ESTIMATION OF DELTA-CQ VALUES WITH CONFIDENCE FROM QPCR DATA

摘要

The invention describes how to estimate delta-Cq values from measured (raw-) Cq values gained from PCR measurements and how to calculate confidence intervals for them. This is realized by the following processing steps: A noise model, which might be constructed on some training PCR data, calculates the distribution of the true target material concentration of a single well for an observed measurement results. Said distribution is calculated for all types of measurement results including "Numeric" raw-Cq values as well as Cq being "Undetected", which denotes that no fluorescence signal was detected during all cycles and thus corresponds to no or very few target molecules. Based on this distribution information from several replicate wells of a gene of interest and several replicate wells of one or more reference genes the distribution of the true delta-Cq value is calculated. In order to come out with a finite variance of the delta-Cq value (which so far would not be the case if all replicates are "Undetected") a lower bound is defined on the delta-Cq value. This lower bound is chosen so small that no useful information about the clinical implications is lost but so large that even for samples with low total material yield a reliable estimation of the delta-Cq value is possible.