Replicates of IPC's

[qPCR user]

Our *calibrator *sample was also *replicated *(see file), to average RT step and qPCR step within each plate. So we have 3 RTs and 2 qPCR cal samples to use as interplate calibrator. Is that ok, or should we average the calibrator values first ourself and load only one cal value ?

[Anders Bergkvist]

Do you not have qPCR and RT replicates of the experiment samples? Only for the calibrator samples? What is the calibrator samples intended to calibrate for? Our experience is that interplate calibrators are intended to calibrate for differences in instrument settings between runs on different plates. In this case the interplate calibrators should be as identical as possible, usually cDNA. With these calibrators you use the "Normalization with Interplate Calibrators" item in the GenEx pre-processing menu. If your calibrators have another purpose you can have qPCR and RT replicates and then compensate for these calibrators by using the "Normalize with reference sample(-s)" item in the GenEx menu.