Real-Time qPCR/qRT-PCR Methods: Standard Curves

Calibration curves for real-time PCR.

BACKGROUND Despite the increasing use of real-time PCR in the diagnosis and management of viral infections, there are no published studies adequately addressing the optimum number of calibrators, the number of replicates of each calibrator, and the frequency with which calibration needs to be repeated. This study was designed to address these issues. METHODS Cycle threshold data (ABI 7700) wa...

Robust regression methods for real-time PCR

Calibration-free assays on standard real-time PCR devices

Quantitative Polymerase Chain Reaction (qPCR) is one of central techniques in molecular biology and important tool in medical diagnostics. While being a golden standard qPCR techniques depend on reference measurements and are susceptible to large errors caused by even small changes of reaction efficiency or conditions that are typically not marked by decreased precision. Digital PCR (dPCR) tech...

Quantitative real-time RT-PCR using hybridization probes and imported standard curves for cytokine gene expression analysis.

Quantitative real-time or kinetic RT-PCR is increasingly used for the quantification of specific mRNA targets, especially in clinical applications. To quantify the mRNA of cytokines and their receptors, which play important roles in the pathogenesis of autoimmune diseases such as multiple sclerosis, we have developed quantitative two-step RT-PCR assays for IL-4, IL-4R, IFN-gamma, IFN-beta, and ...

DAG Expression: High-Throughput Gene Expression Analysis of Real-Time PCR Data Using Standard Curves for Relative Quantification

BACKGROUND Real-time quantitative PCR (qPCR) is still the gold-standard technique for gene-expression quantification. Recent technological advances of this method allow for the high-throughput gene-expression analysis, without the limitations of sample space and reagent used. However, non-commercial and user-friendly software for the management and analysis of these data is not available. RES...