qPCR - Calibration test of Opticon 2

Received new FAM calibration reagent. It comes pre-prepared in a 1x PCR buffer (0.3uM), however there is only enough for a single plate (use 50uL/well). Will run plate in Opticon 2. Run according to the calibration protocol in the Opticon 2 manual (p. 10-4).


Well, there’s actually a signal this time as opposed to the run on 20090806. However, it’s pretty clear that the signals aren’t even close to being uniform. Or, as the manual says “tightly clustered lines”. I’m also not sure why the fluorescence decreases over time, although it could simply be degradation of the fluorophore after being hit with light. I’ve sent the results to Bio-Rad for help interpreting them.