Validation of a qPCR method – Determining limit of detection, limit of quantification and dynamic range

Mikael Kubista 1, Robert Sjöback1, Jens Björkman1, Björn Sjögreen2, Lucas Luke Linz3, Amin Forootan2

1 TATAA Biocenter AB, Sweden
2 MultiD Analyses AB, Sweden
3 LGC Douglas Scientific, USA 

Quantitative Real-Time Polymerase Chain Reaction (qPCR) is the most sensitive and specific technique we have for the detection of nucleic acids. Even though it has been around for more than 30 years and is preferred in research applications, it has yet to win broad acceptance in routine practice. This requires a means to unambiguously assess the performance of specific qPCR analyses. Here we present methods to determine the limit of detection (LoD), the limit of quantification (LoQ) and the Dynamic Range as appli- cable to qPCR. These are based on standard statistical methods as recommended by regulatory bodies and adapted to the logarithmic response characteristic of qPCR.
The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments
SA Bustin, V Benes, JA Garson, J Hellemans, J Huggett, M Kubista, Clinical chemistry 55 (4), 611-622
Prime time for qPCR–raising the quality bar M Kubista Eur. Pharm. Rev. 19, 60-67

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