Performing A Background Run - Applied Biosystems 7900HT User Manual

Chapter 7 Maintaining the Instrument

Performing a Background Run

When to Perform
Purpose of
Background Runs
About the
Background
Component
Materials
Required
7-16 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User Guide
September 1, 2004 11:39 am, CH_Maintenance.fm
Applied Biosystems recommends running a background plate monthly or as often as
necessary depending on instrument use.
A background run measures the ambient fluorescence in a background plate or Low
Density Array containing deionized water. During the run, the 7900HT instrument
performs a continuous scan of the plate for 2 minutes at 60 °C. Afterwards, the SDS
software averages the spectrum recorded during the run and extracts the resulting
spectral component to a calibration file. The software uses the calibration file during
subsequent runs to remove the background signal from the run data.
Because the background signal can change with instrument age, Applied Biosystems
recommends regenerating the Background component calibration every month.
Note: Background runs can also be used to detect and troubleshoot sample block
contamination.
Fluorescence collected by the Applied Biosystems 7900HT Fast Real-Time PCR
System includes a background component, a fluorescent signal that is inherent to the
system. The background component is a composite signal found in all spectral data
that consists of fluorescence from several sources including: the background
electronic signal, the sample block, the water inside the consumable, and the plastic
consumable itself. Because the background signal can interfere with the precision of
SDS data, the 7900HT instrument has been engineered to minimize the background
signal. Additionally, the SDS software also algorithmically eliminates the
background signal from each fluorescent sample to maximize the instrument's
sensitivity.
One of the following:
• Background plate included in a Spectral Calibration Kit and a centrifuge with
plate adapter
• Components for making a standard 384- or 96-well background plate:
– ABI P ™ 394- or 96-Well Optical Reaction Plate
RISM
– An optical adhesive cover or ABI P
– 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 sealer
DRAFT
™ Optical Flat Caps
RISM