4 Peak Wavelengths in the range that the instrument is typically operated Peak wavelength tests 1 F Expected Peak TestRange 586 fe 4 580 to 590 Note For certificates that have only one peak wavelength and a fixed wavelength range of 580 to 590 nm enter the Expected Peak wavelength value and adjust the Test Range values so the range displayed in parentheses is 580 to 590 as demonstrated above 9 Review all of the values you entered and then click OK to save the data The information you just entered will be available in Gen5 each time the Absorbance Plate Test is performed The information must be updated whenever the Test Plate is recalibrated BioTek Instruments Inc Absorbance Plate Test 73 Procedure Gen5 To run the Absorbance Plate Test using Gen5d 1 2 3 In Gen5 select System Diagnostics Test Plates Run If prompted select the desired Test Plate and click OK When the Absorbance Test Plate Options dialog appears check Perform Peak Wavelength Test if it is not already checked Highlight the wavelength s to be included in this test e Not
113. col described on page 101 Test Solutions 1 Prepare the buffer CBB solution e Optional but recommended Using a 0 45 micron filter filter 200 mL of deionized or distilled water e Open and dissolve the contents of 2 CBB capsules do not dissolve the outer gelatin capsule into 200 mL of the water e Stir the solution preferably using a stir table until the CBB is completely dissolved 2 Prepare the MUB stock solution e Add1mL of 100 methanol to the 10 mg vial of MUB e Make sure all of the dye has completely dissolved and is well mixed This yields a 10 mg mL stock solution e Wrap the solution in aluminum foil to prevent exposure to light BioTek Instruments Inc Fluorescence Tests 99 3 Prepare the dilutions Label each with MUB and the concentration Mix this MUB solution 4 5 mL of 100 0 5 mL of 10 mg mL stock solution methanol 1 mg mL 0 88 mL of 1 mg mL solution 4 12 mL of CBB 176 ug mL 0 1 mL of 176 g mL solution 9 9 mL of CBB 1 76 pg mL 0 5 mL of 1 76 g mL solution 4 5 mL of CBB 176 ng mL 17 6 ng mL 100 nM 1 mL of 176 ng mL solution 9 mL of CBB Procedure 1 Create the Gen5 or KC4 protocol see page 101 2 If you have not already done so prepare the test solutions see page 98 3 Refer to the Pipette Map on page 100 and pipette the solutions into a clean 96 well solid black plate 4 Read the plate using the Synergy HT FI_MUB prt protocol 5 Sav
114. column 1 and C10 F12 Pipette 150 uL of the 1 nM SF solution into wells C1 F1 Discard the tips Pipette 150 uL of the 1 nM SF solution into wells C2 F2 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from wells C2 F2 and dispense into wells C3 F3 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from wells C3 F3 and dispense into wells C4 F4 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from wells C4 F4 and dispense into wells C5 F5 Mix the wells using the pipette Aspirate 150 uL from wells C5 F5 and discard 1 2 3 4 5 6 7 8 9 10 11 12 3 3 3 3 Be 3 3 3 3 523 nM nM nM SEE St nM nM nM CBUF CBUF CBUF CBUF CBUF CBUF CBUF CBUF 1 0 0 5 0 25 0 125 0 062 nM nM nM nM nM BUF BUF BUF 1 0 0 5 0 25 0 125 0 062 nM nM nM nM nM BUF BUF BUF 1 0 0 5 0 25 0 125 0 062 nM nM nM nM nM BUF BUF BUF 1 0 0 5 0 25 0 125 0 062 nM nM nM nM nM BUF BUF BUF CBUF CBUF CBUF CBUF CBUF CBUF CBUF CBUF 323 3 3 33 3 3 Bas 513 nM nM nM geU geur nM nM nM BioTek Instruments Inc Gen5 Protocol Reading Parameters Fluorescence Tests 95 The following tables contain the recommended settings for the Gen5 protocols Your tests may require modifications to some of these parameters such as Plate Type or Sensitivity see Troubleshooting on page 93 The Plate Type setting shou
115. cure Protocol2 PE T File Protocol System Help AT EE Ke E Procedur A Procedure Synergy Com2 xi Plate Layo i m Add St E Data Reduction E Plate Type 96 WELL PLATE 7 Advanced Options fa Runtime Prompts Read Report Builder iB File Export Builder Dispense E Power Export Builder Shake T Data Views Protocol Options ___Peky 2 Audit Trail Kinetic Monitor Well Set Temperature Plate Out In Stop Resume r Synchronized Modes gt Figure 33 Editing the Procedure in a new Protocol 3 Perform the steps in the following three sections to define the Procedure customize the Plate Layout and add Data Reduction steps to test Dispenser 1 4 When you re finished select File Save As and give the protocol an identifying file name such as Synergy HT Dispenser 1 prt 5 Repeat steps 2 4 to create a protocol to test Dispenser 2 BioTek Instruments Inc Ss Gen5 Secure Experiment1 created from Synergy HT Dispenser 1 Test_prt File Plate Protocol Window System Help Dispense Module Tests 111 Define the Procedure In brief the protocol s procedure follows the sequence below After each Dispense step the plate is ejected to allow the operator to weigh it and then tare the balance e Dispense 80 uL dye to columns 1 4 e Dispense 20 uL dye to columns 5 8 e Dispense 5 uL dye to columns 9 12 e Shake the plate for 15 seconds e
116. d Warning Potential Biohazards Some assays or specimens may pose a biohazard Adequate safety precautions should be taken as outlined in the assay s package insert Always wear safety glasses and appropriate protective equipment such as chemically resistant rubber gloves and apron Warning Hot Surface The fluorescence lamp assembly is hot when the instrument is turned on Turn off the reader and allow the lamp to cool down before attempting to replace it Warning Unspecified Use Failure to operate this equipment according to the guidelines and safeguards specified in this manual could result in a hazardous condition BioTek Instruments Hazards and Precautions Xv Warning Software Quality Control The operator must follow the manufacturer s assay package insert when modifying software parameters and establishing reading methods Failure to conduct quality control checks could result in erroneous test data Warning Pinch Hazard Some areas of the Dispense Module can present pinch hazards when the instrument is operating These areas are marked with the symbol shown here Keep hands fingers clear of these areas when the instrument is operating Precautions The following precautions are provided to help avoid damage to the instrument service personnel Only qualified technical personnel should perform troubleshooting Caution Service The Synergy HT should be serviced by BioTek authorized A and service procedure
117. defective e Absorbance analog PCB has intermittently failed Scenario 2 Reference channel e The monochromator mirror grating is damaged e Absorbance analog PCB has intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Scenario 3 fluorescence read e Chemistry is too strong e Sensitivity is too high 0506 Light beam saturated too much light Air measurement channel reading reached 65535 This error indicates that filter 6 in absorbance mode has saturated the reference channel Probable Causes e The monochromator mirror grating is damaged e Absorbance analog PCB has intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Synergy HT Operator s Manual 178 Appendix C Error Codes Code 0508 0510 0511 0516 Description and Probable Causes Light beam saturat
118. dels With Injectors ccccceeeeeeeee eee 157 Clean Exposed Surfaces awiied vest wives nasi ntsaversdeonterdectdecadsvans 157 Decontaminate the Fluid LINGS c ccccceceesseeeeeeeseeeesaeeeaas 158 Rinse the Fluid Lines s sssssssssssesrssesssnensnrnreressesesssnenrnns 159 Clean the Internal Tubing and Injector Heads ceeeeeee 159 Clean the Tip Priming Trough and Priming Plate 0 5 160 Alternate Procedure for Models with Injectors ccceseeeseeeeaees 161 154 Appendix A Decontamination Purpose Any laboratory instrument that has been used for research or clinical analysis is considered a biohazard and requires decontamination prior to handling Decontamination minimizes the risk to all who come into contact with the instrument during shipping handling and servicing Decontamination is required by the U S Department of Transportation regulations Persons performing the decontamination process must be familiar with the basic setup and operation of the instrument BioTek Instruments Inc recommends the use of the following decontamination solutions and methods based on our knowledge of the instrument and recommendations of the Centers for Disease Control and Prevention CDC Neither BioTek nor the CDC assumes any liability for the adequacy of these solutions and methods Each laboratory must ensure that decontamination procedures are adequate for the Biohazard s they handle Wear proph
119. der s optics and readings reducing its linear performance range e Dust Readings may be affected by extraneous particles such as dust in the microplate wells A clean work area is necessary to ensure accurate readings Note If you will be installing BioTek s BioStack Microplate Stacker for operation with the Synergy HT you may wish to seat the BioStack and the reader in their aligning plates at this time Refer to the Installation chapter in the BioStack Operator s Manual for more information Synergy HT Operator s Manual 16 Chapter 2 Installation 6 Connect the Power Supply Warning Power Rating The power supply must be connected to a power receptacle that provides voltage and current within the specified rating for the system Use of an incompatible power receptacle may produce electrical shock and fire hazards Warning Electrical Grounding Never use a two prong plug adapter to connect primary power to the power supply Use of a two prong adapter disconnects the utility ground creating a severe shock hazard Always connect the power cord directly to an appropriate receptacle with a functional ground Perform these steps to connect the power supply 1 Connect the power cord to the external power supply Locate the power inlet on the rear of the reader 2 3 Plug the rounded end of the power supply s line cord into the power inlet 4 Plug the power cord into an appropriate power receptac
120. der onto its back feet Attach the shipping panel to the bottom of the reader using the four flat head screws and washers see p 11 and 11 Wrap the plastic bag around the reader and shipping panel Locate the original outer shipping box Place four foam blocks in the four bottom corners of the box Place the inner shipping box inside the outer box see p 8 and 9 Grasp the handles on the shipping pane and carefully lower the reader into the inner shipping box Slide the foam vertical supports into place around the reader Place the accessories box on top Close and seal the inner box with tape Place four foam corner blocks around the inner shipping box Close and seal the outer box with tape Write the RMA number in large clear numbers on the outside of the box Ship the box to BioTek See page 6 for the address Synergy HT Operator s Manual 34 Chapter 2 Installation Perform these steps to prepare the dispense module for shipment 1 If you have not already done so a Contact BioTek s Technical Assistance Center for an RMA Return Materials Authorization number before returning equipment for service See page 6 for contact information b Decontaminate the module according to the instructions in Appendix A c Removethetwo syringes see step 3 on the previous page and store then in their original boxes d Detach the dispense module outlet tubes and communication cable from the reader Replace the two n
121. dified 7 filter test plate Simplified the process for creating Titration Dyes for the Fluorescence SF Sensitivity Test Added information to the pass fail criteria table for the SF Sensitivity Test Clarified that for models without injectors the reader s internal chamber and optical probes are not user accessible for cleaning Updated the Error Codes appendix with recent information Additional minor cosmetic changes throughout Added modified instructions throughout to support Gen5 including Chapter 2 Installation Added instructions for installing software establishing communication with the reader installing testing dispense module components Chapter 3 Getting Started Added introductory information for new Gen5 users Chapter 4 Instrument Qualification Added instructions for performing the System Test Absorbance Plate Test and Dispense Accuracy amp Precision Test Chapter 5 Preventive Maintenance Added instructions for creating the optional Dispense protocol in Gen5 11 2008 Throughout Changed product description from Multi Detection to Multi Mode Changed Bio Stack to BioStack Preface Corrected Service TAC fax number Updated the Intended Use section with respect to IVD labeling Added cautions for Electromagnetic Environment and Compatibility Updated Directives Added Pinch hazard to Hazards and Precautions Added Consult Instructions for use
122. dispenses To create the Dispense protocols start by launching KC4 and setting the current reader to the Synergy HTTR w Injectors Perform the following set of steps three times to create six protocols for the 80 20 and 5 ul tests for Dispenser 1 and Dispenser 2 1 Ifa Data file is open close it now Data Close 2 Select Protocol New and then Protocol Reading The Reading Parameters dialog opens 3 Select the Dispense box and set the Reading Type to Endpoint Reading parameters m Reading Type TM At eader Syner End Point Urea C Kineti Detection method jis C Absorbance l G Spectrum MM Dispense Read amp Dispense 4 Click the Read amp Dispense button The Read amp Dispense dialog opens see the screen shot on the next page 5 Set Read Mode to Plate 6 Click the Dispense button and set the following e Set Dispenser to 1 e Set Priming to Before the dispense e Set the Tip Prime Volume Dispense Volume and Rate as follows Tip Prime Dispense Volume Volume 20 uL 80 UL well 275 uL sec 20 uL 20 uL well 250 uL sec 5 uL well 225 uL sec Synergy HT Operator s Manual 116 Chapter 4 Instrument Qualification 10 11 12 13 14 Dispense step before the dispense D Dispense Steps Jo Shortest after fa Click OK OK to return to the Reading Parameters dialog Select the appropriate Plate Type o
123. e You need to select only those wavelengths most appropriate for your use of the reader Optional Enter any Comments Click Start Test Place the Test Plate in the microplate carrier so that well A1 is in the left rear corner of the carrier as you are facing the carrier Click OK to run the test When the test completes the results report will appear Scroll down through the report every result should show PASS See page 78 for information on results and troubleshooting tips in the event of failures e A sample test report is shown on pages 76 and 77 e Gen5 stores the results in a database they can be retrieved and printed at any time We recommend you print and save the report to document that the test was performed Synergy HT Operator s Manual 74 Chapter 4 Instrument Qualification Setup KC4 To define the Absorbance Test Plate parameters using KC4 1 2 Obtain the certificates that came with the Test Plate Launch KC4 and select System Diagnostics Define Universal Plates Note The terms Universal Plate and Absorbance Plate are equivalent Click Add The Edit Universal Plate dialog will appear Select the appropriate Plate Type and enter the plate s Serial Number If the wavelength values in the top row of the grid are appropriate for your tests carefully enter the standard OD values from the Standards Certificate into the grid Be sure to enter the correct value for each well w
124. e 74 Results amp Troubleshooting TiPS ccceeeeeeeee eee ee teeta eee eee eeeaeeae es 78 Absorbance Liquid Tests nan gninnan E AAA EA A EA a ene ean 80 Stock Solution Formulation ssssssssssssssssssrssrrrnrrrrrrrrrrrrrrrrrnnunnrnrrssras 81 Liquid TeSt ls roii eines se ais valet et E A TEA E AAE 82 LiQUid TeSt 2e oai a aa NE vette need adi a Aa A EAA AA WEEE 83 LIQUich EEE Ee E E A E EAA EN 86 Fl orescence TeStsrciiciosireiii naida ete aed i a a a aa 89 Required MaterialSi eere e a aa a AAE ARTA 90 Test SolUtiONS sree imadni ante ai a i a a Ta aS EEn 91 BioTek Instruments Contents v Pro ed UE a ola A cai e Seed deadewa land boa AA ARA TEENETE 92 Results AnalySiS uen prstane ine a aa ia aeaa 92 Troubleshooting e anaa caters a a AAAA AAE ATASE NEAN ean 93 Pipette Maps cernes aeea ees EAE TA E EN AR eeu EAEE 94 Gen5 Protocol Reading Parameters sssssrssssserrresurrrrnuenrrrnnnenenrnn 95 KC4 Protocol Reading Parameters ssssssssrrrrssrerrrnrsrrrrnnrsrerrnrrsees 96 Fluorescence Tests Using Methylumbelliferone ccccsseeeeeeeeeeeeees 98 Dispense Module TeStsS cccccccccsseesseeeeeeeeeeeeeeeeeeeeeaeeeeeneee nant nant egees 102 Required MaterialS cccccscesscesseeseeeseeeeeeseeesaeeseeeeeegeenasenanenags 103 Test Solution RECIPES cccceeeeseeeeeeeeeeseeeseesaetsaeeseeeaneseetansnanenags 104 Test Setup GEN eis hues tea ag oh ees can dy Satan anda sat aeaadaaeans seeee dans taee 105 Test Setup
125. e and or print the measurement data 6 Calculate and analyze the results as described below Results Analysis Sensitivity Test 1 Calculate the Mean and Standard Deviation for the buffer wells C10 F12 2 Calculate the Mean for the 17 6 ng mL 100 nM MUB solution wells C1 F1 3 Calculate the Detection Limit in ng mL 17 6 Mean MUB Mean Buffer 3 Standard Deviation Buffer Detection Limit must Optic Probe be less than Top 1 5mm 0 31 ng mL Linearity Test 1 Calculate the Mean of the four wells for each concentration in columns 1 5 2 Perform linear regression using these values as inputs Synergy HT Operator s Manual 100 Chapter 4 Instrument Qualification mean of the 100 nM wells mean of the 50 nM wells mean of the 25 nM wells mean of the 12 5 nM wells mean of the 6 25 nM wells 3 Calculate the R Squared value it must be greater than or equal to 0 950 to pass Pipette Map 2 Seal the plate with foil or store it in a black polyethylene bag until use Use a multi channel pipette with just 4 tips installed Perform these instructions carefully and refer to the grid below Pipette 150 uL of CBB buffer into wells C2 F5 and C10 F12 Pipette 150 uL of the 17 6 ng mL 100 nM MUB solution into wells C1 F1 Discard the tips Pipette 150 uL of the 17 6 ng mL solution into wells C2 F2 Mix the wells using the pipette Do not discard the tips Aspirate 150 uL from
126. e incubator to reach its set point before running the System Test To access this feature select System Reader Control and click the Pre Heating tab Gen5 Select System Diagnostics Run System Test KC4 Select System Diagnostics Run Optics Test When the Run Optics Test dialog appears click Start If the test fails during execution a message box will appear in the software Close the box the test report will contain the error code that was generated by the failure When the test is complete a dialog will appear requesting additional information Enter the information if desired and then click OK The test report will appear Scroll down toward the bottom of the report it will show either SYSTEM TEST PASS or SYSTEM TEST FAIL ERROR error code DETECTED Print the report if desired gt A sample test report is shown on the next few pages gt Gen5 and KC4 store the results in a database so the results can be retrieved printed at any time We recommend that you print and save the reports to document that the test was performed If the test failed look up the error code in Appendix C Error Codes to determine its cause If the cause is something you can fix turn off the reader fix the problem and then turn the reader back on and retry the test If the test continues to fail or if the cause is not something you can fix contact BioTek s Technical Assistance Center see page 6 for contact infor
127. e into the read chamber An object may be obstructing the carrier s path 0202 EX Filter Wheel did not home Probable Causes Filter wheel is not inserted into to the EX assembly Filter wheel is not moving due to an obstruction or because the filter is not clipped in Filter wheel is not moving because gear teeth on the filter wheel are binding with the gear teeth on the motor Defective or broken Hall Effect sensor Defective Motor Controller PCB or cable Motors failed due to the bearings 0203 EM Filter Wheel did not Home Probable Causes Filter wheel is not inserted into the EM assembly Filter wheel is not moving due to an obstruction or because the filter is not clipped in Filter wheel is not moving because the gear teeth on the filter wheel are binding with the gear teeth on the motor Defective or broken Hall Effect sensor Defective Motor Controller PCB or cable Motors failed due to the bearings 0204 Monochromator Filter Wheel did not home order sorting filters Probable Causes Filter wheel is not tight and wobbles Filter wheel is not moving because it is too close to the motor gear and is binding Defective or broken optical sensor Defective motor Motor Controller PCB or cable Motors failed due to the bearings Synergy HT Operator s Manual 170 Appendix C Error Codes Code Description and Probable Causes 0206 Z axis Top Probe did not home Probable Causes The optical trigger fl
128. ection Appendix C Error Codes Corrected the range of error codes under Home Sensor Initial Find Errors Removed some text from the Status String Format section that was misleading Synergy HT Operator s Manual xii Preface Document Conventions This icon calls attention to important safety notes A Warning indicates the potential for bodily harm and tells you how to avoid the problem A Caution indicates potential damage to the instrument and tells you how to avoid the problem Bold text is primarily used for emphasis Topics that apply only to specific Synergy HT models are preceded by a notice in italics for example Applies only to Synergy HT models with injectors This icon calls attention to important information BioTek Instruments Intended Use Statement xiii Intended Use Statement The Synergy HT is a single channel absorbance fluorescence and luminescence micro plate reader that uses a dual optics design to perform measurements of samples in a microplate format The performance characteristics of the data reduction software have not been established with any laboratory diagnostic assay The user must evaluate this instrument and PC based software in conjunction with the specific assay This evaluation must include the confirmation that performance characteristics for the specific assay are met e This system is designed for use with PC based software only BioTek s software pack
129. ed too much light Air measurement channel reading reached 65535 This error indicates that during background tests or prior to a read the PMT reached saturation When at rest the PMT is charged to 650 volts to maintain stability for the next read The Synergy is constantly testing the PMT when idle If light reaches the PMT this error can occur Probable Causes e Light leakage or bright light in the read chamber e The EX filter wheel cartridge was removed prior to a Time Resolve read and ambient or light from the lamp reached the PMT e Fluorescence analog PCB has intermittently failed e EM and EX filter wheel overlap If a hole in the EM wheel is required try positions 1 or 4 Basecode v2 14 or higher has greatly reduced the possibility of filter wheel overlap Reference channel light beam saturated too much light Air reading reached 65535 This error can indicate one of the following scenarios e 1 During an absorbance filter calibration or wavelength scan the measurement channel was saturated when storing to memory e 2 During a spectral scan read the measurement channel was saturated for one of the wavelengths Probable Causes Scenario 1 Filter calibration e The monochromator mirror grating is damaged e Absorbance analog PCB has intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole i
130. eet 3 of 3 The format varies depending on the software used Synergy HT Operator s Manual 70 Chapter 4 Instrument Qualification Absorbance Plate Test Description This test uses BioTek s Absorbance Test Plate PN 7260522 to confirm the mechanical alignment optical density accuracy linearity and repeatability and wavelength accuracy of the Synergy HT The Absorbance Plate Test compares the reader s optical density and wavelength measurements to NIST traceable values 7 kS An alternate method that may be used to determine accuracy linearity and repeatability is Liquid Test 2 described on page 83 The Absorbance Plate Test confirms the following Mechanical Alignment The Test Plate has precisely machined holes in its four corners The amount of light that shines through these holes is an indication of how well the reader is aligned A reading of more than 0 015 OD for any of the designated alignment holes indicates that the light is being clipped and the reader may be out of alignment Accuracy Linearity The Test Plate contains neutral density glass filters of known OD values at several wavelengths Actual measurements are compared against the expected values provided in the Test Plate s Standards Certificate Since there are several filters with differing OD values the accuracy across a range of ODs can be established Once it is proven that the reader is accurate at these OD values the reader
131. eiey ede itches A i ia EANGE vii REVISION HISTORY forneare EI A EAA Seiko EErEE AEAEE IRAE ANS viii DOCUMENt CONVENTIONS ree srair ieira Tant eia aaa EA TEENAA EE KARENE viii Intended Use Statement sssssssrsssssssesssurernorrorrrerrnnrenrnnnennannnennennas xiii Quality Control aeaa ntaa ange A A EEE EDA senda a AEREE IAE ADER AA xiii Warranty amp Product Registration ecceee eee e cece eee ee eee eee teense een es xiii Warnings oes ieee Povo dante ce ct ene dice ately ended eters Pen ealeaaledeets ait uce eating aie eebeeeae xiv Hazards and Precautions ccccceccecee cece eee eee eee eee Ene EEE Eee xiv DiFGCtiveS wissen ia a Ad E dah eee ea ie dae Rae ERE Xvi Electromagnetic Interference and Susceptibility cccccceeeeeeeeeeeeees xvii User Safety voice vita tid van ead MO one Tal tea eae aaa oe A xvii Safety SVMDOIS ieeivecu ease deci segisetdevsinch L een ol sce eorr dete E xviii Chapter 1 Introduction sosina ananasa Aaaa n raa raaa aariaa 1 Synergy HT Multi Mode Microplate Reader sssssssssssssssssrerrrrrrrrerrrens 2 FEatUr eSa nein eain a aa ving een a a a aE delete EDAN aA A aa AEREN ani 3 Package CONTENTS icarii oa a aAa AEA AN a aaea 4 Optional AcCeSSOMES csicngrinai inaa a aa aa a aa 5 Product Support amp Service s s sssssssrssssessesrssssrroerrorrennennnenrennennannnennas 6 Technical Assistance Center TAC ssessssssrrsrrsrrsnrrsnrssrssssersesseenns 6 Returning Instruments for Service Repa
132. elow Table 1 Typical Enzyme Substrate Combinations and Stopping Solutions Alkaline Phosphate o nitrophenyl phosphate 3N sodium hydroxide beta o nitrophenyl beta D 1M sodium carbonate Galactosidase galactopyranoside 2 2 Azino di ethylbenzothiazoline R Peroxidase sulfonic acid ABTS citrate phosphate buffer pH 2 8 o phenylenediamine 0 03N sulfuric acid BioTek Instruments Inc Absorbance Liquid Tests 81 Stock Solution Formulation The stock solution for Liquid Tests 1 and 2 may be formulated from the ingredients listed below A or by diluting a dye solution available from BioTek B Either set of instructions should create a solution with an absorbance of about 2 0 when using 200 uL in a flat bottom microwell The OD value result will be proportional to the volume in the well and the amount of FD amp C No 5 dye used You can use a larger or smaller well volume or add more dye or water to adjust the solution Note that too small a well volume may result in increased pipetting related errors Solution A Required Materials e Deionized water e FD amp C Yellow No 5 dye powder typically 90 pure e Tween 20 polyoxyethylene 20 sorbitan monolaurate or BioTek wetting agent PN 7773002 e Precision balance with capacity of 100 g minimum and readability of 0 001 g e Weigh boat e 1 liter volumetric flask Procedure 1 Weigh out 0 092 g of FD amp C yellow No 5 dye powder i
133. em and User s Guide for complete instructions Note KC4 software versions 3 4 and greater offer a Multi Mode option within the Reading Parameters dialog This option allows you to define more than one detection method in a single protocol See KC4 s Help system or User s Guide for more information Synergy HT Operator s Manual 54 Chapter 3 Getting Started To create an Absorbance protocol inKC4 1 Select Data New Plate If prompted to select a protocol select Empty Protocol and click OK If not prompted select Protocol New Select Protocol Reading The Reading parameters dialog will appear Set Detection Method to Absorbance Select a Reading Type of Endpoint Kinetic or Spectrum For a Kinetic protocol click the Kinetic button to define the Run Time and Interval Select or specify the Wavelengths at which the plate will be read Select a Plate Type Enable Temperature Control and or Shaking if necessary Select any Pre Reading parameters Click OK to verify the parameters and return to the main screen Select Protocol Save As and give the protocol an identifying name To create a Fluorescence or Luminescence non Dispense protocol in KC4 1 10 Select Data New Plate If prompted to select a protocol select Empty Protocol and click OK If not prompted select Protocol New Select Protocol Reading The Reading parameters dialog will appear Set Detection Method to Fl
134. ent control and laboratory use part 1 general requirements Synergy HT Operator s Manual xviii Preface Safety Symbols Some of the following symbols may appear on the instrument ONS l Alternating current Courant alternatif Wechselstrom Corriente alterna Corrente alternata Direct current Courant continu Gleichstrom Corriente continua Corrente continua Both direct and alternating current Courant continu et courant alternatif Gleich und Wechselstrom Corriente continua y corriente alterna Corrente continua e corrente alternata Earth ground terminal Borne de terre Erde Betriebserde Borne de tierra Terra di funzionamento Protective conductor terminal Borne de terre de protection Schutzleiteranschluss Borne de tierra de protecci n Terra di protezione On Supply Marche alimentation Ein Verbindung mit dem Netz Conectado Chiuso Off Supply Arr t alimentation Aus Trennung vom Netz Desconectado Aperto sconnessione dalla rete di alimentazione BioTek Instruments Safety Symbols xix Caution refer to accompanying documents Attention voir documents d accompanement Achtung siehe Begleitpapiere Atenci n vease los documentos incluidos Attenzione consultare la doc annessa Warning risk of electric shock Attention risque de choc lectrique Gef hrliche elektrische schlag Precauci n riesgo de sacudida el ctrica Attenzione rischio di scossa elettrica
135. ent chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and are properly fastened e Order sorting filter wheel is jammed and not aligning the filter wheel to block the light path or the filter is degraded and not passing enough light energy 0910 The absorbance measurement channel Dark current value failed See Dark on the system test See criteria in text below This error can indicate one of the following scenarios e The measurement channel failed during a read or spectral scan for one of the following reasons Dark value was lt 100 during a spectral scan using 8 flashes and 8 resets during sweep mode Dark value was lt 100 during a spectral scan using 8 flashes and the number of resets saves for that wavelength during normal and rapid mode e The measurement channel failed lt 100 or gt 20000 during filter calibration or spectral scan with the flash on e The measurement channel failed lt 100 during filter calibration or spectral scan with the flash off Probable Causes If failed lt 100 e Absorbance analog PCB or measurement channel analog PCB is defective e Shielding of the cable between the measurement channel and analog PCB is defective or disconnected e Measurement channel photodetector is defective If failed gt 20000 e Measurement channel photodetector is defective e A faulty analog PCB or faulty internal grounding may cause internal elec
136. ents Inc Recommendations for Achieving Optimum Performance 59 Recommendations for Achieving Optimum Performance e Microplates should be perfectly clean and free of dust or bottom scratches Use new microplates from sealed packages Do not allow dust to settle on the surface of the solution use microplate covers or seals when not reading the plate Filter solutions to remove particulates that could cause erroneous readings e Before preparing your microplates make sure the instrument is on and successfully communicating with the controlling software You may want to run a System Test if the instrument has not been turned off on in a few days Design your Gen5 or KC4 protocol in advance as well to ensure that the intended reading parameters are used and to avoid any last minute corrections e Although the Synergy HT supports standard flat U bottom and V bottom microplates the reader achieves optimum performance with optically clear flat bottomed wells See Appendix D Specifications for more information on the supported plates e Non uniformity in the optical density of the well bottoms can cause loss of accuracy especially with U and V bottom polyvinyl microplates Check for this by reading an empty microplate Dual wavelength readings can eliminate this problem or bring the variation in density readings to within acceptable limits for most measurements e Inaccuracy in pipetting has a large effect on measurements especia
137. er Ports in the Hardware Device Manager area of System Properties e g USB Serial Port COM 5 Click the Current Reader button KC4 will attempt to communicate with the reader If a message appears stating that the software wavelength table does not match the reader filter table this means that communication has been established and KC4 just needs to update its wavelength table Click Yes e If the communication attempt is successful click Close to return to KC4 s main screen e If the communication attempt is not successful i e an error message appears in KC4 try the following gt Isthereader connected to the power supply and turned on gt Isthe communication cable firmly attached to both the reader and the computer gt Did you select the correct model in step 4 gt Choosea different COM port gt If using the USB cable did you install the driver software See 9 Connect the Host Computer If you remain unable to get KC4 and the reader to communicate with each other contact BioTek s Technical Assistance Center See page 6 Synergy HT Operator s Manual 28 Chapter 2 Installation 13 Run a System Test Running a System Test will confirm that the reader is set up and running properly or will provide an error code if a problem has been detected Perform these steps to run the test 1 Gen5 Select System Diagnostics Run System Test If prompted to select a reader select the Sy
138. er into each well on top of the green dye solution If the reader does not support shaking shake the plate in an orbital shaker for about 15 seconds Transfer the plate to the absorbance reader and run the two Read protocols as described under Test Setup KC4 on page 105 Save each Data file using an identifying file name Data Save As When all tests are complete prime both dispensers with at least 5000 uL of deionized water to flush out the green dye solution BioTek Instruments Inc Dispense Module Tests 109 Results Analysis For your convenience we ve included a worksheet at the end of this chapter for recording the dispense weights Delta OD values calcula tions and pass fail Make two copies of this worksheet one for each dispenser tested The pass fail criteria for each set of 32 wells with the same dispense volume is based on the calculated coefficient of variation CV and Accuracy Error e If you created the Gen5 protocols as described on pages 110 113 the CVs are calculated automatically With the experiment open click the Statistics button and set the Well Type to Assay Control Use the Data drop down box to select each Delta OD data set e If you created the KC4 Read protocols as described on page 117 the CV is calculated automatically With the Data file open select the Delta OD data set and click the Statistics button For each volume dispensed 80 20 5 uL for each dispen
139. erformed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 3 5V Flash Power Supply reference voltage is out of range 1399 1453 See self test Voltage Reference Min This tests the voltage across a sense resistor in series of the 3 5 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 4 25V Flash Power Supply reference voltage is out of range 1699 1772 See self test Voltage Reference Low This tests the voltage across a sense resistor in series of the 4 25 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 5 25V Flash Power Supply reference voltage is out of range 2110 2211 See self test Voltage Reference High This tests the voltage across a sense resistor in series of the 5 25 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB BioTek Instruments Inc Error Codes 195 Code Description and Probable Causes 2907 6 00V Flash Power Supply reference voltage is out of range 2411
140. example using Microsoft Excel gt Ina spreadsheet create two columns labeled X and Y Reference Table 2 on the previous page enter the actual absorbance values in column X and enter the expected absorbance values in column Y gt Select Tools Data Analysis Regression Identify column X as the Input X Range and column Y as the Input Y Range gt Click OK to perform the analysis the results of which will be output in a separate sheet gt Note If the Data Analysis command is not available on the Tools menu you may need to install the Analysis ToolPak in Excel Consult Excel s help system for assistance Since it is somewhat difficult to achieve high pipetting accuracy when conducting linear dilutions an R Square value of at least 0 99 is considered adequate BioTek Instruments Inc Absorbance Liquid Tests 85 Data Reduction for Repeatability 5 Calculate the mean and standard deviation for the five readings taken above at each concentration Only one row of data needs to be analyzed 6 For each mean below 2 000 OD calculate the allowed deviation using the repeatability specification for a 96 well plate of 1 0 0 005 OD If above 2 000 OD apply the 3 0 0 005 specification 7 The standard deviation for each set of readings should be less than the allowed deviation Example Absorbance readings of 1 950 1 948 1 955 1 952 and 1 950 will result in a mean of 1 951 and a
141. filters placed in the filter wheels are not in the optimum order to prevent this error e The filters defined in the filter table do not match the filter wheels e The user did not send the filter table information to the reader after making changes to the table in the controlling software BioTek Instruments Inc Code 2D03 2D04 2D0D 2D0E 2D16 2D28 2E01 Error Codes 199 Description and Probable Causes EX or EM filters not adjacent for multi filter set reads This error indicates either the EX or the EM filter wheel is not set up correctly to have adjacent filters for a multi filter set read Note Only 2 filter sets can be sent when developing software Probable Causes e The filters placed in the filter wheels are not in the optimum order to prevent this error e The filters defined in the filter table do not match the filter wheels e The user did not send the filter table information to the reader after making changes to the table in the controlling software Filter plug s not positioned next to EX filter s for light shutter feature during a Dispense well read This error indicates that the EX filter wheel is not set up correctly for the light shutter feature Probable Causes e The filters placed in the filter wheels are not in the optimum order to prevent this error e The filters defined in the filter table do not match the filter wheels e The user did not send the filter table information to the reader a
142. fter making changes to the table in the controlling software Well spacing not valid for dispensing Well spacing in any direction is lt 0 320 or lt 8 13 mm Probable Causes e A384 well plate was selected for a Dispense protocol During a dispense read the reader missed the start of the plate read This error indicates that during a dispense read the reader was not able to start the read Probable Causes e The processor was busy running other tasks e A previous task took longer than estimated Missed start of read event This error indicates that the up counting plate mode timer is already past event start time or the well mode timer is already past start time for the current read event Probable Causes e Basecode 2 10 or lower used two timers to perform this function Bascode 2 12 or higher uses one timer Dispenser module not attached Dispenser is not attached or has lost initialization due to an intermittent connection Probable Causes e Dispenser cable lost connection Top Probe Height Z Axis did not calibrate during the Absorbance Cal This error indicates old basecode Recommend upgrading This error indicates that the probe Z position lt 24 full steps indicating there is not enough margin available to calibrate the probe Z position The distance between the top of the autocalibration jig and the homing opto sensor is lt 24 full steps Probable Causes e Linear way is dirty or needs lubrication e Homing opt
143. g Tips on page 78 Synergy HT Operator s Manual 76 Chapter 4 Instrument Qualification Reader Basecode Date and Time Absorbance Plate User Comments Well C6 Reference 586 Tolerance 3 Read 587 Result PASS Alignment Results Wells Al Read 0 001 Tolerance 0 015 Result PASS Wavelength 405 nm Accuracy Results Wells CL Reference 0 147 Min Limit 0 124 Max Limit 0 170 Read 1 0 144 Result PASS Last Plate Certification Next Plate Certification Due Absorbance Test Plate Results Synergy Serial Number 128787 P N 7090202 v2 24 08 08 2008 03 03 36 PM Peak Absorbance Results Repeatability Results Wells CL Read 1 0 144 Min Limit 0 138 Max Limit 0 150 Read 2 0 144 Result PASS Wavelength Accuracy Results Wells Cal Reference 0 136 Min Limit 0 113 Max Limit 0 159 Read 1 0 134 Result PASS 630 nm 7 Filter Test Plate P N 7260522 S N 161259 January 2008 January 2009 Administrator Test performed during Initial OQ A12 H1 H12 0 002 0 001 0 002 0 015 0 015 0 015 PASS PASS PASS E G3 H6 F5 D4 0 618 be 133 1 701 2 279 2 945 0 586 1 090 1 647 2 168 2 807 0 650 1 176 TESS 2 390 3 083 0 615 1 128 1 696 2 284 2 908 PASS PASS PASS PASS PASS E G3 H6 F5 D4 0 615 1 128 1 696 2 284 2 908 0 604 dead 1 674 2 210 2 816 0 626 1 144 1 718 2 358 3 000 0 615 1 128 1 695 262189 2 903 PASS PASS PASS PASS PASS E G3 H6 F5 D4 0 568 1 040 1 560 1 865 2 400
144. g enough light energy or is blocking the light e Damaged optic spray Measurement channel correction outside limits See criteria in text below See Delta Air Dark on self test This error can indicate one of the following error scenarios e During a system test the Delta was lt 8000 for one of the wavelengths e During a spectral scan or blank read the flash on value minus flash off was lt 8000 normal mode or lt 8000 calibrated resets 8 for sweep mode Probable Causes e Lamp alignment or lamp power supply failure e Absorbance channel analog PCB e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path and white light is passing through or the filter is degraded and is not passing enough light energy or is blocking the light e Damaged optic spray Necessary configuration data missing This error code indicates old basecode Recommend upgrading Probable Causes e The flash memory on the 7080400 PCB is defective or corrupt The basecode software and or assays may need to be re downloaded Failed configuration checksum test This error indicates that during self test or at the end of a plate read the checksums calculated for configuration flash memory page 0 do not match the saved checksums Probable Causes e The flash memory on the 7080400 PCB is defective or corrupt The basecode software may need to be re downloaded BioTek Instruments Inc Error Codes 187
145. gimen is the best way to ensure accurate performance and a long life for your instrument and dispense module BioTek also recommends flushing the module with DI water before conducting the decontamination procedure described in Appendix A Schedule The following charts recommend Preventive Maintenance tasks and the frequency with which each task should be performed 7 Itis important to note that the risk and performance factors associated with your assays may require that some or all of the procedures be performed more frequently than presented in the schedule BioTek Instruments Inc Recommended Maintenance Schedule 125 Tasks for Synergy HT Models without I njectors Quarterly As Needed Inspect Clean Excitation 128 z and Emission Filters Decontamination see Appendix A Before shipment or storage Tasks for Synergy HT Models with Injectors Clean Exposed Surfaces Inspect Clean Excitation and Emission Filters Flush Purge the Fluid Path Optional Run Dispense Protocol Empty Clean Tip Prime Trough Clean Priming Plate Clean Internal Dispense Tubing and Injector Heads Decontamination see Appendix A Before shipment or storage Synergy HT Operator s Manual 126 Chapter 5 Preventive Maintenance Warnings amp Precautions Please read the following before performing any maintenance procedures Warning Internal Voltage Turn off and unplug the instrument for all maintenance and re
146. gy HT Operator s Manual 188 Appendix C Error Codes Code 1501 150F Incubator zone failed temperature range Description and Probable Causes This error indicates that one or more of the incubator zones failed to maintain their temperature The temperature could be too high or too low After turning on the incubator wait at least 10 minutes for it to stabilize Zone encoding is as follows zone 1 1 2 2 3 4 4 8 see table below for more information Zone Location 1 Top Right 2 Top Left 3 Bottom Right 4 Bottom Left Error code Zone 1501 Zone 1 1502 Zone 2 1503 Zone 1 and 2 1504 Zone 3 1505 Zone 1 and 3 1506 Zone 2 and 3 1507 Zone 1 2 and 3 1508 Zone 4 1509 Zone 1 and 4 150A Zone 2 and 4 150B Zone 1 2 and 4 150C Zone 3 and 4 150D Zone 1 3 and 4 150E Zone 2 3 and 4 150F Zone 1 2 3 and 4 Probable Causes e Thermistor is defective for that zone e Heating pad is defective e Motor power supply PCB is defective e Incubation chamber is less than 12 C When facing the front of the unit BioTek Instruments Inc Error Codes 189 Code 1511 151F Description and Probable Causes Incubator Thermistor failed This error indicates that one or more of the incubator zone thermistors are defective After turning on the incubator wait at least 10 minutes for it to stabilize Zone encoding is
147. he dispensers unless instructed to do so as part of installation upgrade or maintenance Synergy HT Operator s Manual 52 Chapter 3 Getting Started KC4 Software BioTek s KC4 software supports all Synergy HT reader models Use KC4 to control the reader and the dispense module perform data reduction and analysis on the measurement values print and export results and more This section provides brief instructions for creating protocols and reading plates It also explains how to use KC4 to perform some functions that are specific to the dispense module Viewing Updating the Filter and Wavelengths Tables The Synergy HT ships with a set of Excitation and Emission filters installed and the reader s onboard software is pre configured with the filter values and their locations When KC4 first establishes communication with the reader it asks for this information and then stores it in a filter table on the computer This table can be viewed in KC4 by selecting System Reader and clicking the Filters Wavelengths button Click the Fluorescence Luminescence tab to view the Excitation and Emission filter wheel information retrieved from the Synergy Wavelengths x p Reader i Regarding the Synergy HTTR w Iniectors on COM2 Filter wheel position Absorbance numbers 1 to 4 Wavelengths table Absorbance Fluorescence Luminescen The Synergy HT performs absorbance reads in the range of 200 to 999 nm
148. he optic spray is damaged or defective giving a noise reading of zero e Absorbance channel analog PCB is defective 0900 The absorbance reference channel Dark current value failed See Dark on the system test See criteria in text below This error can indicate one of the following scenarios e The reference channel failed during a read or spectral scan for one of the following reasons Dark value was lt 100 during a spectral scan using 8 flashes and 8 resets during sweep mode Dark value was lt 100 during a spectral scan using 8 flashes and the number of resets saved for that wavelength during normal or rapid mode e The reference channel failed lt 100 or gt 20000 during filter calibration or spectral scan with the flash on e The reference channel failed lt 100 during filter calibration or spectral scan with the flash off Probable Causes If failed lt 100 e Absorbance analog PCB or reference channel analog PCB is defective e Shielding of the cable between reference channel and analog PCB is defective or disconnected e Reference channel photodetector is defective If failed gt 20000 e Reference channel photodetector is defective e A faulty analog PCB or faulty internal grounding may cause internal electronic noise e There may be an ambient light leak Ensure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The
149. he plate carrier Wipe the instrument s horizontal surface Synergy HT Operator s Manual 150 Chapter 5 Preventive Maintenance 4 If detergent was used wipe the surfaces with a cloth moistened with water 5 Useaclean dry lint free cloth to dry all wet surfaces Reassembling the Components Perform these steps in the order listed to reassemble the components Refer to the page numbers shown for further instructions and photos demonstrating the steps 1 Slidethe microplate carrier back into the instrument p 149 2 Insert the two injector heads into their sockets in the top probe hanger Do not touch the absorbance lens with your fingers Ensure that the injector heads are properly seated in the hanger The knurled plastic should sit flush against the hanger surface as shown below 3 Attach the two internal dispense tubes to the injector heads as shown below Do not overtighten the thumbscrews Continued on the next page BioTek Instruments Inc 10 11 12 13 Cleaning the Internal Components 151 Hereis the top probe hanger ready for reinstallation with injector heads and internal dispense tubes attached Replace the top probe hanger and shoulder screws using the 3 32 Allen wrench p 147 Insert the top optic probe into its socket and replace its holding screw using the 1 8 Allen wrench p 146 Replace the incubator housing and two thumbscrews p 145 and 144
150. he top surface of the plate during the read In Gen5 this option is found in a Read step within a Procedure In KC4 it is in the Reading Parameters dialog Refer to the software documentation for further instructions BioTek Instruments Inc Key Components 37 Lamp Assembly and Filter Wheel Access Excitation filter wheel Emission filter wheel Fluorescence lamp assembly slides in see instructions on page 13 Cartridge for time resolved fluorescence replaces the Excitation filter wheel Figure 20 Accessing the fluorescence lamp assembly and filter wheels e Thefluorescence lamp assembly and the excitation and emission filter wheels are accessible via a hinged door on the front of the instrument To open the door press on its lower left and right corners until the door opens downward A diagram showing the location of the lamp assembly and the orientation of the excitation and emission filter wheels is printed on the inside of the hinged door e For mode s with the Time Resolved Fluorescence feature remove the excitation filter wheel and replace it with the TR cartridge before running a time resolved fluorescence assay See page 41 for more information on the TR cartridge The Synergy HT has two lamps one for standard fluorescence one for absorbance and time resolved fluorescence Standard Fluorescence The 20 watt tungsten halogen lamp s life is rated at an average of 1000 ho
151. he two injectors located inside the instrument Both injectors are positioned directly above the bottom probe and fluid is injected into one well at a time BioTek Instruments Inc Features 3 Features e Operated using BioTek s Gen5 or KC4 Data Analysis Software e Dual optics design with separate fluorescence and absorbance channels e 3mm top and 5 mm bottom fluorescence probes standard configuration e Optional 1 5 mm top bottom 3 mm bottom fluorescence probes custom configurations e Optional Time Resolved Fluorescence TRF capability T models e g SIAFRT e Fluorescence A ranges Standard low noise PMT gt Excitation 300 to 650 nm 200 to 700 nm with T models gt Emission 300 to 700 nm Optional red extended PMT gt Excitation 300 to 650 nm 200 to 800 nm with T models gt Emission 300 to 800 nm e Absorbance A range of 200 to 999 nm e Absorbance OD range from 0 000 to 4 000 OD e Low Medium High and Variable plate shaking speeds with adjustable durations e All models read 6 12 24 48 96 and 384 well microplates e Injector models dispense to 6 12 24 48 and 96 well microplates e Operates from 100 to 240 V 10 50 to 60 Hz e One serial COM port 9 pin female connector e One USB port e 4 Zone incubation to 50 C e Optional dual reagent dispensing capability Synergy HT Operator s Manual 4 Chapter 1 Introduction Package Contents
152. heck to see if the lamp is on when the instrument is on The lamp assembly may need to be replaced See page 13 for more information If the lamp is on be sure to note the test type X axis position incorrect during XY Back sensor test This error indicates that the X axis did not find the XY sensor or found the XY sensor outside its limits Perform a self test to verify the error See Back Sensor on the self test results the Delta should be lt 32 to pass If it is gt 16 this is an indicator that the carrier needs to be serviced soon Probable Causes e X axis rail is dirty The nylon slider bushings are worn and causing too much friction or dirt in the roller bearings is causing the bearings to jam e Y axis rail is dirty The bearings are dirty and worn causing too much friction e Defective or broken optical sensor e Defective Motor Controller PCB e Bent opto flag on carrier e Carrier feet have been adjusted to the point where the XY flag can no longer enter the XY sensor e The shipping screw was not installed before moving the unit and the XY sensor was damaged e Fault motor gear attachment set screw loose Y axis position incorrect during middle sensor test This error indicates that the Y axis did not find the middle sensor or found the middle sensor outside its limits If the carrier is not extended the unit compares the position against the XY sensor otherwise the unit assumes the carrier is at the middle sensor and wil
153. heel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Scenario 4 fluorescence autocalibration e The surface of the autocalibration jig is too reflective Return the jig to BioTek fora replacement jig e The carrier did not move far enough to block the light beam Troubleshoot the carrier movement 0501 Light beam saturated light Air measurement channel reading reached 65535 or PMT Relative Fluorescing Units RFU reached FFFF 99999 This error can indicate one of the following scenarios e 1 Prior to a fluorescence read or during a system test the PMT is tested for operation the gain is set at 153 Is the PMT operating properly e 2 The PMT is constantly being checked for an overload condition e 3 During a spectral scan read the measurement channel reached saturation at one or more of the selected wavelengths e 4 Filter 1 in absorbance mode has saturated the measurement channel Probable Causes Scenarios 1 and 2 fluorescence or system test e The PMT is defective e The connector from the PMT base to the analog board is defective The ground shield is not grounding properly e The PMT base is defective e The PMT analog PCB is defective Scenario 3 Spectral scan read e The
154. ic Compatibility Do not use this device in close proximity to sources of strong electromagnetic radiation e g unshielded intentional RF sources as these may interfere with the proper operation Caution Warranty Failure to follow preventive maintenance protocols may void the warranty See Chapter 5 Preventive Maintenance Synergy HT Operator s Manual xvi Preface C Based on the testing described below and information contained herein this instrument bears the CE mark Directive 2004 108 EC Electromagnetic Compatibility Emissions Class A The system has been type tested by an independent accredited testing laboratory and found to meet the requirements of EN 61326 1 for Radiated Emissions and Line Conducted Emissions Verification of compliance was conducted to the limits and methods of the following CISPR 16 1 CISPR 16 2 and EN 55022 This equipment has been designed and tested to CISPR 11 Class A In a domestic environment it may cause radio interference in which case you may need to mitigate the interference Immunity The system has been type tested by an independent accredited testing laboratory and found to meet the requirements of EN 61326 1 for Immunity Verification of compliance was conducted to the limits and methods of the following N 61000 4 2 Electrostatic Discharge N 61000 4 3 Radiated EM Fields N 61000 4 4 Electrical Fast Transient Burst N 61000 4 5 Surge Immunity N 61000 4 6 Co
155. ide of the box Ship the box to BioTek See page 6 for the address BioTek Instruments Inc Chapter 3 Getting Started This chapter describes some of the Synergy HT s key components and provides an introduction to using Gen5 or KC4 to control the instrument Key Components eniin ect nd eee E Rae tweet pee 36 Power Switch Carrier Eject Button Microplate Carrier 36 Lamp Assembly and Filter Wheel ACCESS cccceeneeeeneeennes 37 Excitation and Emission Filter Wheels cceeee eee eae 38 Installing the Time Resolved Fluorescence Cartridge 41 Configuring the System for Luminescence Measurements 42 The External Dispense Module esceeeeeeeeeeeeeeeeee ee 43 Gen5 SoftWare odeien adrianna Dea Sonu bhas sab eee ceeded take tae 46 Viewing Updating the Filter and Wavelengths Tables 46 Creating Protocols and Experiments 47 Controlling the Dispense Module ccceeeeeee eee eeee eee eas 50 KE4 SOWA EL id t 0ts acon Aena A RTE DORNANA vada ake heat 52 Viewing Updating the Filter and Wavelengths Tables 52 Creating PrOtOCOlS una di2545 i Seaceans nen sani tarita UAA PAARA KaRa a 53 Reading Plat S 2 2 cascsh sabe eituen eataa ches EN ETETE E REEDA KAREA 55 Controlling the Dispense Module cceeeeeee eee eeee eee ees 56 Recommendations for Achieving Optimum Performance 59 36 Chapter 3
156. ilter wheel to align the desired filter with the hole in the supporting bracket Place the bracket on a flat surface with the filter wheel facing down Prepare a multi layered cushion of lens paper Using your finger covered with the lens paper gently push against the filter and its C clip retainer until they pop out To replace a filter or plug 1 2 Hold the metal bracket with the filter wheel facing up Properly orient the filter or plug see page 38 and then drop it into the desired filter wheel location Using your fingers squeeze the sides of the C clip filter retainer and then insert it into the top of the hole containing the new filter Cover your finger with several layers of lens paper and then push down on all sides of the C clip until it sits flush against the filter Clean both sides of the filter with lens paper To reinstall a filter wheel 1 2 3 4 5 Ensure that all filters and or plugs are inserted properly see above Slide the filter wheel back into its chamber Replace the thumbscrew Close the front door Turn on the instrument BioTek Instruments Inc Key Components 41 Installing the Time Resolved Fluorescence Cartridge For Synergy HT models that support time resolved fluorescence the TR cartridge must be installed in place of the Excitation filter wheel before a TRF assay can be run The TR cartridge allows light from the xenon flash bulb to be input t
157. ing filter wheel is jammed not aligning the correct bandpass filter with the light path and white light is passing through or the filter is degraded and is not passing enough light energy or is blocking the light e Damaged optic spray OFO1 OF06 Reference channel correction outside limits See criteria in text below See Delta Air Dark on self test This error can indicate one of the following scenarios e During the system test the Delta was lt 500 for one of the wavelengths e During sweep mode only the blanking on air uses one flash to test the light performance A ratio is used to determine the performance of the lamp The ratio used is Reference channel blank for wavelength flash on reference data corrected reference dark offset The ratio will fail if lt 0 5 or gt 2 0 e During a spectral scan or blank read the flash on value minus flash off is lt 500 normal mode or lt 500 calibrated resets 8 for sweep mode Probable Causes e Lamp alignment or lamp power supply failure e Reference channel beam splitter lens cracked e Defective reference channel analog PCB absorbance channel analog PCB or the cable in between e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path and white light is passing through or the filter is degraded and is not passing enough light energy or is blocking the light e Damaged optic spray Synergy HT Operator s Manual 18
158. int free cloth Before placing a plate in the instrument blow the bottom of the plate with an aerosol If the test fails again the optical probe s may need to be cleaned Contact BioTek TAC for instructions e Ifa test fails because one or more wells overranged reduce the Sensitivity value in the protocol by 1 5 counts and re read the plate e If the Corners Test continues to fail the hardware may be misaligned Contact BioTek TAC e Review the instructions under Pipette Map page 94 for SF or 100 for MUB to verify that you correctly prepared the plates e Does the Plate Type setting in the protocol match the plate you used Synergy HT Operator s Manual 94 Chapter 4 Instrument Qualification Pipette Map gt x 2 Seal the plates with foil or store them in black polyethylene bags until use If the base of a plate is touched clean the entire base with alcohol 95 ethanol and then wipe with a lint free cloth Before placing a plate in the instrument blow the bottom of the plate with an aerosol duster For the Corners test Pipette 200 uL of the 3 3 nM SF solution into wells A1 A3 A10 A12 H1 H3 and H10 H12 Optional for plates with black sidewalls Pipette 200 uL of buffer in the wells surrounding the 3 3 nM wells CBUF in the grid below For the Sensitivity Linearity tests Use a multi channel pipette with just four tips installed Pipette 150 uL of buffer into wells C2 F5 NOT
159. ioTek Instruments Inc 8 Install the Dispense Module 21 Figures 10 and 11 Possible locations for the dispense module to the left or on top of the instrument Outlet tubes Syringe valves Figure 12 Initial setup of the dispense module Synergy HT Operator s Manual 22 Chapter 2 Installation 12 13 Remove the two syringes from their protective boxes They are identical and interchangeable Each syringe should already be assembled in one piece but if for some reason there are two separate pieces assemble them now insert the white tip of the syringe plunger into the barrel of the syringe and gently push it all the way into the barrel Install both syringes e Hold thesyringe vertically with the threaded end at the top and the knurled steel end at the bottom e Screw the threaded end of the syringe into the bottom of the syringe valve Finger tighten only e Carefully pull down the knurled steel end of the syringe until it is resting inside the holein the bracket e Passa metal thumbscrew up through this hole and thread it into the bottom of the syringe Hold the syringe from rotating while tightening the thumbscrew Finger tighten only The installed syringes should resemble the following O Hes T s Syringe DA J Pp gt oo a Figure 13 The dispense module with the syringes installed Continued on the next page BioTek Instruments Inc 8 Install the Di
160. ion on creating Gen5 protocols See page 53 for more information on creating KC4 protocols Figure 22 The TR cartridge for time resolved fluorescence assays Synergy HT Operator s Manual 42 Chapter 3 Getting Started Configuring the System for Luminescence Measurements For best results when taking luminescence measurements the Excitation filter wheel should have no empty locations and it should have at least one plug also referred to as a dummy filter installed to prevent light from reaching the samples Remove the Excitation filter wheel See page 39 and examine its contents ensure that there are no empty locations and there is at least one plug installed If your tests require that the light emitted from the samples remain unfiltered the Emission filter wheel should have an empty location in it Remove the Emission filter wheel and examine its contents ensure that there is an empty location If you made any changes to either filter wheel you must update Gen5 s or KC4 s filter table Select PLUG to indicate the presence of a plug and HOLE to indicate an empty location Click Send Values or Filters to download the information to the reader Reader Setup Synergy Com6 x r Reader Probe Size Absorbance Fluorescence Luminescence Dispenser 1 Dispenser 2 Excitation Sara Get Values Bandwidth Filter Center Bandwidth p Excitation Synergy HTTR w lnjecto
161. ionized DI or distilled water and wipe all surfaces that have been cleaned with the bleach solution or alcohol Use a clean dry cloth to dry all wet surfaces Continued on the next page Synergy HT Operator s Manual 158 Appendix A Decontamination 10 Reassemble the instrument as necessary If the dispense module is installed detach the outlet tubes from the rear panel of the instrument If it is not installed attach just the dispense module s communication cable to the instrument Remove the supply bottles and their holders Perform the procedures described below through page 160 to decontaminate the fluid lines in the dispense module the internal tubing and injector heads and the tip priming trough and priming plate Decontaminate the Fluid Lines 1 10 11 12 13 Place a beaker with 20 ml of 0 5 sodium hypochlorite solution or 70 isopropyl alcohol near SYRINGE 1 on the dispense module Place the SYRINGE 1 inlet tube in the beaker If you have not already done so detach the dispense module s outlet tubes from the instrument s rear panel Place the ends of the outlet tubes in an empty beaker and set the beaker on the work surface Launch Gen5 or KC4 select System Reader Control and click the Dispenser tab Select Dispenser 1 enter a Volume of 5000 ul and keep the default dispense Rate Place the priming plate on the carrier it is not used but the reader requires its presence
162. ir sssssssssssssssssssserrrrrerrrerrrers 6 Contacting BioTek for Applications Support cceeeeeeeeeeeeee eee eaeenas 6 Chapter 2 Installation c ccsccsseseeeseeseeneeeeeneeneeneeeeeaseaseasesseseeeeensensense 7 1 Unpack and Inspect the Reader sssssssssssssssssssrsrrsnnrrnrrrrrrrrrrrarrennn 8 2 Remove the Shipping Panel cccecceeeeeee esse esse esse eaeeeeeeeeenaeeeaeeaas 10 3 Remove the Microplate Carrier Shipping SCrew cccceceeeeeeeeeeaeees 12 4 Install the Fluorescence Lamp Assembly ccccceeeeeeeeseeeeeeeeeeeeeeaeees 13 5 Select an Appropriate Location sssssssessrsrrrrrrrrrrrrrrrrrrrsrrssrrsrns 14 6 Connect the Power Supply s ssssssssssrssssessessssnsrnsrnorrnerrnnrennnenennnan 15 7 Unpack and Inspect the Dispense Module ccceceeceeeeeeeeeeeeeeaeeaas 16 8 Install the Dispense Module cccceeeeeeee esse eee eeee eee eneeeneeaeeeeeeeaenes 19 9 Connect the Host Computer ccc ecce cece eee eee eee eee eee neat ene e teen ene eee es 23 10 Install Software on the Host Computer ssssssssssrrrrrrrrrrrsrrssessens 24 11 Turn on the REAdeP cececce cess eeeeeeeeeeeeeseeeeeeaeeeeeeanetanetaneeaeees 24 12 Establish Communication cc ecce cece eee eee e eee eee teen ne ener e es 25 USING GenSan listers sie Gia llde a a A sh heen ea teaver EOE 26 USING KCA a A se r A dasaaenadeivansaieda an tthe atenhseagelcde sessed 26 132 RUD ad Systemi TeSt orprsiris irrisa orai
163. is also considered to be linear Repeatability This test ensures the reader meets its repeatability specification by reading each neutral density filter on the Test Plate twice with the filter in the same location Wavelength Accuracy BioTek s Absorbance Test Plate with the part number 7260522 contains a glass filter in position C6 This filter is used to check the wavelength accuracy of the reader The filter is scanned across a specified wavelength range in 1 nm increments The wavelength of maximum absorbance is compared to the expected peak wavelength supplied on the Test Plate s certificate BioTek Instruments Inc Absorbance Plate Test 71 Test Plate Certificates To run this test on the Synergy HT you ll need BioTek s 7 Filter Absorbance Test Plate PN 7260522 with its accompanying certificates e The Standards Certificate contains standard OD values for the filters at several different wavelengths see the sample below e The Peak Wavelength Certificate contains one or more Peak Wavelength values for the glass filter in position C6 on the plate Each value has a valid test range associated with it For example a Peak Wavelength value may be 586 nm with a test range of 580 to 590 nm or tolerance values of 6 4 This test plate can be used for testing the reproducibility linearity and alignment of your BioTek autoreader The following calibration data has been recorded by a N 1 S T traceable spect
164. is returned in response to a Run Area Scan c command it indicates the scanned point left to right that caused the error BioTek Instruments Inc Appendix D Specifications This appendix contains BioTek s published specifications for the Synergy HT SPECICATION S vasvwoy severe aa e a T EEEREN 208 MICFO DALES ii vies maa bund e T R eid een A h 208 Hardware and Environmental ccccccccceeeeeeeeeeeeuceeeeeeeeneneeuaas 208 ADSORDAN CO sirian beds e due eaa dawn ets a heise aaa 209 Fluorescence ronseis esa cee wannliee a aei dave da be eda a A 211 INGCUBDAUI ON eee Riad a ce teeaue eG Wada aue tee a ail ix a a nae aoa ane 213 Shakire n a sae a a wud sedete ee a a aa aaa KI aA 213 208 Appendix D Specifications Specifications Microplates All models accommodate standard 6 12 24 48 96 and 384 well microplates with 128 x 86 mm geometry and support e Absorbance mode plates up to 0 8 20 30 mm high e Fluorescence Luminescence modes plates up to 1 25 31 75 mm high e PCR tube trays up to 1 25 31 75 mm high may require an adapter All models read 6 12 24 48 96 and 384 well microplates Injector models dispense to 6 12 24 48 and 96 well microplates Hardware and Environmental Light Source Absorbance Xe flash light source 10 W max average power Life 1 billion flashes Fluorescence Tungsten quartz halogen 20 W Life 1000 hours Dimensions 16 Dx 15 W x
165. ispenser e Add a Read step to specify the detection method and filter sets or wavelength values enable time resolved fluorescence and set the Top Probe Vertica Offset value e Todefinea Kinetic read place an Endpoint Read step inside a Kinetic Start End loop Dispenser 1 Dispense Step x Step Label lt default gt Full Plate Detection Method Fluorescence x IT Time Resolved Tip Prime Volume 10 ul Priming prior to dispense 7 Dispense Volume 50 ul Rate 250 x ul sec cm Read Type Endpoint m Filter Sets 4 C41 2 v Excitation fszsjz0 fszsjz0 Emission fessj40 gt feasj40 Optics Position fo xl fo Sensitivity 50 75 Help Top Probe Vertical Offset 1 mm Add Step Read Dispense Description Read 405 630 4 End Kinetic Plate Type ERS Rata Y Start Kinetic Run 0 10 00 Interval 0 01 39 Procedure Synergy Com6 Figure 28 Clockwise from upper left defining a Dispense step defining a Read step defining a Kinetic read Kinetic Endpoint Read steps BioTek Instruments Inc Gen5 Software 49 Open the Plate Layout dialog and assign blanks samples controls and or standards to the plate Open the Data Reduction dialog to add data reduction steps Categories include Transformation Well Analysis Curve Analysis Cutoff
166. ission TR Time Resolved Cartridge see page 41 Filter direction within a filter wheel is important and the direction differs depending on the filter wheel There is a diagram on the inside of the front panel door indicating this Each filter is marked with an arrow indicating the proper direction of light Refer to the figures on the previous page for proper filter orientation To remove a filter wheel 1 2 I mportant Turn off the instrument Using your thumbs push down on the bottom corners of the hinged door on the front of the instrument Observe the two thumbscrews within the compartment The left thumbscrew holds the Excitation filter wheel in place the right secures the Emission filter wheel Remove the thumbscrew and slide the filter wheel s supporting metal bracket straight out of the compartment Note The Emission filter wheel will spring out when removed This is because a shutter behind the wheel closes quickly to protect the PMT Synergy HT Operator s Manual 40 Chapter 3 Getting Started Important When removing or replacing a filter or C clip filter retainer do not usea sharp instrument Use several layers of lens paper and your finger to renove and replace filters and clips Using a sharp instrument such as a flat screwdriver will scratch the filter surface and make it unusable Do not touch the filters with your bare fingers To remove a filter or plug 1 Turn the f
167. l Synergy HT Operator s Manual 138 Chapter 5 Preventive Maintenance Perform these steps to remove both sets of internal dispense tubes and injector heads 1 Open the cable clamp to release the tubes 2 Locate the tubing ports on the reader s rear wall Turn each tube s thumbscrew counterclockwise and gently pull the tube from the port 3 Locate the injector heads Turn each tube s thumbscrew counterclockwise to disconnect the tube from the injector head BioTek Instruments Inc Cleaning the Internal Components 139 4 Turn the injector heads counterclockwise and gently pull them out of their sockets Note Be sure to seat the injector tips securely when reinstalling See the photo on page 150 Synergy HT Operator s Manual 140 Chapter 5 Preventive Maintenance Cleaning the I nternal Tubes and Injector Heads As discussed on page 124 some reagents can crystallize and clog the tubing and injector heads Daily flushing and purging can help to prevent this but more rigorous cleaning may be necessary if reagent has been allowed to dry in the tubing and or injectors To clean the tubes 1 Soak the internal tubes in hot soapy water to soften and dissolve any hardened particles 2 Flush each tube by holding it vertically under a stream of water froma faucet To clean the injector heads 7 Do not remove the o ring from the injector head see photos below
168. l beep and the failure will be indicated on the test report generated via Gen5 or KC4 The report also contains the reader s serial number and the part number and version number of the basecode software When the instrument is turned on the System Test automatically runs but no report is generated To run the test and generate a report you must use Gen5 or KC4 The absorbance measurement system is checked using the six wavelengths specified in the reader s internal absorbance wavelength table Before running the test set these wavelengths to the ones you most frequently use if they are not already set To view modify the wavelength table via Gen5 KC4 see the instructions in the Getting Started chapter Note The System Test runs automatically when the instrument is turned on If this power up System Test fails the instrument will peep repeatedly If this happens press the carrier eject button to stop the beeping and then initiate a System Test through Gen5 or KC4 to retrieve the error code Synergy HT Operator s Manual 66 Chapter 4 Instrument Qualification Procedure To run the System Test 1 2 Turn on the reader and launch Gen5 or KC4 If necessary set Gen5 s or KC4 s wavelength table to the six wavelengths you most frequently use See the Getting Started chapter for instructions If your assays use incubation we recommend enabling Temperature Control and allowing th
169. l gt Dispense to wells A5 H8 Tip prime before this dispense step 20 uL Dispense 20 uL at rate 250 uL sec Suggested comment Weigh the plate 20 ul test RECORD the weight TARE the balance Place the plate back on the carrier Click OK to continue Dispenser lt select 1 or 2 depending on the protocol gt Dispense to wells A9 H12 Tip prime before this dispense step 5 uL Dispense 5 uL at rate 225 uL sec Suggested comment Weigh the plate 5 ul test RECORD the weight PIPETTE 150 ul well of DI water into all 12 columns Place Plate Out In the plate back on the carrier Click OK to perform the Read steps Medium intensity for 15 seconds Detection Method Read Type Read Speed Two Wavelengths Step label Wells Detection Method Read Type Read Speed Two Wavelengths Read Step label 80 ul Read_Disp 1 or _Disp 2 Wells A1 H4 Ea i Absorbance Endpoint Normal 405 and 750 nm 20 and 5 ul Read_Disp 1 or _Disp 2 A5 H12 Absorbance Endpoint Normal 630 and 750 nm BioTek Instruments Inc Dispense Module Tests 113 Customize the Plate Layout Optional The results analysis worksheet at the end of this chapter requires the calculation of the Standard Deviation Mean and CV of the ODs read for each dispense volume in each plate six sets of calculations By identifying the wells by their dispense volumes in the Plate Layout Gen5 will calculate these values for
170. l verify or home the sensor Perform a self test to the verify error See Middle Sensor on the self test results the Delta should be lt 32 to pass If it is gt 16 this is an indicator that the carrier needs to be serviced soon Probable Causes e Y axis rail is dirty The bearings are dirty and worn causing too much friction e Defective or broken optical sensor e Defective Motor Controller PCB e Shipping screw was not installed before moving the unit and the XY sensor was damaged e Something moved the carrier while it was extended e Faulty motor gear attachment set screw loose BioTek Instruments Inc Code 2500 2505 2600 280x 2900 Error Codes 193 Description and Probable Causes X axis went by flash location too soon during sweep mode This error indicates that during sweep mode the unit calculated the exact time for the flash to occur and the flash failed to flash at that time Probable Causes e Defective motor driver PCB e One of the carrier axis motors is defective Monochromator position more than 1 full step past flash point This error indicates that during a spectral scan the monochromator went more than 1 step past the flash point Probable Causes e Monochromator motor is defective e Motor power supply PCB lost or added counts from monochromator motor Physical limit exceeded for area scan request This error can occur only during computer control indicating that the area scan reque
171. ld match the plate you are actually using Protocol Name Synergy HT FI_B prt Parameter Setting Plate Type Greiner SensoPlate Mfr 655892 Read Wells Corners Read step All wells Sensitivity Read step Wells C1 F12 Filters EX 485 20 nm EM 528 20 nm Sensitivity Linearity Read step 100 Delay After Plate Movement Measurements Per Data Point Protocol Name Synergy HT FI_T prt Parameter Setting Plate Type Costar 96 black opaque Mfr 3915 Sensitivity Read step Wells C1 F12 Sensitivity Corners Read step 80 Sensitivity Linearity Read step 100 Synergy HT Operator s Manual 96 Chapter 4 Instrument Qualification KC4 Protocol Reading Parameters The following tables contain the recommended settings for the KC4 protocols Your tests may require modifications to some of these parameters such as Plate Type or Sensitivity see Troubleshooting on page 93 The Plate Type setting should match the plate you are actually using Sensitivity Filter Set 1 Corners 80 Filter Set 2 Sensitivity Linearity 100 Nb Samples Per Well Both filter sets 40 Delay Before Sampling Both filter sets 350 msec Plate Type Greiner SensoPlate Mfr 655892 Note You may need to add this Plate Format in KC4 see the dimensions on the next page Read Wells Sensitivity Filter Set 1 Corners 80 Filter Set 2 Sensitivity Linearity 100 Top Probe Vertical Offset
172. le gm 1D GJO e Power inlet for injector models as shown Power inlet for non injector models e 24 VDC center positive Figure 6 Power inlet on the rear of the instrument BioTek Instruments Inc 7 Unpack and Inspect the Dispense Module 17 7 Unpack and Inspect the Dispense Module This section applies to Synergy HT modds with injectors only Important Save all packaging materials If you need to ship the dispense module to BioT ek for repair or replacement you must use the original materials Using other forms of commercially available packaging or failing to follow the repackaging instructions may void your warranty During the unpacking process inspect the packaging module and accessories for shipping damage If the reader is damaged notify the carrier and your BioTek representative Keep the shipping boxes and the packaging materials for the carrier s inspection BioTek will arrange for repair or replacement of your reader immediately before the shipping rdiated claim is settled Perform these steps to unpack and inspect the dispense module and accessories 1 Open the outer shipping box Remove the foam cap inner shipping box and accessories box Top foam end cap Accessories box Inner shipping box Dispense module Bottom foam end cap Outer dj shipping box Figure 7 Unpacking the dispense module s outer shipping b
173. lect System Reader Control and click the Dispenser tab The Tip Prime Trough value shows KC4 s estimate of how much fluid is in the trough 4 Click Dump all Tip Prime Trough s The Tip Prime Trough value will reset to 1500 uL remaining meaning the trough is empty Note When running a Read amp Dispense protocol KC4 may prompt you to empty the tip prime trough In this case KC4 will automatically open the System Reader Control Dispenser dialog Synergy HT Operator s Manual 58 Chapter 3 Getting Started Syringe Maintenance Position KC4 provides access to special syringe setup functions for maintenance and calibration purposes If a syringe needs to be installed or replaced it must first be moved to its Maintenance Position To do this using KC4 1 InKC4 select System Readers click the Configuration button and then click the appropriate Dispenser tab 1 or 2 2 Click Move Syringe to maintenance position The syringe plunger will move to its furthest from home position The syringe can then be disconnected form the drive bracket and unscrewed from the valve See Install Dispense Module Components in Chapter 2 Installation for information on installing renoving the syringes 3 Click OK to close the dialog Important Do not change the syringe positions or calibrate the dispensers unless instructed to do so as part of installation upgrade or maintenance BioTek Instrum
174. light or positional verify The order sorting filter wheel is homed and moved to the open hole position The monochromator is moved until the optical system detects saturation It is then moved to the point where there is no saturation and then moved back to the saturation point This error is indicating the saturation did not clear or appear Probable Causes e Flash lamp is missing flashes or is not flashing e The optic system does not detect the saturation Analog PCB e Order sorting filter wheel is binding against the motor gear not allowing the open hole to line up correctly e Order sorting filter motor failed due to the bearings e Bearings within the grating mirror are causing the monochromator to jam Z axis Top Probe failed position verify Probable Causes e The optical trigger flag has moved or is loose e The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft Reference Field Change Notice L0031 e Fiber optic cable is not tied to the upper probe assembly and has moved into the top probe s path not allowing the top probe to reach the optical sensor Synergy HT Operator s Manual 174 Appendix C Error Codes Code Description and Probable Causes 0407 Probe Changer failed position verify Probable Causes The optical trigger flag has moved or is loose The probe changer assembly is not allowed to move due to a foreign object preventing its movement 0408 Syringe
175. light shutter feature INVALID MODE O05 invalid processing mode received DISP READ DEF NOT SET 06 dispense read protocol parameters not set or not valid FEATURE NOT AVAILABLE POT disp read feature not enabled by dip switch INVALID EX WAVE MOB excitation wavelengths must be the same for TR assay TIP PRIME TUB VOL OO total tip prime vols in assay larger than tub capacity NUM EVENTS OB total number of events defined invalid INVALID EVENT MGM invalid event type received INVALID GEOMETRY OD wells spaced too close for dispense operation PLATE START LATE OE plate mode event missed its scheduled start time INVALID TIP PRIME OF tip prime conflicts with tip prime rules NUM READ EVENTS LO number of read events defined invalid READ START TIME MLS read event start time out of range NUM SAMPLES MLD amp number of samples out of range SAMPLING INTERVAL PLESA sampling interval out of range NUM PLATE READS 14 number of plate mode kinetic reads out of range PLATE MODE READ TIME POA plate mode kinetic interval time too short READ START LATE LEN read event missed its scheduled start time NUM DISP EVENTS 2i number of dispense events defined invalid TIP PRIME VOLUME TDD tip prime volume out of range DISP VOLUME TO volume requested out of range DISP RATE 24 dispense rate requested out of range DISP START TIME eg dispense start time kinetic interval out of range DISP INTERVAL 26r dispense kinetic
176. lly if smaller volumes of liquid are used For best results use at least 100 ul per well in a 96 well plate and 25 uL in a 384 well plate e Dispensing solution into 384 well plates often traps air bubbles in the wells which may result in inaccurate readings A dual wavelength reading method usually eliminates these inaccuracies however for best results remove the air bubbles by degassing the plate in a vacuum chamber before reading e Theinclination of the meniscus can cause loss of accuracy in some solutions especially with small volumes A gitate the microplate before reading to help bring this problem within acceptable limits Use Tween 20 if possible or some other wetting agent to normalize the meniscus for absorbance measurements Some solutions develop menisci over a period of several minutes This effect varies with the brand of microplate and the solution composition As the center of the meniscus drops and shortens the light path the density readings change The meniscus shape will stabilize over time Synergy HT Operator s Manual 60 Chapter 3 Getting Started e To keep the dispense system in top condition flush and purge the fluid lines with deionized DI water every day or upon completion of an assay run whichever is more frequent Some reagents may crystallize or harden after use clogging the fluid passageways Flushing the tubing at the end of each day letting the DI water soak them overnight and then purging
177. ls to the syringes These tubes are short pieces of opaque PTFE Teflon tubing connected to stainless steel probes on one end and threaded fittings on the other end Three way valves switch the syringe flow from the inlet tubes to the outlet tubes 4 Outlet tubes transport fluid from the syringes into the instrument through the tubing ports on the Synergy HT s rear panel The outlet tubes are opaque PTFE tubes with threaded fittings on each end that are used to deliver fluid from the syringes to the instrument OO Synergy HT Operator s Manual 44 Chapter 3 Getting Started Inside the Synergy HT two Teflon tubes transport fluid from the tubing ports on the rear of the instrument to the two injectors As shown below both injectors are positioned directly above the bottom fluorescence optical probe Syringe 1 Syringe 2 Bottom probe Figure 24 Close up view of the injectors inside the instrument Note The tubing and injectors should be cleaned at least quarterly See Chapter 5 Preventive Maintenance for more information BioTek Instruments Inc Key Components 45 Priming the System Before an assay requiring fluid dispense is run the system should be fully primed with the reagent or other fluid used by the assay At the start of the assay and optionally at the start of each dispense to a well an additional injector tip prime can be performed The tip prime compensates for any fluid loss at
178. mation BioTek Instruments Inc System Test 67 Gen5 System Test Report Reader Synergy Serial Number 128787 Basecode P N 7090202 v2 24 Date and Time 08 08 2008 10 12 58 AM User Administrator Company BioTek Comments System Test run during the IQ Test Results Operator ID Notes SYSTEM SELF TEST 7090202 Version 2 23 128787 1111 1110 Bias current offset 0 9 counts PASS Offset voltage 1541 counts PASS 750V measurement 90 1 counts PASS 750V noise 66 counts 500V measurement 4 6 counts 500V noise 3 counts Lambda 200 Gain Resets Channel Ref Air 14235 Dark 9869 Delta 4366 Lambda Gain Channel Ref Air 12954 Dark 9871 Delta 3083 Lambda Gain Resets Channel Ref Air 12755 Dark 9871 Delta 2884 Lambda Gain Resets Channel Ref Air T2783 Dark 9869 Delta 2914 Lambda Gain Resets Channel Ref Air T2705 Dark 9868 Delta 2837 Lambda Gain Channel Ref Air 13084 Dark 9865 Delta 3219 Figure 30 1 Sample output for the System Test Sheet 1 of 3 The format varies depending on the software used Synergy HT Operator s Manual 68 Chapter 4 Instrument Qualification Channel Ref Noise Max 9866 Noise Min 9865 Delta 1 Voltage Reference Mtr Min Low High Max TR 2045 1425 1734 2157 2465 3333 INCUBATOR SELF TEST Temperature Setpoint 7 Current Average A D Test PASS Zone 1 37 Min 36 S ae Range Thermistor PASS Zone 2 37
179. ming as well Endpoint read time is from plate start to plate stop Kinetic read time is from A1 to A1 read positions Single Dual Endpoint 96 well plate 630 nm 630 450 nm Normal Read Mode 57 sec 106 sec Rapid Read Mode 47 sec 86 sec Sweep Read Mode 23 sec 37 sec Single Dual Endpoint 384 well plate 630 nm 630 450 nm Normal Read Mode 159 sec 309 sec Rapid Read Mode 121 sec 232 sec Sweep Read Mode 38 sec 65 sec Single Kinetic 96 well plate 630 nm Normal Read Mode 49 sec Rapid Read Mode 39 sec Sweep Read Mode 14 sec BioTek Instruments Inc Specifications 211 Single Kinetic 384 well plate 630 nm Normal Read Mode 150 sec Rapid Read Mode 111 sec Sweep Read Mode 26 sec Fluorescence Read Timing Because of the possible wide variations in setup the following benchmark conditions are specified Excitation Filter 485 20 nm Emission Filter 528 20 nm Samples per well 10 Delay before sampling 350 ms Delay between samples 1ms 96 well read 89 sec 384 well read 275 sec Optical Probes The Synergy HT is configured with a variety of probe sizes 1 5 and 3 mm probes can be installed in either the top or bottom positions the 5 mm probe can only be installed in the bottom position Sensitivity 5 mm probe Sodium Fluorescein SF Bottom reading 10 pg ml solution of Sodium Fluorescein in PBS 150 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 530 25 Hellma 96 well quartz
180. n of 32 absorbance readings is calculated Pass Fail criteria depends on the per well volume dispensed 2 0 for 80 uL 7 0 for 20 uL and 10 0 for 5 uL The plate is read in an absorbance reader at 405 750 nm for columns 1 4 and at 630 750 nm for columns 5 12 The two tests are performed simultaneously and use the same plate Data reduction can be performed automatically or manually depending on the test setup BioTek Instruments Inc Dispense Module Tests 103 Required Materials Absorbance reader with 405 630 and 750 nm filters The reader must have an accuracy specification of 1 0 0 010 OD or better and a repeatability specification of 1 0 0 005 OD or better Note The Synergy HT reader may be used if it has passed the Absorbance Plate Test and the Absorbance Liquid Tests described earlier in this chapter Shaker if the absorbance reader does not support shaking Precision balance with capacity of 100 g minimum and readability of 0 001 g 50 200 ul hand pipette and disposable tips Deionized water Supply bottles 250 mL beaker New 96 well flat bottom microplates BioTek s Green Test Dye Solution PN 7773003 undiluted or one of the alternate test solutions provided on the next page 100 mL graduated cylinder and 10 mL pipettes if not using BioTek s Green Test Dye Solution Gen5 or KC4 software installed on the host PC Calculation worksheet on the last page of this chapter Syne
181. n ascending order Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Values entered are corrupted or incorrect e Incorrect calibration of the syringe Default calibration data set to do not use This error indicates that the default calibration data flag is set Syringe needs to be calibrated Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Factory default data is set the syringe needs to be calibrated before use or calibration values need to be entered from the syringe drive e Values entered are corrupted or incorrect During a dispense read the reader missed the start of the well read This error indicates that during a dispense read the reader was not able to start the read for a particular well Probable Causes e The processor was busy running other tasks e A previous task took longer than estimated Filter bandpass overlap in intermediate switching position for multi filter set reads This error indicates that the software cannot move either the EX or EM filter wheel without causing the filters to overlap and saturate the PMT Note Only 2 filter sets can be sent when developing software Equation used for overlapping If EM wavelength is gt EX if EX wavelength 42 bandpass gt EM wavelength 1 2 bandpass then fail Else if EX wavelength is gt EM if EM wavelength 1 2 bandpass gt EX wavelength 12 bandpass then fail Probable Causes e The
182. nce IQ OQ PQ package PN 7090521 p 5 2 Updated liquid test procedures Added new Chapter 6 Maintenance and Troubleshooting which includes Sample reproducible page from maintenance logbook Procedures for maintenance and routine cleaning Instructions for changing injector positions Updated decontamination procedure Appendix A Modified Appendix B Computer Control Corrected Appendix C Error Codes Changed Dispenser Module to Dispense Module throughout E 02 2004 Chapter 1 Introduction Removed reference to NB version p 1 3 Added specifications to reflect the use of an additional PMT type R4220PHA to Hardware Features p 1 3 Moved filter plug 7082073 from Optional Accessories list to Package Contents p 1 4 Added required specifications for microplates used in Luminescence mode BioTek Instruments Revision History ix Rev Date Changes E p 1 5 Added 6 to 96 well plates and fluorescence and luminescence read modes to Injector Model features p 1 10 Removed option from reference to Incubation specifications p 1 11 Chapter 3 Installation Removed references to NB version from description of reader model options Setting Communication Parameters in KC4 p 3 12 Chapter 5 Performance Verification Qualification Tests Updated Figure 5 1 Sample System Test p 5 6 and 5 7 Updated Appendix C Error Codes 07 2004
183. nducted Disturbances N 61000 4 11 Voltage Dips Short Interruptions and Variations m o m m a Directive 73 23 EEC Low Voltage Safety The system has been type tested by an independent testing laboratory and was found to meet the requirements of EC Directive 73 23 EEC for Low Voltage Verification of compliance was conducted to the limits and methods of the following EN 61010 1 Safety requirement for electrical equipment for measurement control and laboratory use Part 1 General requirements Directive 2002 96 EC Waste Electrical and Electronic Equipment Disposal Notice This instrument contains printed circuit boards and wiring with lead solder Dispose of the instrument according to Directive 2002 96 EC on waste electrical and electronic equipment WEEE or local ordinances Directive 98 79 EC In Vitro Diagnostics if labeled for this use e Product registration with competent authorities e Traceability to the U S National Institute of Standards and Technology NIST Optical density measurementsare traceable to NIST BioTek Instruments Electromagnetic Interference and Susceptibility xvii Electromagnetic Interference and Susceptibility USA FCC CLASS A Warning Changes or modifications to this unit not expressly approved by the manufacturer could void the user s authority to operate the equipment This equipment has been tested and found to comply with the limits for a Class A digital devi
184. nergy HT and click OK KC4 Select System Diagnostics Run Optics Test When the Run Optics Test dialog appears click Start When the test is complete a dialog will appear to request additional information Enter the information if desired and click OK The results report will appear Scroll down toward the bottom the text should read SYSTEM TEST PASS e You may wish to print the report and store it with your Installation records e Thesoftware stores system test information in its database you can retrieve it at any time If an error code is returned turn to Appendix C Error Codes and look up the code If the problem is something you can fix do so now and run another System Test If the problem is something you cannot fix or if the test continues to fail contact BioTek s Technical Assistance Center See page 6 for contact information Models with injectors Keep the software open and proceed to 14 Test Injector System All other models The installation and setup process is complete Close the software and turn to page 31 to read about Product Registration and Operational Performance Qualification BioTek Instruments Inc 14 Test the Injector System 29 14 Test the Injector System This section applies to Synergy HT modds with injectors only Perform these steps to test the injector system 1 If necessary press the button above the power switch to eject the microplate carrier Place
185. ng Updating the Filter and Wavelengths Tables 0005 52 Creating Protocols tic ccdicsecatrces teens eaten ons civ N A pace deel sawedee 53 Reading Plate S shiici cases se daavansnevegsaaccarnata a dedi saweegs Deca AARDE ENS 55 Controlling the Dispense MOdUle ceceeeeeee eee ee eee eee eee eae eaeeaeaes 56 Recommendations for Achieving Optimum Performance eeee ee 59 Chapter 4 Instrument Qualification s sssss15225 5 61 OVERVIEW ev deveined neice eeu weed aN alee aad ears 62 TQ OO PQ MEA A A ea shaeadiuetiexhe na detesntuneneciser eee 62 Recommended Qualification Schedule oo ceeccecee eee e eee eeeeeee eee eeeeaeenas 64 System TESE siete aN sata ie nda Rees a EA naa evans vende 65 DESCrIPElON Se sieves cea teehee whet Say woe erste sata taaeva vines Taw RNA 65 Procedure orgias randi acts oes a e a inched a a Hees iendy ot Aana EAEn 66 Absorbance Plate Testerna enean gaip A AA RANEE A A EAA TENERTE AAEN AEA AE ia 70 DESCrIPUlOM inent cee ra a AARAA N Yeeweaetee E ANANE AANER 70 Test Plate Certificates ssssserssrrsrrsrrssnsenaensensennernsrrorrnernenrenran 71 SABE ne AAT cian keieaceleemencaey T dhean Beinn xeiltes ahornn bes 72 ProGeduirres GENS vise cachetdcals s cash a vidaites cubase Da E Aa Goats DERA AAE EEE En 73 Setup KC4 tena itive rid sdaeane REE AE ova leads Paw ea eine aN 74 Procedure KC4in nt a tea ince Cea e A etre dia Baines aaa noi aveue te
186. nitialization If the dispense module was connected to the reader before the reader was turned on or if a System Test was run via Gen5 the dispense module should initialize automatically If for any reason the module does not initialize automatically you can initialize it from Gen5 1 In Gen5 select System Reader Control Synergy Com lt gt and click the Dispenser tab Select the desired Dispenser number 1 or 2 and click the I nitialize button The syringe drive will move to its home position and its sensors will be verified Upon successful completion the I nitialized field should show Yes Prime Utility Before running an experiment with a Dispense step the dispense module and its associated tubing must be primed with the fluid to be used Gen5 provides a special utility for this task To prime the dispense module 1 Fill the supply bottle with a sufficient volume of the fluid to be used for the prime and the assay Insert the appropriate inlet tube into the bottle Important Place the priming plate on the carrier In Gen5 select System Reader Control Synergy Com lt gt and click the Dispenser tab Select the Dispenser number 1 or 2 associated with the supply bottle Enter the Volume to be used for the prime from 5 to 5000 uL The minimum recommended prime volume is 1100 UL Select a prime Rate in uL second Click Prime to start the process When the process is complete carefully renove the
187. nto a weigh boat 2 Rinse the contents into a 1 liter volumetric flask 3 Add 0 5 mL of Tween 20 or 5 mL of BioTek s wetting agent 4 Make up to 1 liter with DI water cap and shake well Solution B Required Materials e BioTek QC Check Solution No 1 PN 7120779 25 mL or PN 7120782 125 mL e Deionized water e 5 mL Class A volumetric pipette e 100 mL volumetric flask Procedure 1 Pipette a 5 mL aliquot of BioTek QC Check Solution No 1 into a 100 mL volumetric flask 2 Make up to 100 mL with DI water cap and shake well Synergy HT Operator s Manual 82 Chapter 4 Instrument Qualification Liquid Test 1 This procedure confirms repeatability and alignment and will reveal any problems with the reader s optics A 96 well flat bottom microplate is required for this test Corning Costar 3590 is recommended Be sure to use a new microplate because fingerprints or scratches may cause variations in readings 1 Using freshly prepared stock solution Solution A or B on the previous page prepare a 1 2 dilution using deionized water one part stock one part deionized water the resulting solution is a 1 2 dilution Pipette 200 uL of the concentrated solution into the first column of wells in the microplate Pipette 200 uL of the diluted solution into the second column of wells After pipetting the diluted test solution into the microplate and before reading the plate we strongly recommend
188. o sensor is defective Synergy HT Operator s Manual 200 Appendix C Error Codes Code 2E02 2E03 3000 3001 3100 3201 3202 3203 3204 3306 Description and Probable Causes Probe Z calibration failure This error indicates that the probe Z position is lt 24 full steps indicating there is not enough margin available to calibrate the probe Z position The distance between the top of the autocalibration jig and the homing opto sensor is lt 24 full steps Probable Causes e Linear way is dirty or needs lubrication e Homing opto sensor is defective Incubator This error indicates old basecode Recommend upgrading Time Resolve filter block is not installed when a TR function is requested Time Resolve filter block in the Excitation filter slot when a non TR fluorescence dispense read is selected Probable Causes e The Time Resolved cartridge is not installed in the EX position e The Hall Effect sensor is defective e Instrument does not have the TR functionality and the DIP switch setting 4 on the 7080400 PCB is open and the DIP switch setting 1 is open on the 7090410 PCB Plate read took longer than kinetic interval Probable Causes e User defined interval is incorrect No absorbance A D ready transition No fluorescence A D ready transition No incubation A D ready transition No voltage reference channel A D ready transition Required carrier outside when locked in If the carrier is inside the read
189. o the fluorescence optical system within the Synergy instrument Excitation wavelengths are selected by adjusting the monochromator from 200 to 999 nm in 1 nm increments with a fixed bandwidth of 10 nm The Synergy HT automatically detects the presence of the TR cartridge At the start of a time resolved fluorescence assay the operator will be prompted to install the TR cartridge if it is missing To install the TR cartridge 1 Important Turn off the instrument 2 Using your thumbs push down on the bottom corners of the hinged door on the front of the instrument Observe the two thumbscrews within the compartment The left thumbscrew holds the Excitation filter wheel in place See the figure on page 37 3 Remove the left thumbscrew and slide the filter wheel s supporting metal bracket straight out of the compartment 4 SlidetheTR cartridge into the compartment and replace the thumbscrew Close the front door and turn on the instrument To specify time resolved fluorescence in a Gen5 protocol check the Time Resolved box in a Read step in the procedure To specify time resolved fluorescence in a KC4 protocol check the Time Resolved box in the Reading parameters dialog When defining a filter set using either software package click the Options button to specify the length of time to delay before collecting readings and the length of time for which readings will be taken See page 47 for more informat
190. ock in excitation slot lt TR block in gt plate read took longer than kinetic interval lt device gt never saw A D ready transition lt motor gt required carrier outside when locked in overload at lt well gt A01 P247 Test Type Codes lt lowest digit in returned error code gt FAIL VTEST HI FAIL VTEST LO FAIL WELL TEST FAIL BKGRND TEST 17 WO 57 Ng higher PMT voltage level incorrect lower PMT voltage level incorrect PMT saturation at well PMT saturation during background overload test Vref Test Channel Codes lt lowest digit in returned error code gt TEST LAMP EST 24V q ie n q MOTOR Bi ST XF MIN a ie n 5 l XF LOW ti ST XF HIGH TEST XF MAX EST XF HTRF 17 vo W397 47 vga vg 77 Ng lamp current test 24V power drive test motor drive test xenon flash min power test xenon flash low power test xenon flash high power test xenon flash max power test xenon flash TR power test Motor Codes lt lowest digit in returned error code gt Carrier X Axis Carrier Y Axis Excitation Filter Wheel Emission Filter Wheel Monochromator Filter Wheel Monochromator Probe Height Axis Probe Changer Dispenser Syringe 1 Dispenser Syringe 2 QO 7 vo gy 47 57 g 77 Ng Ng Synergy HT Operator s Manual 204 Appendix C Error Codes Incubator Codes
191. onnection between the Dispenser and Synergy is too long or has intermittently been lost e The syringe valve did not open e While moving in the negative direction the syringe opto sensor triggered off then on indicating that the syringe went past the opto sensor Syringe on sensor when it should be off This error indicates that the current position gt 1050 1 16 steps and the syringe is on the opto sensor when it should be off Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Linear way is dirty or needs lubrication e Defective or broken optical sensor e Syringe was not installed correctly or was not cleaned causing stress on syringe movement by not allowing the syringe to move to the home position e The syringe valve did not open e The glue between the lead screw and the motor has separated BioTek Instruments Inc Code 2B03 2B04 2B05 2B0A 2C01 2C02 Error Codes 197 Description and Probable Causes Opto sensor clear count obtained This error indicates that during syringe initialization the syringe motor tried to move off the opto sensor and moved gt 1050 1 16 steps and did not see the opto transition Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Syringe was not installed correctly or was not cleaned causing stress on syringe movement by not allowing the syringe to move off the home position e Linear way is dirty or needs lu
192. opsy 0 999 1 509 1 808 2 284 0 599 1 081 1 611 1 922 2 516 0 566 1 037 1 557 1 866 23385 PASS PASS PASS PASS PASS Figure 32 1 Sample output for the Absorbance Plate Test page 1 of 2 Note The format varies depending on the software used to run the test BioTek Instruments Inc Absorbance Plate Test 77 Repeatability Results Product support center Wells GL E G3 H6 F5 D4 Read 1 0 134 0 566 12037 L557 1 866 2 385 Min Limit 0 128 0 555 1 022 T530 1 842 2 308 Max Limit 0 140 0 577 t4052 1 578 1 890 2 462 Read 2 OPMS o 0 566 TaT 1 558 1 867 2 385 Result PASS PASS PASS PASS PASS PASS Reviewed Approved By Date For Technical Support In the U S In Europe BioTek Instruments Inc BioTek Instruments GmbH Tel 800 242 4685 Tel 49 0 7136 9680 Fax 802 655 3399 Fax 49 0 7136 968 111 All Others Tel 802 655 4040 Fax 802 655 3399 email TAC biotek com www biotek com service Figure 32 2 Sample output for the Absorbance Plate Test page 2 of 2 Note The format varies depending on the software used to run the test Synergy HT Operator s Manual 78 Chapter 4 Instrument Qualification Results amp Troubleshooting Tips The Absorbance Test Plate Report contains results for the following Peak Absorbance When the test is performed the C6 filter is scanned at the test range s defined by the user in the Absorbance Test Plate dialog To verify wavelength accuracy
193. or PCB e Ifthe error persists contact BioTek TAC A D calibration STBY line never went low A D calibration STBY line went low but never transitioned to a high This error indicates a failure with one of the PCBs when trying to initialize the A D or the cable to the defective PCB has lost continuity This error can indicate one of the following A D circuits e Fluorescence analog PCB e Absorbance analog PCB Probable Causes e Fluorescence analog PCB or connection between the analog PCB and motor power supply PCB e Absorbance analog PCB or the cable between the analog PCB and motor power supply PCB Band pass overlap in filter set This error indicates an open hole or overlapping filter set within the optical path of the excitation and emission filter wheels designated by the protocol Equation used for overlapping If EM wavelength is gt EX if EX wavelength 42 bandpass gt EM wavelength 1 2 bandpass then fail else if EX wavelength is gt EM if EM wavelength 1 2 bandpass gt EX wavelength 1 2 bandpass then fail Note Only 2 filter sets can be sent when developing software not Gen5 or KC4 1F01 is for the first set 1F02 is for the second set Invalid parameter value selected This error can occur only during computer control indicating that an invalid assay configuration was sent to the instrument PMT signal too low This error indicates that during the 750V test the PMT signal was less than 1 If this is true
194. or Plate gt Click the Dispense Shake and Read buttons to add steps to the protocol gt When finished click OK to return to the Reading parameters dialog Click OK to verify the parameters and return to the main screen Select Protocol Save As and give the protocol an identifying name Reading Plates To read a plate using KC4 L 2 oe On ae Select Data New Plate If prompted to select a protocol select a protocol and click OK If not prompted select Protocol Open and select a protocol Select Data Read Plate The Plate Reading dialog will appear Click Start Reading The door will open and the plate carrier will extend Place the plate on the carrier and click START READING to begin the read When the read is complete the measurement values will appear in KC4 Select Data Save As and give the file an identifying name Synergy HT Operator s Manual 56 Chapter 3 Getting Started Controlling the Dispense Module This section applies to Synergy HT modds with injectors only KC4 is used to perform several dispense module specific functions including initializing priming and purging KC4 also contains certain configuration items that must be set before using the dispense module Read the following sections to become familiar with these functions and configuration items Initialization If the dispense module was connected to the reader before the reader was turned on or if an Optics Test was r
195. ord the weight and tare the balance e Place the plate on the carrier dispense 20 uL well to columns 5 8 e Remove the plate and weigh it Record the weight and tare the balance e Place the plate on the carrier dispense 5 uL well to columns 9 12 BioTek Instruments Inc Dispense Module Tests 107 e Remove the plate and weigh it Record the weight e Manually pipette 150 pL of deionized or distilled water into all 12 columns on top of the green test dye solution e Place the plate on the carrier for a 15 second shake the 80 uL read at 405 750 nm and the 20 and 5 uL read at 630 750 nm 9 When processing is complete select File Save As and save the experiment using an identifying file name 10 Repeat steps 4 9 using the Synergy HT Dispenser 2 prt protocol 11 See page 109 for instructions for analyzing the results 12 When all tests are complete prime both dispensers with at least 5000 uL of deionized or distilled water to flush out the green dye solution Synergy HT Operator s Manual 108 Chapter 4 Instrument Qualification Test Procedure KC4 2 KS Note If you are using one of BioTek s keypad based readers such as the ELx800 or ELx808 make sure the reader is not running in Rapid mode To check the setting select UTIL gt READ and cycle through the options until READ IN RAPID MODE appears Set it to NO Perform the following set of steps two times to te
196. orresponds to the 6 calibration values Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e No configuration data entered e Values entered are corrupted or incorrect Syringe calibration checksum failed This error indicates that the calibration data has been entered but the checksum failed This corresponds to the 6 calibration values Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Values entered are corrupted or incorrect Synergy HT Operator s Manual 198 Appendix C Error Codes Code 2C03 2C05 2C07 2D00 2D02 Description and Probable Causes During the validation of the syringe calibration data the pL step failed This error indicates that when the software calculated the p1l step the results were either lt 0 0100 or gt 9 9999 Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Values entered are corrupted or incorrect During the validation of the syringe calibration data the calibration data failed This error can indicate one of the following error scenarios e The minimum calibration value is not equal to 5 e The minimum measured value is not between 2 and 6 e The second measured value is not within 20 of the calibrated value e The third through fifth measured values are not within 10 of the calibrated value e The calibration values are not in ascending order e The measured calibration values are not i
197. ot touch the lens with your fingers nspect the block for spills or other contamination Carefully clean with mild detergent if necessary Synergy HT Operator s Manual 148 Chapter 5 Preventive Maintenance Important When cleaning the absorbance lens with the swab apply very little pressure to the lens Applying too much pressure can push the lens out of its holder reinstallation must be performed by BioTek service personnel If the lens does fall out contact BioTek TAC 11 Useacotton swab moistened with alcohol to gently clean the lens on the top probe hanger 12 Slidethe microplate carrier out of the way Use a cotton swab moistened with alcohol to clean the lens on the instrument surface BioTek Instruments Inc Cleaning the Internal Components 149 Cleaning the Reader s Internal Surface 1 3 If you have not already done so unplug the instrument and remove its shroud see page 135 for instructions Follow the instructions under Cleaning the Optical Probes to at a minimum disconnect the incubator wires detach the ground wire lower the top optic probe hanger and remove the incubator housing steps 1 through 6 Manually slide the microplate carrier to the left to engage the support pin and then away from the center surface Microplate carrier fully extended Moisten do not soak a clean cotton cloth with alcohol water or with water and mild detergent Wipe all sides of t
198. ox Synergy HT Operator s Manual 18 Chapter 2 Installation 2 Using no sharp tools open the box containing the dispense module Renove the two reagent bottle holders and the cardboard shipping insert Lift out the module and place it on a level surface Cardboard insert Reagent bottle holders 2 Dispense module Inner shipping box Figure 8 Unpacking the dispense module s inner shipping box 3 Open the accessories box Remove and identify its contents See Figure 9 on the next page 2 inlet tubes packaged in plastic cylinders 4 outlet tubes packaged in plastic bags PN 7082120 2 syringes packaged in boxes 1 priming plate 2 reagent bottles 1 injector tip priming trough small plastic cup 1 plastic tool storage bag with Velcro strips 2 metal thumbscrews 1 stylus wire packaged in asmall plastic cylinder 1 dispense module cover 1 dispense module cable BioTek Instruments Inc 7 Unpack and Inspect the Dispense Module 19 Top foam Dispense module end cap cable Dispense module cover Inlet tubes 2 Syringes 2 Outlet tubes 4 Bottom foam end cap with cutouts Accessories box Figure 9 Unpacking the dispense module s accessories Synergy HT Operator s Manual 20 Chapter 2 Installation 8 Install the Dispense Module This section applies to Synergy HT modds with injectors only 2 Refer to the figures on the next two page
199. pair operations Warning Wear protective gloves when handling contaminated instruments Gloved hands should be considered contaminated at all times keep gloved hands away from eyes mouth nose and ears Warning Mucous membranes are considered prime entry routes for infectious agents Wear eye protection and a surgical mask when there is a possibility of aerosol contamination Intact skin is generally considered an effective barrier against infectious organisms however small abrasions and cuts may not always be visible Wear protective gloves when handling contaminated instruments Important Do not immerse the instrument spray it with liquid or use a wet cloth on it Do not allow water or other cleaning solution to run into the interior of the instrument If this happens contact BioTek s Technical Assistance Center Important Do not apply lubricants to the microplate carrier or carrier track Lubrication on the carrier mechanism or components in the carrier compartment will attract dust and other particles which may obstruct the carrier path and cause the reader to produce an error Caution The buildup of deposits left by the evaporation of spilled fluids within the read chamber can impact measurements Be sure to keep System Test records before and after maintenance so that changes can be noted Caution Models with injectors Before removing the reader s cover to expose internal parts purge the dispense module turn
200. penser tested Tip You can take advantage of Gen5 s Power Export feature to create Microsoft Excel spreadsheets for performing the Results Analysis calculations See the Gen5 Help System for information on the Power Export Builder Test Setup KC4 e Using KC4 create three Dispense protocols for the Synergy HT If you are using a BioTek absorbance reader create two additional Read protocols See page 115 for instructions e If you are not using a BioTek absorbance reader prepare your reader to perform two reads with the following characteristics For your convenience we ve included a worksheet at the end of this chapter for recording the dispense weights Delta OD values calcula tions and pass fail Make two copies of this worksheet one for each dispenser tested Tip You can take advantage of KC4 s PowerReports feature to create Microsoft Excel spreadsheets for performing the Results Analysis calculations See KC4 s User Guide for information on PowerReports Synergy HT Operator s Manual 106 Chapter 4 Instrument Qualification Test Procedure Gen5 1 Prime both dispensers with deionized or distilled water e Ensure that the tip priming trough BioTek PN 7082118 is installed in the microplate carrier e Place a clean priming plate BioTek PN 7132158 or 7092135 on the carrier e Fill a supply bottle with deionized or distilled water and insert the inlet tube e InGen5 selec
201. peration selected INVALID PROFILE soa motor profile selected has too many steps for move NO CATCH PLATE TAN catch plate not in carrier for prime or purge Dispenser Configuration Data Error Test Codes lowest digit in returned error code DATA NOT SET MENA disp cnfg data not set CHKSUM FAILED ND disp cnfg data checksum failed ULPERSTEP OUT OF RANGE mgA syringe pl step factor not within allowed limits INJECTOR POSITION ERR ma invalid injector position or position not set VOLUME CAL DATA ERR RD volume cal data is invalid PULLBACK VOLUME ERR nge pull back volume too large DEFAULT VOL CAL DATA SET we indicates that default cal volume data set and should be updated via the serial host with the cal data values from the attached syringe drive TWO BOTTOM INJECTORS ig only one injector can be at bottom position BioTek Instruments Inc Status String Format 205 Dispense Read Runtime Definition Error Test Codes lowest two digits in returned error code SAMPLE START LATE MQW individual sample missed its scheduled start time FILTERSETS OUT OF RANGE MOY more than two filters selected for disp read method FILTER SWITCHING OVERLAP Q2 filter bandpass overlap condition exists in the intermediate switching position for multi filterset reads FILTERS NOT ADJACENT NOB ex or em filters not adjacent for multi filterset reads EX FILTER PLUG POSITION 04 filter plug s not positioned next to ex filter s for
202. plate Propidium Iodide PI Bottom reading 62 5 ng ml solution of Propidium Iodide in PBS 50 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 645 40 Corning Costar 96 well black sided clear bottom plate Synergy HT Operator s Manual 212 Appendix D Specifications 3 mm probe Sodium Fluorescein SF Bottom reading 20 pg ml solution of Sodium Fluorescein in PBS 150 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 530 25 Hellma 96 well quartz plate Propidium Iodide PI Bottom reading 125 ng ml solution of Propidium Iodide in PBS 50 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 645 40 Corning Costar 96 well black sided clear bottom plate Methylumbelliferone MUB Top reading 0 16 ng ml solution of Methylumbelliferone in CBB 300 uL per well signal to noise ratio greater than 2 0 Excitation 360 40 Emission 460 40 Corning Costar black strips 1 5 mm probe Sodium Fluorescein SF Bottom reading 40 pg ml solution of Sodium Fluorescein in PBS 150 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 530 25 Hellma 96 well quartz plate Propidium Iodide PI Bottom reading 250 ng ml solution of Propidium Iodide in PBS 50 uL per well signal to noise ratio greater than 2 0 Excitation 485 20 Emission 645 40 Corning Costar 96 well black sided clear bottom plate Me
203. ppendix A for decontamination instructions Caution Remove the microplate and tip prime trough if equipped from the carrier before shipment Spilled fluids can contaminate the optics and damage the instrument Important The instrument s packaging design is subject to change over time If the instructions in this section do not appear to apply to the packaging materials you are using please contact BioT ek s Technical Assistance Center for guidance Replace the microplate carrier shipping screw and the shipping panel before repackaging the reader for shipment Please contact BioT ek if you have misplaced either of these items If you need to ship the Synergy HT and or the dispense module to BioTek for service or repair be sure to use the original packaging materials Other forms of commercially available packaging are not recommended and can void the warranty The shipping materials are designed to be used no more than five times If the original materials have been damaged lost or used more than five times contact BioTek to order replacements PN 7093001 for the reader PN 7083001 for the dispense module See page 6 for contact information Perform these steps to prepare the reader for shipment 1 Contact BioTek s Technical Assistance Center for an RMA Return Materials Authorization number before returning equipment for service See page 6 for contact information 2 Decontaminate the reader and if attached
204. r set it to 96 WELL PLATE Set First Well and Last Well as follows First Last Well A1 H4 A5 H8 A9 H12 Leave all other parameters set to their defaults Click OK to return to the Main Menu Select Protocol Save As and give the protocol an identifying name that includes the dispenser number and the dispense volume such as Dispenser 1_80 ul Keep the protocol open Select Protocol Reading again and return to the Read amp Dispense dialog Double click on the DISPENSE item as shown below to open the Dispense dialog and change the Dispenser to 2 Read amp Dispense ISPENSE Syringe 80yl at 275 m Return to the main menu and select Protocol Save As Modify the file name so that it includes dispenser number 2 e g Dispenser 2_80 ul BioTek Instruments Inc Dispense Module Tests 117 To create the Read protocols start by launching KC4 and setting the current reader to the absorbance reader Perform the following set of steps two times to create two protocols for the 80 uL and 20 amp 5 ul reads 1 2 10 If a Data file is open close it now Data Close Select Protocol New and then Protocol Reading The Reading Parameters dialog will open See the sample screen shots on the next page If the current reader is the Synergy set Detection Method to Absorbance Select two Wavelengths as follows Note KC4 automatically creates a multi plate transformation to subtr
205. recommend cleaning the internal tubes and injector heads along with the optical probes Instructions for removing and cleaning these components are provided on pages 137 through 140 e Before starting this procedure gather some supplies gt gt gt gt Small container of isopropyl alcohol Small container of deionized or distilled water Lens cleaning tissue Cotton swabs Synergy HT Operator s Manual 142 Chapter 5 Preventive Maintenance Take a moment to identify the components discussed in this section Thumbscrew to access ground wire Thumbscrews 2 black for holding the incubator housing in place Screw for lowering and raising the top optical probe Top optical probe Top probe hanger with two shoulder screws Sas Figure 38 Internal components to be removed adjusted for cleaning the optic probes Incubator A housing BioTek Instruments Inc Cleaning the Internal Components 143 Once the shroud has been removed and the internal tubes and injector heads have been removed and cleaned see page 140 follow these instructions to remove a few more components and then clean the optical probes 1 Disconnect the heater and thermistor wires To do this depress the small tab pictured below and separate the connectors 2 Remove the thumbscrew located in the left rear of the instrument and set it aside This exposes the ground wire Synergy
206. rgy HT Operator s Manual 104 Chapter 4 Instrument Qualification Test Solution Recipes 80 UL of test solution with 150 uL of deionized water should read between 1 300 and 1 700 OD at 405 750 nm It is assumed that the solutions used are at room temperature If you do not have BioTek s Green Test Dye Solution PN 7773003 prepare a green test dye solution using one of the following methods Using BioTek s Blue and Yellow Concentrate Dye Solutions Ingredient Quantity Concentrate Blue Dye Solution PN 7773001 125 mL QC Yellow Solution PN 7120782 125 mL Deionized water 90 0 mL Using FD amp C Blue and Yellow Dye Powder Ingredient Quantity per Liter FD amp C Blue No 1 0 200 grams FD amp C Yellow No 5 0 092 grams Tween 20 1 0 mL Sodium Azide N3Na 0 100 gram BioTek Instruments Inc Dispense Module Tests 105 Test Setup Gen5 To perform the Dispense Accuracy and Precision Test using Gen5 you ll need to create two protocols containing the necessary Dispense and Read steps one protocol per dispenser You can create these protocols once and then reference them in new experiments each time you run the test See page 110 for instructions for creating these protocols For your convenience we ve included a worksheet at the end of this chapter for recording the dispense weights Delta OD values calcula tions and pass fail Make two copies of this worksheet one for each dis
207. rophotometer WAVELENGTH nm am a 040 0 881 0 783 C E E FS E e e Set 2453 Serial 161259 Figure 31 Sample Standards Certificate showing OD Wavelength combinations for each of six locations on the Absorbance Test Plate Before the Absorbance Plate Test can be performed the standard OD values and the peak wavelength value s must be entered into Gen5 or KC4 Instructions for defining the Test Plate s characteristics and for running the test are provided for both Gen5 and KC4 on the following pages Synergy HT Operator s Manual 72 Chapter 4 Instrument Qualification Setup Gen5 To define the Absorbance Test Plate parameters using Gen5 Note The Gen5 Reader Diagnostics Utility must be installed 1 Obtain the certificates that came with the Test Plate 2 Start Gen5 and select System Diagnostics Test Plates Add Modify Plates Click Add The Absorbance Test Plate dialog will appear 4 Select the appropriate Plate Type and enter the plate s Serial Number Enter the Last Certification and Next Certification dates from the calibration sticker on the Test Plate 6 If the wavelength values in the top row of the grid are appropriate for your tests carefully enter the OD values from the Standards Certificate into the grid Make sure you enter the correct value for each well wavelength combination e If you need to change the wavelength values click the Wavelength List button Cli
208. ror Codes 177 Code Description and Probable Causes 0504 Light beam saturated too much light Air measurement channel reading reached 65535 This error indicates that filter 4 in absorbance mode has saturated the reference channel Probable Causes e The monochromator mirror grating is damaged e Absorbance analog PCB has intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply 0505 Measurement channel light beam saturated too much light Fail if gt 3 steps of error This error can indicate one of the following scenarios e 1 When the monochromator is trying to find the center of the white light home position the monochromator is not able to find home or it found home somewhere other than where it found home before e 2 Filter 5 in absorbance mode has saturated the reference channel e 3 During a fluorescence read a well saturated the PMT Probable Causes Scenario 1 Monochromator did not find home e Flash bulb skipped a flash due to a defective lamp connection or power supply e Order sorting filter wheel is jammed not aligning the through hole with the light path e Monochromator is
209. rs on COM2 Absorbance Fluorescence Luminescence r Emission Center i a 360 x 40 1 sco gt 40 Se aoe Center Bandwidth Center Bandwidth Get Filters ps wil 2 2 ss alao karma so z zipo serne 530 x 25 4 Eo I 40 ole Pua h U pdating Gen5 s filter table for complete U pdating KC 4 s filter table for complete instructions see page 46 instructions see page 52 e When defining a filter set in a Read e When defining a filter set in a KC4 step in aGen5 procedure selecting Luminescence or Multi Mode protocol Hole indicates the empty location in selecting Lum E indicates the empty the Emission filter wheel See page 47 location in the Emission filter wheel for information on Read steps and See page 53 for information on procedures protocols p Fiter Sets m Filters eH Filter Set Escitutions Mode Emission Hole Optics Position ea Optics Position Options ae Sensitivity BioTek Instruments Inc Key Components 43 The External Dispense Module This section applies to Synergy HT modds with injectors only The dispense module pumps fluid from the supply bottles to injector heads located inside the instrument Fluid is injected into one well at atime Figure 23 Dispense module components Two 250 uL syringes draw fluid from the supply bottles Inlet tubes transport fluid from the supply vesse
210. s Absorbance mode e The rail nylon bushings or bearings are dirty causing the carrier to jam e The Autocal jig is not in the carrier e Order sorting filter wheel is jammed and is not able to turn to the open hole Fluorescence mode e The rail nylon bushings or bearings are dirty e The Autocal jig is not in the carrier or mirrors are facing the wrong direction EX EM filter wheel did not home This error code indicates older basecode Recommend upgrading This error indicates the filter wheel did not rotate to activate the Hall Effect home sensor Probable Causes e The Filter Wheel is removed when the instrument is powered on and the wheel positioning is changed e The filter wheel is jammed Reseating the filter wheel may correct this Monochromator order sorting filter wheel did not home This error is caused when the Order sorting Filter Wheel cannot rotate to the home sensor position Probable Causes e The motor or motor driver circuit is defective e The home sensor or sensor circuit is defective Synergy HT Operator s Manual 172 Appendix C Error Codes Code Description and Probable Causes 0305 Saturation transition failed in the Monochromator motor movement light beam never found During the instrument initialization the monochromator is homed by rotating the monochromator mirror until the white light full light is detected This requires a fully functional Flash lamp detection system Probable Causes
211. s allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Scenario 2 Spectral scan read e The monochromator mirror grating is damaged e Absorbance analog PCB intermittently failed e Missed flashes or an erratic flash lamp e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply Measurement channel light beam saturated too much light Air reading reached 65535 This error indicates that during a read or system test one of the filters from 1 to 6 in absorbance mode has saturated the measurement channel The last number is the lambda table position number Probable Causes e The monochromator mirror grating has a defect e Absorbance analog PCB intermittently failed e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Voltage to the lamp has increased due to failure of the lamp power supply or the motor power supply PCB sent the wrong voltage request to the lamp power supply BioTek Instruments Inc Error Codes 179 Code 0600
212. s for guidance while performing these steps Perform these steps to install the dispense module L Place the dispense module to the left side or on top of the reader See the photos on the next page On the rear panel of the Synergy HT identify the SYRI NGE 1 and SYRINGE 2 tubing ports Remove the nylon screws from both ports Open two of the plastic bags containing the outlet tubes labeled as PN 7082120 Remove the clear plastic fitting covers from the tubes Put the other two bags in a safe place they are spares Place the nylon screws and the plastic fitting covers in the plastic tool storage bag Use the supplied Velcro strips to attach the bag to the rear panel of the dispense module Remove the two inlet tubes from their protective plastic canisters Identify the two circular syringe valves on the dispense module Each is labeled with a left pointing arrow See Figure 12 on the next page When installing the inlet and outlet tubes do not use any tools Finger tighten only 10 11 Screw the fitting of one inlet tube into the right side of the Syringe 1 valve Screw one end of one outlet tube into the left side of the Syringe 1 valve Screw the other end of the outlet tube into the SYRI NGE 1 port on the rear of the Synergy HT Repeat steps 7 through 9 to attach the inlet and outlet tubing for Syringe 2 Seat the outlet tubes in the clip to the left of the Syringe 2 valve Continued on page 21 B
213. s on internal components Caution Environmental Conditions Do not expose the system to temperature extremes For proper operation ambient temperatures should remain between 18 40 C Performance may be adversely affected if temperatures fluctuate above or below this range Storage temperature limits are broader Caution Sodium Hypochlorite Do not expose any part of the instrument to the recommended diluted sodium hypochlorite solution bleach for more than 20 minutes Prolonged contact may damage the instrument surfaces Be certain to rinse and thoroughly wipe all surfaces Caution Power Supply Only use the power supply shipped with the instrument Operate this power supply within the range of line voltages listed on it Caution Shipping Panel and Carrier Shipping Screw The shipping panel and carrier shipping screw must be removed before operating the reader They must be reinstalled before repackaging the reader for shipment See Chapter 2 Installation Caution Disposal This instrument contains printed circuit boards and wiring with lead solder Dispose of the instrument according to Directive 2002 96 EC on waste electrical and electronic equipment WEEE Caution Electromagnetic Environment Per IEC 61326 2 6 it is the user s responsibility to ensure that a compatible electromagnetic environment for this instrument is provided and maintained in order that the device will perform as intended Caution Electromagnet
214. s wake venievieeies AKARCA ees sheave 27 14 Test the Injector SyStem siisii a i AE vende leuac ee eee easel neeek sans 28 Register With BiOT eK cviic vine senacie arnee Naan bard teneawed eens aaea Na NN dowd 30 Synergy HT Operator s Manual iv Preface Operational Performance Qualification cccecceeeeeee eee eeae teat eeeeeneeeae ees 30 Repackaging and Shipping Instructions sceeeeee eee eee eee eee ee ea ees 31 Chapter 3 Getting Started ccsccceseeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeseeeeeeeeeneneae 35 Key COMpOnents ross Iese RAEE E sever veal avr add tesa ESETET 36 Power Switch Carrier Eject Button Microplate Carrier sasse 36 Lamp Assembly and Filter Wheel ACCESS ccccceeeeeseeeaeeeeeeeeeeeenees 37 Excitation and Emission Filter WheelS ccccceceeeseeeeeeeeeeeeeeeeaeees 38 Installing the Time Resolved Fluorescence Cartridge e0ee 41 Configuring the System for Luminescence Measurements 42 The External Dispense MOdUIe cccecceee eee e eee esse eee nate neenenenee ened 43 Gen Software orioa grin heise aana AAKA AANE cowie tetsawieas CENAA NEARNE 46 Viewing Updating the Filter and Wavelengths Tables sances 46 Creating Protocols and Experiment ccceeeeeeeee estes eee eee eee enenes 47 Controlling the Dispense MOdUule cecceeeeee eee ee eee eeeeeeeaetaeeaenes 50 KC4 SoftWare niiin edivntersasend cence chee Vdnis tated ies eeeedvar Padawalale tacts ves 52 Viewi
215. second lowest digit in returned error code Range Error o Thermistor Error NLS A D Error wee Note Affected zones are encoded in the lowest digit returned one bit per zone starting from bit 0 Data Flash Codes second lowest digit in returned error code Readback Error ba Oe data readback didn t match data written Copy Error alee final data readback didn t match original passed in A D Device Codes lt lowest digit in returned error code gt Absorbance measurement RA Fluorescence measurement a Incubation measurement SEs Voltage reference 4 A D Device Codes lt second lowest digit in returned error code gt A D ready line ng A D converter never saw ready line transition A D noise 3200 belles A D samples inconsistent Probe Z Calibration Codes lt lowest digit in returned error code gt Jig Windup DAK not enough windup at jig Syringe Motor Error Test Codes lt lowest digit in returned error code gt OFF SENSOR ree syringe off sensor when it should be on ON SENSOR me syringe on sensor when it should be off SENSOR CLEAR COUNT wore sensor clear count out of range FMEA TEST FAIL 47 clear count measured during aspirate operation deviated more than allowed range from initial value MOVE OVERRANGE Sa number of microsteps requested for move too large SYR NOT HOME No syringe logical position not at home INVALID POSITION pres invalid position passed to position syringe function INVALID OPERATION bits invalid syringe o
216. ser 1 2 e Calculate the Standard Deviation of the 32 wells e Calculate the Mean of the 32 wells e Calculate the CV Standard Deviation Mean x 100 e Calculate the Accuracy o Error Actual Weight Expected Weight Expected Weight 100 Expected Weights for 32 wells 80 ul 2 560 g 20 ul 0 640 g 5 ul 0 160 g It is assumed that one gram is equal to one milliliter Dispense To pass To pass Accuracy Volume CV must be Error must be lt 10 0 lt 20 0 Failures If any tests fail prime the fluid lines and rerun the test s If the test s fail again the injector heads may require cleaning see Chapter 5 Preventive Maintenance If tests continue to fail contact BioTek s Technical Assistance Center Synergy HT Operator s Manual 110 Chapter 4 Instrument Qualification Creating the Test Protocols Using Gen5 This section contains instructions for creating two Gen5 protocols specifically for performing the Synergy HT Dispense Precision and Accuracy test Refer to the Gen5 Help system to learn more about using Gen5 and for complete instructions for creating protocols To create the protocols in Gen5 1 Start by selecting System Reader Configuration and add configure the Synergy if it is not already there 2 Select File New Protocol A menu tree will appear e To edit a protocol category double click its branch in the tree as shown below S Gen5 Se
217. shipping pane that has two handles for lifting Locate and grasp the handles Carefully lift the reader out of the box and place it on a level surface Remove the protective plastic bag 4 Placeall packing material back into the shipping box for reuse if the reader needs to be shipped again See Package Contents in Chapter 1 for assistance with identifying the contents of the accessories box Accessories box Shipping panel a Inner shipping box eal Figure 1 B Unpacking the reader s inner shipping box BioTek Instruments Inc 2 Remove the Shipping Panel 11 2 Remove the Shipping Panel Perform these steps to remove the shipping panel from the bottom of the reader 1 Carefully tip the reader onto its back 2 Using aslotted screwdriver remove the four screws and washers attaching the shipping panel to the bottom of the reader See Figure 2 on the next page 3 Carefully set the reader upright 4 Locate the supplied plastic tool storage pocket Place the screws and washers inside the bag Use the supplied Velcro strips to attach the pocket to the back of the reader for storage Do not block any air vents See Figure 3 on the next page 5 Place the panel back into the inner shipping box for storage Important Reattach the shipping panel before repackaging the Synergy HT for shipment Synergy HT Operator s Manual 12 Chapter 2 Installation
218. spense Module 23 14 Locate the dispense module cable Plug one end into the port on the left side of the dispense module Plug the other end into the Dispenser Port on the rear panel of the Synergy HT One end of the cable connected to the port on the side of the dispense module SYRINGE 1 The other end connected to the reader s Dispenser Port Figure 14 Dispense module connected to the reader rear view 15 Locate the injector tip cleaning stylus packaged in a small plastic cylinder Attach the cylinder to the back of the dispense module for storage Synergy HT Operator s Manual 24 Chapter 2 Installation 9 Connect the Host Computer The Synergy HT is equipped with two types of communication ports Serial RS 232 and USB Both ports are located on the rear pane of the reader e Both types of cables are included in the accessories box Determine which cable is supported by the host computer e Connect one end to the appropriate port on the reader see photo below and the other end to the appropriate port on the host computer Figure 15 RS 232 serial and USB ports on the rear panel injector model shown 10 Install the Software on the Host Computer The Synergy HT is controlled by BioTek s Gen5 or KC4 software running on a host computer There is a certain sequence of events that must be followed to ensure that the software is properly installed and configured Please follow
219. st both dispensers Using KC4 prime the fluid lines with deionized water e Fill a reagent bottle with deionized water and insert the inlet tube e Select System Reader Control and click the Dispenser tab Set the Dispenser number 1 or 2 enter a Volume of 4000 uL and click Prime Place the priming plate on the carrier and click OK e Remove the inlet tube from the bottle and run another prime with the volume set to 2000 uL Empty the plate and replace it on the carrier Fill a reagent bottle or beaker with 20 mL of the green dye solution Prime the fluid lines with 2000 ul of the green dye Remove and empty the priming plate Perform these steps carefully When each dispense protocol is finished you will weigh the plate record the weight and tare the balance 10 11 12 13 Place a new 96 well microplate on the balance and tare the balance Place the plate on the carrier and run the 80 pl Dispense protocol created for the dispenser under test Weigh the plate Record the 80 uL dispense weight and then tare the balance Place the plate on the carrier and run the 20 pl Dispense protocol created for the dispenser under test Weigh the plate Record the 20 uL dispense weight and then tare the balance Place the plate on the carrier and run the 5 pl Dispense protocol created for the dispenser under test Weigh the plate Record the 5 uL dispense weight Pipette or dispense 150 pL of deionized wat
220. stallation and setup process is complete Turn to the next page to read about product registration and Operational Performance Qualification BioTek Instruments Inc Operational Performance Qualification 31 Operational Performance Qualification Your Synergy HT Multi Detection Microplate Reader was fully tested at BioTek prior to shipment and should operate properly following the successful completion of the installation and setup procedures described throughout this chapter If you suspect that problems occurred during shipment if you received the reader back from BioTek following service or repair and or if regulatory requirements dictate that Operational Performance Qualification is necessary turn to Chapter 4 Instrument Qualification now to learn about BioTek s recommended OQ PQ procedures for the Synergy HT Note An I nstallation Operational Performance 1Q OQ PQ package for the Synergy HT is available for purchase PN 7090521 Contact your local BioTek dealer for more information Synergy HT Operator s Manual 32 Chapter 2 Installation Repackaging and Shipping I nstructions Warning If the reader and or dispense module has been exposed to potentially hazardous material decontaminate it to minimize the risk to all who come in contact with the reader during shipping handling and servicing Decontamination prior to shipping is required by the U S Department of Transportation regulations See A
221. sted is too large for the unit The X axis tripped the XY sensor The Corning Costar 12 well plate part number 430345 has a width dimension that is causing a 2600 error with Synergy Simply changing the width to 84320 will alleviate the error message and will not affect the results In KC4 v3 4 r16 the width was changed to 84320 and the 2600 error does not appear when running a protocol that specifies the Corning Costar 12 Well plate type lt Motor gt currently in use This error indicates that the lt motor gt is not available for this model or already has a task assigned to it At the beginning of the motor_setup function the basecode checks to see if the motor is currently in use or is not available This error can occur with any motor request See table below for errors Error Motor 2800 X axis 2801 Y axis 2802 EX motor 2803 EM motor 2804 Order sorting filter wheel 2805 Monochromator 2806 Probe Height 2807 Probe changer 2808 Syringe 1 2809 Syringe 2 Probable Causes e User selected the wrong model in the controlling software e The incorrect basecode was downloaded to the instrument Fluorescence Lamp off voltage out of range 0 730 This error indicates that during a self test the lamp did not indicate that it was off Probable Causes e The sense resistor is damaged on the 7090402 PCB e Cable between the lamp PCB and motor power supply PCB is defective e Motor power s
222. successful completion of the OQ procedure in combination with results that are comparable to previous PQ and OQ tests confirms that the equipment is operating according to specification initially and over time Performance Qualification confirms that the reader consistently meets the requirements of the tests performed at your laboratory The recommended PQ procedure consists of performing the System Test Absorbance Plate Test a series of Liquid Tests and if the external dispense module is used the Dispense Accuracy and Precision Tests Your facility s operating policies may also require that you routinely perform an actual assay to confirm that the reader will consistently give adequate results for the assays to be run on it These tests should be performed routinely the recommended interval is monthly or quarterly depending on the test This frequency may be adjusted depending on the trends observed over time The successful completion of the PQ procedure confirms that the equipment is performing consistently under normal operating conditions Synergy HT Operator s Manual 64 Chapter 4 Instrument Qualification Recommended Qualification Schedule The following schedule defines the factory recommended intervals for qualifying a Synergy HT used two to five days a week The schedule assumes the reader is properly maintained as outlined in Chapter 5 Preventive Maintenance i Initially Initially Annually Monthly Quar
223. syringe drive Probable Causes Linear way is dirty or lack of lubrication is causing the bearings to jam The lead screw is loose on the motor shaft because the glue is no longer bonding it to the motor shaft Defective optical sensor Defective motor Motor Controller PCB or cable Cable between the dispenser and the Synergy is defective too long or there is a lost connection For certain older dispensers opto cable 7330506 may need to be replaced with 7120734 BioTek Instruments Inc Code 0300 0301 0302 0303 0304 Error Codes 171 Description and Probable Causes Saturation transition failed in the X axis movement light beam never found This error indicates that during the X axis movement the light beam saturation transition max light to no light was never found during autocalibration Probable Causes Absorbance mode e The rail nylon bushings or bearings are dirty causing the carrier to jam e The Autocal jig is not in the carrier e Order sorting filter wheel is jammed and is not able to turn to the open hole Fluorescence mode e The rail nylon bushings or bearings are dirty e The Autocal jig is not in the carrier or mirrors are facing the wrong direction Saturation transition failed in the Y axis movement light beam never found This error indicates that during the Y axis movement the light beam saturation transition max light to no light was never found Probable Cause
224. t Input X Range to reference the above mentioned X values Click OK to perform the analysis the results of which will be output in a separate sheet Note If the Data Analysis command is not available on the Tools menu you may need to install the Analysis ToolPak in Excel Consult Excel s help system for assistance Since it is somewhat difficult to achieve high pipetting accuracy when conducting linear dilutions an R Square value of at least 0 99 is considered adequate BioTek Instruments Inc Fluorescence Tests 89 Fluorescence Tests The Corners Test uses fluorescent compounds to verify that the plate carrier is properly aligned in relation to the optical probe s Because the Synergy HT s fluorescence optics are different from the absorbance optics the Corners Test is also required We recommend running the test for both the top and bottom probes if equipped The Sensitivity Test uses fluorescent compounds of varying concentrations to test the fluorescence reading capability of the reader The ability to detect specific compounds at low concentrations ensures that the filters optical path and PMT are all in working order This test verifies that the difference between the means of wells with known lower limits of concentration of the substance under investigation is statistically distinguishable from the mean of wells with pure diluent The Linearity Test verifies that the system is linear that is signal
225. t lt Channel gt This is not consistent with other error codes This error indicates that the absorbance reference channel gain for a specific wavelength during Time Resolved readings is out of the range necessary to ensure the lambda performance to specifications The second to last number is the lambda table position number Probable Causes e The absorbance reference channel PCB is defective e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path or the through hole is allowing white light to pass e Monochromator is defective e Flash lamp alignment or Flash lamp power supply is defective e Lamp is too bright Measurement channel gain out of range for the selected wavelength Fail if Air reading gt 60000 with a Gain 1 This error indicates that during a filter calibration or testing the reader prior to a read one of the filters saturated the measurement channel The last number is the lambda table position number Probable Causes e The monochromator mirror grating is damaged e Order sorting filter wheel is jammed not aligning the correct bandpass filter with the light path the filter is degraded and not passing enough light energy or filter is blocking the light e Absorbance analog PCB has intermittently failed e Missed flashes or an erratic flash lamp e Voltage to the lamp has decreased due to failure of the lamp power supply or the motor power supply PCB sent the
226. t Check the neutral density filters on the Test Plate to ensure there is no debris that may have shifted between readings and caused changes gt Check the microplate carrier to ensure it is clear of debris Linearity of the optical density readings is confirmed by default if the optical density readings are accurate To further verify this perform a regression analysis on the Test Plate OD values in a program such as Microsoft Excel as follows 1 Launch Excel 2 Create a spreadsheet and label one column Assigned and the next column Observed 3 Enter the Assigned OD data for each glass filter in the first column from the Standards Certificate provided with the Test Plate Analyze one wavelength at a time 4 Enter the Observed OD values for the same glass filters in the adjacent column 5 Under Tools select Data Analysis and then Regression Define the Assigned values as the Input Y Range and the Observed OD as the Input X Range Note If the Data Analysis command is not available on the Tools menu you may need to install the Analysis ToolPak in Microsoft Excel Consult Microsoft Excel Help for assistance 6 Click OK and the Summary Output sheet will be displayed An R Square value of at least 0 990 is expected Synergy HT Operator s Manual 80 Chapter 4 Instrument Qualification Absorbance Liquid Tests Conducting Liquid Tests confirms the Synergy HT s ability to perform
227. t System gt Reader Control If prompted select the reader under test e Click the Dispenser tab Select Dispenser 1 or 2 If the Initialized status is No click Initialize e Enter a prime Volume of 4000 uL and click Prime e When finished remove the priming plate empty it and put it back on the carrier Remove the inlet tubes from the supply bottles Prime both dispensers with the Volume set to 2000 uL This prevents the water from diluting the dye Fill a beaker with at least 20 mL of the green dye solution Prime both dispensers with 2000 uL of the solution When finished remove the priming plate from the carrier Create a new experiment based on the Synergy HT Dispense 1 prt file described on page 110 Place a new 96 well microplate on the balance and tare the balance Place the plate on the microplate carrier Important Gen5 will provide prompts and instructions for processing the plates follow the steps carefully When each dispense step is finished you will weigh the plate record the weight tare the balance with the plate on it and then place the plate back on the carrier for the next step Select Plate Read and click READ Gen5 will prompt you to empty the tip priming trough When ready click OK at the Load Plate dialog to begin the experiment Follow the prompts displayed on the screen the sequence is as follows e Dispense 80 uL well to columns 1 4 e Remove the plate and weigh it Rec
228. t values may be affected by extraneous particles such as dust in the microplate wells A clean work area is necessary to ensure accurate readings When operated in a safe environment according to the instructions in this document there are no known hazards associated with the instrument However the operator should be aware of certain situations that could result in serious injury these may vary depending on the instrument model See Hazards and Precautions Hazards and Precautions Hazards A Warning Power Rating The instrument s power supply or power cord must be connected to a power receptacle that provides voltage and current within the specified rating for the system Use of an incompatible power receptacle may produce electrical shock and fire hazards Warning Electrical Grounding Never use a two prong plug adapter to connect primary power to the external power supply Use of a two prong adapter disconnects the utility ground creating a severe shock hazard Always connect the power cord directly to a three prong receptacle with a functional ground Warning Internal Voltage Always turn off the power switch and unplug the power supply before cleaning the outer surface of the instrument Warning Liquids Avoid spilling liquids on the instrument fluid seepage into internal components creates a potential shock hazard Wipe up all spills immediately Do not operate the instrument if internal components have been exposed to flui
229. tarting on page 168 for more information You can look up an error code to determine its probable cause e An A indicates a more serious error with the memory or processing In this case the instrument may be unable to respond turn the instrument off and on again You may be able to use the instrument after restarting it See Fatal Errors on the next page If an error code is displayed run a System Test for diagnostic purposes e If you re using Gen5 select System Diagnostics Run System Test e If you re using KC4 select System Diagnostics Run Optics Test 2 The error code tables in this manual serve a varied audience which includes personnel equipped to service instruments Use these tables to assist you with diagnosing problems and solving them if possible If you need help or more information contact BioTek s Technical Assistance Center See page 6 for contact information For errors that are displayed during operation of the Synergy HT with the BioStack Microplate Stacker refer to the BioStack Operator s Manual BioTek Instruments Inc Error Codes 167 Fatal Errors Code A100 A200 A300 A301 A302 A304 A400 A500 A502 A600 A700 A800 A900 AAO1 AA02 Fatal errors indicate conditions that require immediate attention If a fatal error is displayed contact BioTek s Technical Assistance Center for further instructions Description Task control block not available
230. terly Absorbance Tests Absorbance Plate Test Liquid Test 1 or Liquid Test 2 Liquid Test 3 Fluorescence Tests Corners Test Sensitivity Linearity Tests Tests for Injector Models Injector System Test Dispense Accuracy and y Precision Tests Regarding Liquid Tests 1 and 2 e If you have an Absorbance Test Plate run Liquid Test 1 e If you do not have an Absorbance Test Plate run Liquid Test 2 Liquid Test 3 is optional it is provided for sites requiring verification at wavelengths lower than those attainable with the Absorbance Test Plate Important The risk factors associated with your assays may require that the Operational and Performance Qualification procedures be performed more frequently than shown above BioTek Instruments Inc System Test 65 System Test Description The System Test begins with a check of the stepper motor driven transmission axes within the instrument each is sequentially homed and verified The two measurement systems Absorbance and Fluorescence are checked for noise and signal levels The incubation system is monitored to make sure all zones have thermistor readings within expected ranges The analog power supply levels are measured to make sure all are within expected limits A configuration data area in memory is tested to make sure all of the calibration information is present and checksums correctly If any area tests outside of programmed limits the reader wil
231. the Detection Limit in pg mL 1000 Mean SF Mean Buffer 3 Standard Deviation Buffer Optic Probe Detection Limit must be less than Bottom 5 mm 26 pM 10 pg mL Bottom 3 mm 53 pM 20 pg mL 106 pM 40 pa ml 53 pM 20 pa mt Top 1 5 mm 106 pM 40 pg mL Typical performance BioTek Instruments Inc Fluorescence Tests 93 Linearity Test 1 Calculate the Mean of the four wells for each concentration in columns 1 5 rows C F only 2 Perform linear regression using these values as inputs a a ee 1000 mean of the 1000 pM 1 0 nM wells 500 mean of the 500 pM 0 5 nM wells 250 mean of the 250 pM 0 25 nM wells 125 mean of the 125 pM 0 125 nM wells mean of the 62 5 pM 0 0625 nM wells 3 Calculate the R Squared value it must be greater than or equal to 0 950 to pass Troubleshooting If any tests fail please try the suggestions below If the test s continue to fail print the results and contact BioTek s Technical Assistance Center e Are the solutions fresh The open buffer and stock solutions should be discarded after seven days e Are the Excitation Emission filters clean Are they in the proper locations and in the proper orientation in the filter wheels e Are you using new clean plates We suggest you re run the test with a new clean microplate For the bottom optics test if the base of a plate is touched clean the entire base with alcohol 95 ethanol and then wipe with a l
232. the instrument is on turn it off and allow the lamp to cool down before attempting to replace it Perform these steps to install the fluorescence lamp assembly 1 Locate the lamp assembly in the accessories box The lamp is attached to a metal bracket that also holds a condenser lens and a heat absorber Two cables are attached to the back of the lamp Open the hinged door on the front of the reader by pressing on its lower left and right corners The lamp compartment is on the far left Orient the lamp assembly as shown below Slide the assembly all the way into the compartment Plug the lamp cables into the power source located to the right of the lamp Either cable can be plugged into either socket Close the hinged door Figure 5 Installing the fluorescence lamp assembly replacement lamp PN 7080500 BioTek Instruments Inc 5 Select an Appropriate Location 15 5 Select an Appropriate Location Install the Synergy HT on a level surface in an area where ambient temperatures between 182 and 40 C can be maintained The reader is sensitive to extreme environmental conditions Avoid the following e Excessive humidity Condensation directly on the sensitive electronic circuits can cause the reader to fail internal self checks The specified relative humidity range for this reader is from 10 to 85 non condensing e Excessive ambient light Bright sunlight or strong incandescent light may affect the rea
233. the read chamber can impact performance of both the fluorescence and absorbance functions Be sure to perform a System Test before and after maintenance so that any changes in performance can be noted Synergy HT Operator s Manual 134 Chapter 5 Preventive Maintenance Required Materials Warning Always wear protective gloves and safety glasses when performing cleaning maintenance procedures For all tasks e Protective gloves e Safety glasses For removing the shroud and some of the internal components e Phillips head screwdriver e 1 8 Allen wrench e 3 32 Allen wrench For cleaning the internal dispense tubes and injector heads as well as for wiping the surface under the plate carrier e Mild detergent e Clean lint free cotton cloths e Deionized or distilled water e Stylus stored in a plastic cylinder affixed to the rear of the dispense module or reader PN 2872304 For cleaning the optical probes e Clean cotton swabs e Isopropyl alcohol e Lens cleaning tissue BioTek Instruments Inc Cleaning the Internal Components 135 Removing the Reader s Shroud The Synergy HT s shroud cover must be removed to expose the internal components Caution Before removing the shroud Purgethe dispense module see page 129 for instructions and then turn off and disconnect the reader from its power supply the PC and the dispense module 1 Disconnect power and all cables Set the e
234. the reader 4 Click OK to save the settings and close this dialog The settings become available for selection in the Read step dialog in a Procedure BioTek Instruments Inc Gen5 Software 47 Creating Protocols and Experiments In Gen5 a Protocol contains instructions for controlling the reader and optionally instructions for analyzing the data retrieved from the reader At a minimum a protocol must specify the Procedure for the assay you wish to run After creating a protocol create an Experiment that references the protocol You ll run the experiment to read plates and analyze the data Figure 27 An Experiment containing measurement data based on a pre defined protocol Synergy HT Operator s Manual 48 Chapter 3 Getting Started The instructions below briefly describe how to create a simple protocol in Gen5 See Gen5 s Help system for complete instructions To create a Protocol in Gen5 1 Select File New Protocol 2 Open the Procedure dialog If prompted to select a reader select the Synergy HT and click OK 3 Add Steps to the procedure for shaking or heating the plate dispensing fluid reading the plate and more Click the Validate button to verify that the reader supports the defined steps and then click OK Tips e Add a Dispense step to define the volume and rate at which fluid will be dispensed and from which d
235. ther Errors 2A00 4000 Code 2A00 2A01 2B01 2B02 Description and Probable Causes XY axis movement did not find the middle sensor This error indicates old basecode Recommend upgrading This is indicating the middle sensor signal did not transition Home carrier to middle sensor This error indicates that the signal did not transition when homing the carrier to the middle sensor Error 0201 will occur during initialization where 2A01 occurs any other time Possible causes e Defective belt pulley dirty rail bushings bearings or motor e The carrier hit the top probe e The carrier was moved while the plate was being placed on the carrier causing the carrier to move off the middle sensor Syringe motor axis did not find the home opto sensor transition This error can indicate one of the following scenarios e A motor was not able to move to its home position as registered by feedback from an optical sensor e Prior to aspirating the syringe was not within the homing sensor To minimize the air bubble the syringe must be in the opto sensor Note This error can apply to either Syringe 1 or Syringe 2 Probable Causes e Linear way is dirty or needs lubrication e Defective or broken optical sensor e Syringe was not installed correctly or was not cleaned causing stress on syringe movement by not allowing the syringe to move to the home position e The glue between the lead screw and the motor has separated e C
236. thylumbelliferone MUB Top reading 0 31 ng ml solution of Methylumbelliferone in CBB 300 uL per well signal to noise ratio greater than 2 0 Excitation 360 40 Emission 460 40 Corning Costar black strips Optional Time Resolved Fluorescence e Delay 0 or 20 us to 16000 us e Integration interval 20 to 16000 us e Times adjustable in 10 us increments BioTek Instruments Inc Incubation Specifications 213 e Temperature control range from 4 over ambient to 50 C e Temperature variation 0 50 C across the plate 37 C 250 uL per well with the plate sealed Shake e Low Medium High and Variable shaking speeds e Shake duration is programmable by the user Injector Model The following specifications apply to Synergy HT models with injectors Plate Type Dispense Volume Range Accuracy Precision Dispenses to standard 6 12 24 48 and 96 well microplates with 128 x 86 mm geometry 5 1000 uL with a 5 20 uL tip prime Dispensing deionized water with 0 1 Tween 20 at room temperature 1 uL at 5 50 uL 2 at 51 1000 uL Dispensing a 200 uL solution of deionized water 0 1 Tween 20 and dye at room temperature lt 2 0 for volumes of 50 200 uL lt 4 0 for volumes of 25 49 uL lt 7 0 for volumes of 10 24 uL lt 10 0 for volumes of 5 9 uL Synergy HT Operator s Manual 214 Appendix D Specifications BioTek Instruments Inc Appendix E Instrument Dimensions for
237. to specification with liquid samples Liquid testing differs from testing with the Absorbance Test Plate in that liquid in the wells has a meniscus whereas the Test Plate s neutral density glass filters do not The optics characteristics may differ in these two cases thus alerting the operator to different types of problems e Liquid Test 1 confirms repeatability and alignment of the reader when a solution is used in the microplate If these tests pass then the lens placement and optical system cleanliness are proven e Liquid Test 2 can be used to test the alignment repeatability and accuracy of the reader if an Absorbance Test Plate is not available e Liquid Test 3 is provided for sites requiring proof of linearity at wavelengths lower than those attainable with the Absorbance Test Plate This test is optional because the reader has good front end linearity throughout its wavelength range For Liquid Tests 1 and 2 the tester is instructed to prepare the stock dye solution described on the next page The purpose of the formulation is to create a solution that absorbs light at 2 0 OD full strength when dispensed at 200 uL in a flat bottom microplate well Alternatively any solution that gives a stable color will suffice This includes substrates incubated with an enzyme preparation and then stopped with an acidic or basic solution Some enzyme substrate combinations that may be used as alternates to the described dye are shown b
238. tronic noise e There may be an ambient light leak Make sure the plate carrier door and the front hinged door are properly closed e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and are properly fastened e Order sorting filter wheel is jammed not aligning the filter wheel to block the light path or the filter is degraded and is not passing enough light energy 0911 0916 The absorbance measurement channel Dark current value failed See Dark on the system test See criteria in text below The last number is the lambda table position number This error can indicate one of the following scenarios e The measurement channel failed lt 100 during optic test with the flash on e The measurement channel failed lt 100 during a read or blank read not in sweep mode with the flash off e The measurement channel failed lt 100 or the Dark value has changed more than 10 from the last self test data during a read or blank read with the flash on Probable Causes e Absorbance analog PCB or measurement channel analog PCB is defective e Shielding of the cable between measurement channel and analog PCB is defective or disconnected e Measurement channel photodetector is defective or the optic spray is damaged Synergy HT Operator s Manual 184 Appendix C Error Codes Code 0A01 OA06 0A11 0A16 Description and Pro
239. truments and may not all be applicable to any single given reader Home Sensor Initial Find Errors 0100 0307 Code 0100 0101 0200 These errors occur when the optical sensor for the axis in question never electrically transitions from a high state to a low state The causes can range from a simple disconnected cable obstructed axis plate or shipping screw limiting travel or a defective sensor The last digit of the error identifies the axis Description and Probable Causes Abort Error This error indicates that the read or task has been aborted Abort Error This error indicates that the read or task has been aborted 0101 indicates that the abort was a software abort Possible Causes e User aborted read from Gen5 or KC4 e User aborted from another serial interface X axis motor did not find the home opto sensor transition This error indicates that a motor was not able to move to its home position as registered by feedback from an optical sensor Note This error does not look for the mid or XY sensor See 2A01 for mid sensor See 2400 for XY sensor Probable Causes e X axis rail is dirty where the nylon slider bushings are worn and causing too much friction or dirt in roller bearings causing bearings to jam e The support pin on the carrier has moved preventing it from properly sitting between the two roller bearings on the bearing block e Defective or broken optical sensor e Defective Motor
240. un via KC4 the dispense module should initialize automatically If for any reason the module does not initialize automatically you can initialize it from KC4 1 4 In KC4 select System Readers click the Configuration button and then click the appropriate Dispenser tab 1 or 2 Click the I nitialize button The syringe drive will move to its home position and its sensors will be verified Upon successful completion the Status should show Initialized Repeat for the other syringe Dispenser 2 Prime Utility Before running a Read amp Dispense protocol the dispense module and its associated tubing must be primed with the fluid to be used KC4 provides a special utility for this task To prime the dispense module 1 oie Tee g Fill the supply bottle with a sufficient volume of the fluid to be used for the prime and the assay Insert the appropriate inlet tube into the bottle I mportant Place the priming plate on the carrier In KC4 select System Reader Control and click the Dispenser tab Select the Dispenser 1 or 2 associated with the supply bottle Enter the Volume to be used for the prime from 5 to 5000 uL The minimum recommended prime volume is 1100 UL Select a prime Rate in uL second Click Prime to start the process When the process is complete carefully renove the priming plate from the carrier and empty its contents If the priming plate is empty the prime volume was too low
241. uorescence or Luminescence Select a Reading Type of Endpoint or Kinetic For a Kinetic protocol click the Kinetic button to define the Run Time and Interval If supported by the current Synergy HT model checking Time Resolved enables the Time Resolved Fluorescence feature Define the Filter Sets by selecting the filter s Optics Position and Sensitivity Select a Plate Type Enable Temperature Control and or Shaking if necessary Select any Pre Reading parameters Click OK to verify the parameters and return to the main screen Select Protocol Save As and give the protocol an identifying name BioTek Instruments Inc KC4 Software 55 To create a Fluorescence or Luminescence Dispense protocol in KC4 for the Synergy HTTR w Injectors 1 ee an 10 11 Select Data New Plate If prompted to select a protocol select Empty Protocol and click OK If not prompted select Protocol New Select Protocol Reading The Reading parameters dialog will appear Set Detection Method to Fluorescence or Luminescence Check the Dispense box Select a Reading Type of Endpoint or Kinetic Define the Filter Sets up to 2 by selecting the filter s Optics Position and Sensitivity Select a Plate Type Enable Temperature Control if necessary Select any Pre Reading parameters Click the Read amp Dispense button The Read amp Dispense dialog will appear gt Select a Read Mode of Well
242. up If you suspect that a problem occurred during shipment if you have received the equipment after returning it to the factory for service and or if regulatory requirements dictate that you qualify the equipment on a routine basis you should perform the procedures outlined in this chapter This chapter contains BioTek Instruments recommended Installation Qualification IQ Operational Qualification OQ and Performance Qualification PQ procedures for all models of the Synergy HT Multi Mode Microplate Reader A Product Qualification Package PN 7090521 for the Synergy HT is available for purchase The package contains complete procedures for performing Installation Qualification Operational Qualification Performance Qualification and Preventive Maintenance procedures Microsoft Excel spreadsheets are provided for performing the calculations and checklists data sheets and logbooks are provided for recording results Contact your local BioTek dealer for more information IQ 0Q PQ Installation Qualification confirms that the reader and its components have been supplied as ordered and ensures that they are assembled and configured properly for your lab environment e The recommended IQ procedure consists of setting up the instrument and its components as described in Chapter 2 Installation and performing the System Test For models with injectors a quick Injector Test is also performed to ensure that the dispense
243. upply PCB is defective Synergy HT Operator s Manual 194 Appendix C Error Codes Code 2901 2902 2903 2904 2905 2906 Description and Probable Causes Lamp reference voltage out of range 1459 1917 See self test Voltage Reference Lamp This tests the voltage across a sense resistor in series of the lamp It is monitoring the current through the lamp This test is performed when the instrument is first turned on and then tested periodically during background functions Probable Causes e The lamp is weak or defective change the lamp BioTek PN 7080500 e The sense resistor is damaged on the 7090402 PCB e The regulator mounted on the 7090402 PCB is defective e Verify the lamp is actually on or off Perform a self test This error can give false errors If the lamp is on disregard this error e Reference Field Change Notice L0045 24V reference voltage out of range 1769 2162 See self test Voltage Reference 24V This tests the voltage across a sense resistor in a series of the 24 volts it is monitoring the current This test is performed during self test Probable Causes e The motor power supply PCB is defective e The sense resistor is damaged on the motor power supply PCB 40 Volt Motor reference voltage out of range 2027 2069 See self test Voltage Reference Mtr This tests the voltage across a sense resistor in series of the 40 volts it is monitoring the current This test is p
244. urs and it is user replaceable The intensity of the bulb will slowly drop over time until the instrument s run time self check detects a low lamp current signal and Gen5 or KC4 displays an error message The lamp PN 7080500 should be replaced at this time Absorbance and Time Resolved Fluorescence The xenon flash lamp life is rated at an average of 1 billion flashes This bulb should outlive the useful life of the reader If there is a problem with the lamp however the intensity may drop and the run time self check will detect a low signal level and generate an error message If this happens the instrument will require service Contact BioTek for assistance this lamp is not user replaceable Synergy HT Operator s Manual 38 Chapter 3 Getting Started Excitation and Emission Filter Wheels All Synergy HT models are equipped with one Excitation filter wheel and one Emission filter wheel for use with fluorescence and luminescence measurements A monochromator is used for absorbance measurements A filter in the Excitation wheel selects the narrow band of light to which the sample will be exposed A filter in the Emission whed selects the band of light with the maximum fluorescence signal to be measured by the photomultiplier PMT Each filter wheel is labeled EX or EM and can contain up to four filters and or black plugs A filter can be used in either wheel but it must be oriented properly as described
245. w noise PMT detector through an empty filter position in the Emission filter wheel A filter can also be left in place if light filtering is necessary Absorbance measurements are made by switching to a xenon flash lamp and a monochromator for wavelength selection The use of a xenon flash lamp allows for both UV and visible light absorbance measurements The monochromator provides wavelength selection from 200 to 999 nm in 1 nm increments The Synergy HT has a 4 Zone temperature control from 4 C over ambient to 50 C that ensures superior temperature uniformity necessary for kinetic assays Internal plate shaking is also supported All Synergy HT models support the reading of 6 12 24 48 96 and 384 well microplates with standard 128 x 86 mm geometry Absorbance mode reads plates up to 0 8 20 3 mm in height fluorescence mode reads plates up to 1 25 31 75 mm Polymerase Chain Reaction PCR tubes up to 1 25 31 75 mm are also readable with the use of existing adapter plates The Time Resolved TR option allows time resolved fluorescence measurements by using the xenon flash light source in conjunction with the PMT measurement detector A special cartridge installed in the Excitation filter wheel location is required Models with injectors support dual reagent dispensing to 6 12 24 48 and 96 well microplates with standard 128 x 86 mm geometry An external dispense module pumps fluid from the supply bottles to t
246. with SF and the concentration Mix this SF solution with buffer 0 53 mL of 1 32 nM stock solution 13 47 mL 50 2 uM Synergy HT Operator s Manual 92 Chapter 4 Instrument Qualification Procedure 1 2 3 Create the Gen5 protocols see page 95 or KC4 protocols see page 96 Prepare the test solutions see page 91 Perform the tests using the Bottom optics e Pipette the solutions for the Corners Sensitivity and Linearity Tests into a clean 96 well quartz or glass bottom microplate see the map on page 94 e Read the plate using the Synergy HT FI_B prt protocol Perform the tests using the Top optics e Pipette the solutions for the Corners Sensitivity and Linearity Tests into a new 96 well solid black or quartz microplate see the map on page 94 e Read the plate using the Synergy HT FI_T prt protocol Save and or print the measurement data Calculate and analyze the results as described below Results Analysis Corners Test 1 Calculate the Mean of the twelve wells containing the 3 3 nM SF test solution A1 A3 A10 A12 H1 H3 H10 H12 2 Calculate the Standard Deviation of the same twelve wells 3 Calculate the CV Standard Deviation Mean 100 The CV must be less than 3 0 to pass Sensitivity Test 1 Calculate the Mean and Standard Deviation for the buffer wells C10 F12 2 Calculate the Mean for the 1000 pM 1 0 nM SF solution wells C1 F1 3 Calculate
247. wrong voltage request to the lamp power supply Synergy HT Operator s Manual 180 Appendix C Error Codes Absorbance Reader Noise Errors 0700 0A16 Typical noise data during the system self check is under 5 counts in both measurement and reference channels Failure of this test indicates basic instability The instrument should be returned for service to correct the problem Code Description and Probable Causes 0700 PMT Bias current offset See Bias current offset on self test Fail if gt 5 counts This error indicates that the PMT analog PCB noise level is elevated and is not able to compensate This is not affected by the PMT or PMT base The PMT gain is set to 0 Probable Causes e PMT analog board failed to compensate for elevated noise e Motor power supply PCB and PMT analog PCB not properly grounded defective connection e Motor power supply PCB is noisy causing the PMT analog PCB to be noisy Absorbance reference channel noise test at max gain See noise Delta on self test fail if lt 20 This error indicates significant variations in background electronic noise were detected when blocking the light and increasing the gain to maximum Probable Causes e Electrical noise may be penetrating the measurement chamber The bottom and top shrouds are part of the electrical shielding Verify that the shrouds are installed and properly fastened e The coaxial cable ground between the reference channel and the
248. xternal dispense module aside 2 Place the Synergy HT on a work surface that allows you to easily access all sides of the instrument 3 Remove four black Phillips head screws one at the bottom rear corner on each side and two at the top center of the rear panel Remove two screws from the top center of the rear panel P Note When reinstalling the shroud press down firmly on the top to maintain a good light seal while tightening the top screws Synergy HT Operator s Manual 136 Chapter 5 Preventive Maintenance 4 Stand facing the front of the instrument Grasp both sides of the shroud slide it toward you and pull it straight off the instrument Set the shroud aside BioTek Instruments Inc Cleaning the Internal Components 137 Removing the Internal Tubes and Injector Heads Take a moment to identify the components described in this section iilini ch oie ciate ee ON Internal tubes 2 black for delivering fluid to the injector heads Injector heads 2 white for dispensing fluid into microwells Cable clamp for holding the tubes in place Figure 37 Internal components for the injection system SYRINGE 1 SYRINGE 2 I mportant When reinstalling the i ite internal dispense tubes be sure to align the tubing ports with the injector heads as shown in this diagram Look for the SYRINGE 1 and SYRINGE 2 labas on the instrument s rear pane
249. ylactic gloves when handling contaminated instruments Gloved hands should be considered contaminated at all times keep gloved hands away from eyes mouth and nose Eating and drinking while decontaminating instruments is not advised Mucous membranes are considered prime entry routes for infectious agents Wear eye protection and a surgical mask when there is a possibility of aerosol contamination Intact skin is generally considered an effective barrier against infectious organisms however small abrasions and cuts may not always be visible Wear protective gloves when performing the decontamination procedure BioTek Instruments Inc Required Materials 155 Required Materials For all Synergy HT models e Sodium hypochlorite NaClO or bleach e 70 isopropyl alcohol as an alternative to bleach e Deionized or distilled water e Safety glasses e Surgical mask e Protective gloves e Lab coat e Biohazard trash bags e 125 ml beakers e Clean lint free cotton cloths Additional materials for models with injectors e Phillips head screwdriver e Small brush for cleaning the tip priming trough and priming plate e Optional Mild detergent Synergy HT Operator s Manual 156 Appendix A Decontamination Procedure for Models without Injectors The sodium hypochlorite bleach solution is caustic wear gloves and eye protection when handling the solution Do not immerse the instrument spray it with liquid or use
250. ylon screws into the Syringe 1 and 2 tubing ports on the rear of the reader Refer to the illustrations in 7 Unpack and Inspect the Dispense Module starting on page 17 when performing these steps 10 11 Remove the two inlet tubes from the syringe valves and store them in their plastic canisters Remove the two outlet tubes from the syringe valves Attach the clear plastic fitting covers to the fittings of the outlet tubes Place the tubes in a plastic bag Place the dispense module inside the inner shipping box Slide the cardboard shipping insert down around the module Pack the reagent bottle holders in bubble wrap and place them on top of the module Seal the box with tape Locate the original accessories shipping box and foam end caps Place the bottom foam end cap into the box Place the syringes the inlet tubes and the outlet tubes inside the cutouts of the bottom foam end cap in the accessories box Place the dispense module cover on top of the accessories Cover the accessories with the top foam end cap place the dispense module cable inside the top of the end cap and seal the box with tape Locate the original outer shipping box and foam end caps Insert the bottom foam end cap Lower the dispense module box into the end cap Insert the accessories box alongside the dispense module box Insert the top foam end cap Close and seal the outer box with tape Write the RMA number in large clear numbers on the outs

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