Robust CT-Prediction Algorithm for RT-PCR

Abstract

Real-time PCR is being used increasingly as the method of choice for
mRNA quantification, allowing rapid analysis of gene expression from
low quantities of starting template. 2009 pandemics increased the need
for efficient diagnostic tools that can provide rapid results for a
number of pathogens without compromising sensitivity and
specificity. Introduction of fluorescence-based Real-Time PCR (RT-PCR)
is increasingly used to detect multiple pathogens simultaneously and
rapidly by gene expression analysis of PCR amplification data.

Real-time PCR data are analyzed after setting an arbitrary threshold
that must intersect the signal curve in its exponential phase. The
point at which the curve crosses the threshold is called Threshold
Cycle (CT) This simple and arbitrary however not an elagant definition
of CT value sometimes leads to conclusions that are either false
positive or negative. Therefore, the purpose of this paper is to
present a stable and consistent alternative approach for the
definition and determination of CT value that leads to near zero false
positives and negatives.