Using Passive Reference Dyes for Normalization and Troubleshooting in qPCR

Three statements are often made about what the use of a passive reference achieves with respect to normalizing non-PCR related fluorescence variation: It normalizes for pipetting errors; it normalizes for well-to-well optical variations on the instrument; it normalizes for changes in concentration or volume of reaction mix during the qPCR run. This technical note discusses the accuracy of these statements and explores other benefits of using passive reference dyes in qPCR.