Hospital Stool Panel RT-PCR Kit

The EAEC PCR Kit has been tested on Roche LightCycler 480II/480 z, Agilent Technologies Mx3005P, Applied Biosystems ABI 7500, Bio-Rad CFX96, and QIAGEN Rotor-Gene Q real-time PCR instruments.  The Hospital Stool Panel RT-PCR is a multiplex real-time PCR assay for the direct, qualitative detection and differentiation norovirus (genogroup I and II), rotavirus and C. difficile toxin genes A (tcdA) / B (tcdB) in human stool samples. Detection takes place in one-step real-time RT-PCR format where the reverse transcription and PCR takes place in the same reaction tube. The isolated RNA (norovirus, rotavirus) is transcribed into cDNA by a reverse transcriptase. Subsequently amplification of the gene fragments specific for norovirus (ORF1/ORF2 junction region), rotavirus (NSP3), and C. difficile toxin A/B (tcdA/TcdB) occurs if present. The amplified targets are detected with hydrolysis probes, which are labeled at one end with a quencher and at the other end with a fluorescent reporter dye (fluorophore). In the presence of a target, the probes hybridize to the amplicons. During the extension step, the Taq-polymerase breaks the reporter-quencher proximity. The reporter emits a fluorescence signal which is detected by the optical unit of a real-time PCR instrument. The fluorescence signal increases with the amount of formed amplicons. The Hospital Stool Panel RT-PCR assay contains an Internal Control RNA (ICR) as an internal control of the sample preparation procedure and to determine possible PCR inhibition.