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Updated: Nov 3, 2025
Author Spotlight: Advancements in Multiplex Detection of Respiratory Viruses
Published on: November 10, 2023
Development and validation of a new triplex real-time quantitative reverse Transcriptase-PCR assay for the clinical
Vijay J Gadkar1, David M Goldfarb1, Virginia Young2
1Department of Pathology & Laboratory Medicine, Division of Microbiology, Virology & Infection Control, BC Children's and Women's Hospital + Sunny Health Center, Vancouver, Canada; Department of Pathology & Laboratory Medicine, Faculty of Medicine, University of British Columbia, Vancouver, Canada.
Abstract:
To increase the repertoire of PCR based laboratory developed tests (LDTs) for the detection of SARS-CoV-2, we describe a new multiplex assay (SORP), targeting the SARS-CoV-2's, Spike and ORF8 genes. The widely used human RNaseP internal control was modified to specifically co-amplify the RNaseP mRNA. The SORP triplex assay was tested on a cohort (n = 372; POS = 144/NEG = 228) of nasopharyngeal flocked swab (NPFS) specimens, previously tested for the presence of SARS-CoV-2 using a PCR assay targeting E and RdRp genes. The overall sensitivity and specificity of the SORP assay was: 99.31% (95% CI: 96.22-99.98%), 100.0% (95% CI: 98.4-100%) respectively. The SORP assay could also detect a panel of variants of concern (VOC) from the B1.1.7 (UK) and B1.351 (SA) lineage. In summary, access to a repertoire of new SARS-CoV-2 LDT's would assist diagnostic laboratories in developing strategies to overcome some of the testing issues encountered during high-throughput SARS-CoV-2 testing.
