Applied Biosystems 7900HT Fast Real-Time PCR System and SDS ...

Chapter 7Maintaining the InstrumentRunning theVerification Plateor TGF-β CardVerifyingInstrumentPerformance1. In the plate document in the SDS software, select the Instrument tab.2. In the lower portion of the Instrument tab, select the Real-Time tab.3. If the instrument tray is inside the 7900HT instrument, click .The instrument tray rotates to the OUT position.4. Place the TaqMan RNase P Instrument Verification Plate or TGF-β card into theinstrument tray.Note: The A1 position is located in the top-left corner of the instrument tray.5. In the Real-Time tab, click . The 7900HT instrument begins the run.Note: Before starting the PCR run, the instrument may pause (up to 15 min) toheat the heated cover to the appropriate temperature.6. When the run is complete:a. Analyze the run data as explained on page 6-12.b. Set the baseline and threshold values for the analyzed data as explained onpage 6-46.c. Verify the performance of the 7900HT instrument as explained below.Analyzing Data from RNase P RunsThe install specification of the Applied Biosystems 7900HT Fast Real-Time PCRSystem demonstrates the ability to distinguish between 5,000 and 10,000 genomeequivalents with a 99.7% confidence level for a subsequent sample run in a singlewell.The following equation verifies the 7900HT instrument install specifications:[( CopyUnk 1)–3( σ CopyUnk1)] > [( CopyUnk 2)–3( σ CopyUnk2)]where:• CopyUnk 1 is the Average Copy Number of Unknown #1 (10,000 replicatepopulation)• σ CopyUnk1 is the Standard Deviation of Unknown #1 (10,000 replicate population)• CopyUnk 2 is the Average Copy Number of Unknown #2 (5000 replicatepopulation)• σ CopyUnk2 is the Standard Deviation of Unknown #2 (5000 replicate population)Note: The values above can be obtained from the experimental report window.Note: Up to six wells from each replicate group in a 96-well TaqMan RNase PInstrument Verification Plate (both standard 96-well and Fast 96-well plates) can beignored to meet specifications.Note: Up to 10 wells from each replicate group in a 384-well TaqMan RNase PInstrument Verification Plate can be ignored to meet specifications.7-34 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User GuideDRAFTSeptember 1, 2004 11:39 am, CH_Maintenance.fm