The article describes a novel method for real-time quantitative reverse transcriptase polymerase chain reaction (QC RT-PCR) using the 5' nuclease assay. The method involves reverse transcription, amplification, detection, and quantitation of cystic fibrosis transmembrane transductance regulator (CFTR) target mRNA. A fluorogenic probe is designed to detect the CFTR amplicon, and an internal control template with the same primer sequences but a different internal sequence is used for normalization. The internal control probe hybridizes to the internal control amplicon. The method provides a high-throughput format for QC RT-PCR, allowing for rapid and accurate quantitation of target mRNA. The article details the design and validation of the internal control, the use of a sequence detector for real-time monitoring of fluorescence emission, and the precision and dynamic range of the assay. The method is demonstrated to be sensitive, accurate, and suitable for quantifying large numbers of samples in a short time.The article describes a novel method for real-time quantitative reverse transcriptase polymerase chain reaction (QC RT-PCR) using the 5' nuclease assay. The method involves reverse transcription, amplification, detection, and quantitation of cystic fibrosis transmembrane transductance regulator (CFTR) target mRNA. A fluorogenic probe is designed to detect the CFTR amplicon, and an internal control template with the same primer sequences but a different internal sequence is used for normalization. The internal control probe hybridizes to the internal control amplicon. The method provides a high-throughput format for QC RT-PCR, allowing for rapid and accurate quantitation of target mRNA. The article details the design and validation of the internal control, the use of a sequence detector for real-time monitoring of fluorescence emission, and the precision and dynamic range of the assay. The method is demonstrated to be sensitive, accurate, and suitable for quantifying large numbers of samples in a short time.