Fig. 2.
Optimal primer and probe annealing temperature determination for the ddPCR assay using a temperature gradient PCR. The highest amplitude of fluorescence and the best separation between bands, positive for wild-type (not shown) or mutant (green droplets) and negative (gray lowermost threshold), was observed in column D06, wherein annealing temperature was 58,1 °C. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)