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

Chapter 7Maintaining the InstrumentPerforming a Background RunWhen to PerformPurpose ofBackground RunsAbout theBackgroundComponentMaterialsRequiredApplied Biosystems recommends running a background plate monthly or as often asnecessary depending on instrument use.A background run measures the ambient fluorescence in a background plate or LowDensity Array containing deionized water. During the run, the 7900HT instrumentperforms a continuous scan of the plate for 2 minutes at 60 °C. Afterwards, the SDSsoftware averages the spectrum recorded during the run and extracts the resultingspectral component to a calibration file. The software uses the calibration file duringsubsequent runs to remove the background signal from the run data.Because the background signal can change with instrument age, Applied Biosystemsrecommends regenerating the Background component calibration every month.Note: Background runs can also be used to detect and troubleshoot sample blockcontamination.Fluorescence collected by the Applied Biosystems 7900HT Fast Real-Time PCRSystem includes a background component, a fluorescent signal that is inherent to thesystem. The background component is a composite signal found in all spectral datathat consists of fluorescence from several sources including: the backgroundelectronic signal, the sample block, the water inside the consumable, and the plasticconsumable itself. Because the background signal can interfere with the precision ofSDS data, the 7900HT instrument has been engineered to minimize the backgroundsignal. Additionally, the SDS software also algorithmically eliminates thebackground signal from each fluorescent sample to maximize the instrument’ssensitivity.One of the following:• Background plate included in a Spectral Calibration Kit and a centrifuge withplate adapter• Components for making a standard 384- or 96-well background plate:– ABIPRISM 394- or 96-Well Optical Reaction Plate– An optical adhesive cover or ABI PRISM Optical Flat Caps– Pipettor, 100-µL (with pipet tips)– Centrifuge, with plate adapter• Components for making a TaqMan Low Density Array background plate:– Microfluidic card spectral calibration reagents– Centrifuge– Microfluidic card sealer7-16 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User GuideDRAFTSeptember 1, 2004 11:39 am, CH_Maintenance.fm