Analytical and Bioanalytical Chemistry

, Volume 401, Issue 10, pp 3221–3227 | Cite as

A standard additions method reduces inhibitor-induced bias in quantitative real-time PCR

  • Stephen L. R. Ellison
  • Kerry R. Emslie
  • Zena Kassir
Paper in Forefront

Abstract

A method of calibration for real-time quantitative polymerase chain reaction (qPCR) experiments based on the method of standard additions combined with non-linear curve fitting is described. The method is tested by comparing the results of a traditionally calibrated qPCR experiment with the standard additions experiment in the presence of 2 mM EDTA, a known inhibitor chosen to provide an unambiguous test of the principle by inducing an approximately twofold bias in apparent copy number calculated using traditional calibration. The standard additions method is shown to substantially reduce inhibitor-induced bias in quantitative real-time qPCR.

Keywords

Nucleic acids Quantitative PCR Calibration Standard additions 

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Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Stephen L. R. Ellison
    • 1
  • Kerry R. Emslie
    • 2
  • Zena Kassir
    • 2
  1. 1.LGC LimitedTeddingtonUK
  2. 2.National Measurement InstituteLindfieldAustralia

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