Performing A Pure Dye Run - Applied Biosystems 7900HT User Manual

Chapter 7 Maintaining the Instrument

Performing a Pure Dye Run

When to Perform
Purpose of Pure
Dye Runs
Components of
the Pure Dye
Spectra
7-20 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 performing spectral calibration:
• Every 6 months depending on instrument use
• After changing sample block formats (see
Always run a background plate before performing a Pure Dye
IMPORTANT!
calibration.
Pure dye data is generated from the results of a pure dye run in which the SDS
software collects spectral data from a set of dye standards during a 2-min hold at
60 °C. The software stores the spectral information for the pure dye standards in a
calibration file located in the SDS directory. After the run, the software extracts each
component dye spectrum from the collected data in the pure spectra run file.
Because the age and use of instrument components can affect pure
IMPORTANT!
spectra readings, Applied Biosystems recommends updating the pure spectra data
files once or twice annually depending on instrument use.
The Applied Biosystems 7900HT Fast Real-Time PCR System monitors fluorescent
signals generated by several dyes inside the preloaded pure dye plate. For the
standard 384-well, standard 96-well, or Fast 96-well pure dye plates, the dyes are:
FAM ™ , NED ™ , ROX ™
For the 7900HT System TaqMan Low Density Array Upgrade, the 7900HT
instrument monitors fluorescent signals generated by three dyes: FAM
ROX ™ . Your installation kit contains these dyes, and you introduce them into three
separate microfluidic pure dye cards to perform pure dye runs.
1 2 3 4
Figure 7-4 Standard Pure Dye Spectra of the 7900HT Instrument
The 7900HT instrument supports the detection of custom pure dyes (dyes
Note:
other than those provided by Applied Biosystems). To add custom pure dyes to the
Pure Dye set for your instrument, see
page 7-27.
DRAFT
, SYBR ® Green, TAMRA
5 6 7
"Adding Custom Dyes to the Pure Dye Set" on
page 7-6)
™ , TET ™ , and VIC ® .
Dye
1 FAM
2 SYBR Green
3 TET
4 VIC
JOE
5 NED
6 TAMRA
7 ROX
™ , VIC ® , and
Peak (nm)
~520
~520
~540
~550
~550
~570
~580
~610