Labonnet Ltd, 2 Hamelacha St, Ramat-Hasharon 47445, Israel.
Nucleic Acids Res. 2012 Feb;40(4):1395-406. doi: 10.1093/nar/gkr778. Epub 2011 Oct 19.
Quantitative real-time PCR (qPCR) is the method of choice for specific and sensitive quantification of nucleic acids. However, data validation is still a major issue, partially due to the complex effect of PCR inhibition on the results. If undetected PCR inhibition may severely impair the accuracy and sensitivity of results. PCR inhibition is addressed by prevention, detection and correction of PCR results. Recently, a new family of computational methods for the detection of PCR inhibition called kinetics outlier detection (KOD) emerged. KOD methods are based on comparison of one or a few kinetic parameters describing a test reaction to those describing a set of reference reactions. Modern KOD can detect PCR inhibition reflected by shift of the amplification curve by merely half a cycle with specificity and sensitivity >90%. Based solely on data analysis, these tools complement measures to improve and control pre-analytics. KOD methods do not require labor and materials, do not affect the reaction accuracy and sensitivity and they can be automated for fast and reliable quantification. This review describes the background of KOD methods, their principles, assumptions, strengths and limitations. Finally, the review provides recommendations how to use KOD and how to evaluate its performance.
实时荧光定量 PCR（qPCR）是核酸特异性和灵敏定量的首选方法。然而，数据验证仍然是一个主要问题，部分原因是 PCR 抑制对结果的复杂影响。如果未检测到 PCR 抑制，可能会严重影响结果的准确性和灵敏度。通过预防、检测和校正 PCR 结果来解决 PCR 抑制问题。最近，出现了一种新的计算方法家族，称为动力学异常值检测（KOD），用于检测 PCR 抑制。KOD 方法基于将一个或几个描述测试反应的动力学参数与描述一组参考反应的参数进行比较。现代 KOD 可以通过仅半个循环的扩增曲线偏移来检测 PCR 抑制，特异性和灵敏度>90%。这些工具仅基于数据分析，补充了改进和控制分析前阶段的措施。KOD 方法不需要劳动力和材料，不影响反应的准确性和灵敏度，并且可以自动化进行快速可靠的定量。本文综述了 KOD 方法的背景、原理、假设、优缺点。最后，本文提供了如何使用 KOD 以及如何评估其性能的建议。
