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Table 3 Details of the six quality controls used in the real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay

From: Sensitivity and specificity of a real-time reverse transcriptase polymerase chain reaction detecting feline coronavirus mutations in effusion and serum/plasma of cats to diagnose feline infectious peritonitis

 

quality controls

goal of quality controls

1

PCR positive controls (quantitatively¸ using synthetic DNA covering the real-time PCR target region (Integrated DNA Technologies IDT, Coralville, IA, USA))

functionality of PCR test protocols

2

PCR negative controls (PCR-grade nuclease free water)

absence of contamination in reagents

3

negative extraction controls (extraction positions filled with lysis solution and PCR-grade nuclease free water only)

absence of cross-contamination during the extraction process

4

RNA pre-analytical quality control targeting feline ssr rRNA (18S rRNA) gene complex

quality and integrity of the RNA as a measure of sample quality

5

a swab-based environmental contamination monitoring control

absence of contamination in laboratory

6

spike-in internal positive control (using lambda phage DNA)

absence of PCR inhibitory substances as a carryover from sample matrix