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Calibrate the ViiA

™

7 System

Applied Biosystems ViiA

™

7 Real-Time PCR System Quick Reference Guide

Calibrate the ViiA

™

7 System

Calibration types

• Regions of interest (ROI)

• Background

• Uniformity

• Dye (only for dyes used in your experiments)

• Normalization

• Instrument verification

Materials required

384-well sample block

Array-card sample block

96-well sample block

• 384-Well Spectral Calibration Plate with FAM

™

Dye (PN 4432271)

• 384-Well Spectral Calibration Plate with VIC

Dye (PN 4432278)

• 384-Well Spectral Calibration Plate with ROX

™

Dye (PN 4432284)

• 384-Well Spectral Calibration Plate with SYBR

Green Dye (PN 4432290)

• 384-Well Spectral Calibration Plate with TAMRA

™

Dye (PN 4432296)

• 384-Well Spectral Calibration Plate with NED

™

Dye (PN 4432302)

• 384-Well Region of Interest (ROI) and Background Plates (PN 4432320)

• 384-Well Normalization Plates with FAM

™

/ROX

™

and VIC

/ROX

™

Dyes

(PN 4432308)

• TaqMan RNase P 384-Well Instrument Verification Plate (PN 4455280)

• ViiA

™

7 System 384-Well Plate Adapter

(PN 4457087)

• Centrifuge with plate adapter

• Powder-free gloves

• Safety goggles

• Array Card Spectral Calibration Dye Kit (PN 4432314)

• TaqMan

RNase P Array Card Instrument Verification Reagents Kit

(PN 4432265)

• ViiA

™

7 System Array Card Adapter

(PN 4454166)

• Centrifuge with centrifuge array-card carrier

clips and buckets

• Powder-free gloves

• Safety goggles

• 200-µL pipettor, with pipette tips

• Array-card staker/sealer fixture

• 96-Well Spectral Calibration Plate with FAM

™

Dye (PN 4432327)

• 96-Well Spectral Calibration Plate with VIC

Dye (PN 4432334)

• 96-Well Spectral Calibration Plate with ROX

™

Dye (PN 4432340)

• 96-Well Spectral Calibration Plate with SYBR

Green Dye (PN 4432346)

• 96-Well Spectral Calibration Plate with TAMRA

™

Dye (PN 4432352)

• 96-Well Spectral Calibration Plate with NED

™

Dye (PN 4432358)

• 96-Well Region of Interest (ROI) and Background Plates (PN 4432364)

• 96-Well Normalization Plates with FAM

™

/ROX

™

and VIC

/ROX

™

Dyes

(PN 4432370)

• TaqMan

RNase P 96-Well Instrument Verification Plate (PN 4432382)

• ViiA

™

7 System 96-Well Plate Adapter (PN

4459845)

• Centrifuge with plate adapter

• Powder-free gloves

• Safety goggles

Содержание ViiA 7 Real

Страница 1: ...Maintain the ViiA 7 System 15 Power off the ViiA 7 System 22 For details on operating and maintaining your ViiA 7 System refer to the Applied Biosystems ViiA 7 Real Time PCR System User Guide PN 4442661 Description The Applied Biosystems ViiA 7 Real Time PCR System uses fluorescent based polymerase chain reaction PCR reagents to provide Quantitative research detection of target nucleic acid sequen...

Страница 2: ...55280 ViiA 7 System 384 Well Plate Adapter PN 4457087 Centrifuge with plate adapter Powder free gloves Safety goggles Array Card Spectral Calibration Dye Kit PN 4432314 TaqMan RNase P Array Card Instrument Verification Reagents Kit PN 4432265 ViiA 7 System Array Card Adapter PN 4454166 Centrifuge with centrifuge array card carrier clips and buckets Powder free gloves Safety goggles 200 µL pipettor...

Страница 3: ...on mix from the freezer then allow it to thaw to room temperature in the dark approximately 5 minutes IMPORTANT Keep the calibration mix out of the light The fluorescent dye in the mix is photosensitive Prolonged exposure to light can diminish the fluorescence of the dye b Fill the array card with calibration mix For a normalization calibration fill two cards with separate normalization solutions ...

Страница 4: ...t IMPORTANT Plates should be loaded and unloaded by trained operators who have been warned of the moving parts hazard 4 Start the run a In the Setup tab select Please check the box when the calibration plate has been loaded then click Next b In the Run tab click START RUN to start the calibration IMPORTANT Do not attempt to open the access door duing the run The door is locked while the ViiA 7 Sys...

Страница 5: ...ck Reaction mix components For 384 well 96 well or Fast 96 well sample blocks Centrifuge with plate adapter Plate or 96 well and Fast 96 well blocks only tube strip or single tube Optical adhesive film For array card sample blocks Centrifuge with centrifuge array card carrier clips and buckets Array card staker sealer fixture Array card Workflow Design the experiment page 6 Prepare the reactions p...

Страница 6: ...reen All experiments except Genotyping Define Targets Genotyping Define SNPs All experiments Define Samples Optional Standard Curve Relative Standard Curve and Comparative CT Define Biological Replicate Groups All experiments except Presence Absence Select the Passive Reference dye Relative Standard Curve and Comparative CT Select the Reference Sample and the Endogenous Control 5 Assign targets an...

Страница 7: ...on change the default analysis settings Start Cycle and End Cycle in Preferences c For cycling stages Edit the number of cycles Enable or disable AutoDelta For an AutoDelta step enter the Starting Cycle d For a melt curve stage select the ramp increment Step and Hold Click the Step and Hold field select the minutes or seconds then use the up or down arrow keys or click the up or down buttons in th...

Страница 8: ...ons Mix the master mix thoroughly by swirling the bottle Resuspend the assay mix by vortexing then centrifuge the tube briefly Thaw any frozen samples resuspend them by vortexing then centrifuge the tubes briefly Verify that the liquid is at the bottom of each well of the plate If not centrifuge the plate again at a greater rpm and for a longer time IMPORTANT Do not allow the bottom of the plate t...

Страница 9: ... between the film and the entire surface of the reaction plate e While using the applicator to hold the edge of the film in place grasp one end of the end tab and pull up and away sharply Repeat this step for the other end tab f To ensure a tight evaporation free seal repeat step d While applying firm pressure run the edge of the applicator along all four sides of the outside border of the film No...

Страница 10: ...Note The fill port is the larger of the two holes on the left arm of each fill reservoir IMPORTANT Do not allow the tip to contact and possibly damage the coated foil beneath the fill port 3 Pipette fluid into the fill reservoir Dispense the fluid so that it sweeps in and around the fill reservoir toward the vent port Pipette fluid into the fill reservoir but do not go past the first stop of pipet...

Страница 11: ...Do not try to save time by doing one spin for 2 minutes Centrifuging twice is important for a good fill into the array card d Open the centrifuge and check that the fluid levels in the reservoirs of the array card have decreased by the same amount Also check for bubbles in all wells If necessary centrifuge the array card for an additional minute to fill unfilled wells Do not exceed three 1 minute ...

Страница 12: ...ting properly on the sample block and affect amplification IMPORTANT Keep the filled array card out of the light The fluorescent dye in the array card is photosensitive Prolonged exposure to light can diminish the fluorescence of the dye Start and monitor the experiment 1 Load the plate or array card a Touch on the instrument touchscreen to eject the plate adapter b Place the plate or array card o...

Страница 13: ...wder free gloves reach into the ViiA 7 System and remove the plate from the heated cover then close the instrument door e Perform a background calibration to confirm that the sample block has not been contaminated View analysis reports Plot Applicable experiment type Purpose Amplification All experiment types if the experiment includes amplification To set baseline and threshold To compare the dat...

Страница 14: ...n separate files for each data type d Select the contents of the file Include any of the following information in your export file Sample setup data Raw data Amplification data Multicomponent data Results e Depending on the file type enter or select export file properties name type and location 2 Start the export a Optional Save export settings Select the Save Export Set As check box and name the ...

Страница 15: ...y archive or back up your experiment files Power off the computer that controls the instrument wait 30 seconds then power on the computer Clean the surface of the instrument with a lint free cloth IMPORTANT Do not use organic solvents to clean the ViiA 7 System Perform a ViiA 7 System self test Monthly Check the lamp status If necessary replace the tungsten halogen lamp Perform a background calibr...

Страница 16: ...self If you suspect a problem contact an Applied Biosystems Service Representative CAUTION PHYSICAL INJURY HAZARD During instrument operation the sample block can be heated to 100 C Before performing the following procedure be sure to wait until the sample block reaches room temperature CAUTION Before using a cleaning or decontamination method other than those recommended by the Applied Biosystems...

Страница 17: ...eionized water into each contaminated well b Pipette the water up and down several times to rinse the well then pipette the water to a waste beaker c Using a cotton swab scrub inside of each contaminated well d Using a lint free cloth absorb the excess deionized water 3 Install the sample block a Load the sample block into the ViiA 7 System IMPORTANT After installing the sample block confirm that ...

Страница 18: ...lls again step 2 to rinse the wells of the sample block e Perform a background calibration to verify that you have eliminated the contamination If the contamination remains remove the sample block and clean the contaminated wells step 2 then clean the contaminated wells again using 10 bleach solution a Pipette a small volume of 10 bleach solution into each contaminated well b In each contaminated ...

Страница 19: ...on order When you are instructed to perform calibrations in the following sections perform them in this order 1 ROI calibration 2 Background calibration 3 Uniformity calibration 4 Dye calibration 5 Normalization calibration Change the sample block IMPORTANT If you are installing a sample block of a different format you must also change the heated cover and plate adapter to match the new consumable...

Страница 20: ...ment Properties pane The installation is successful if the instrument powers on and if the Block Type field displays the correct type of sample block 4 Perform calibrations Perform calibrations in the order shown in A note on calibration order on page 19 Change the plate adapter IMPORTANT If you are installing a plate adapter of a different format you must also change the heated cover and sample b...

Страница 21: ...new heated cover into the instrument IMPORTANT When the heated cover is seated correctly the arrows on the front handle align as shown below If the arrows do not align push the heated cover further into the ViiA 7 System until the handle locks into place b Close the access door c Plug in and power on the instrument 3 Verify success a In the ViiA 7 Software select Instrument Instrument Console b In...

Страница 22: ... not blank touch to place the ViiA 7 System into stand by mode b Toggle the power button on the rear of the ViiA 7 System 2 Power off the ViiA 7 System computer a In the desktop select Start Shut Down b In the Shut Down Windows dialog box select Shut Down then click OK c If necessary Power off the monitor For Research Use Only Not intended for any animal or human therapeutic or diagnostic use LIMI...

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