Clinical evaluation of a multiplex real-time RT-PCR assay for detection of SARS-CoV-2 in individual and pooled upper respiratory tract samples

The aim of this study was to identify and validate a sensitive, high-throughput, cost-effective SARS-CoV-2 real-time RT-PCR assay be used as surveillance diagnostic tool for in university program. We conducted side-by-side clinical evaluation newly developed multiplex (EZ-SARS-CoV-2 Real-Time RT-PCR) with the commercial TaqPath COVID-19 Combo Kit, which has an Emergency Use Authorization from FDA. EZ-SARS-CoV-2 incorporates two assays targeting N gene, internal control human RNase P PCR inhibition single reaction. Nasopharyngeal (NP) anterior nares (AN) swabs were tested individuals pools both ABI 7500 Fast QuantStudio 5 detection platforms. analytical sensitivity 250 copies/ml or approximately 1.75 genome copy equivalents per performance evaluated using NP AN samples other laboratories. ranged between 94 96% across platforms, specificity 94.06%. positive predictive value 94%, negative 96%. Pooling five specimens yielded 93% sensitivity. overall agreement these high, supported by Cohen’s kappa 0.93. attributes high sample matrix pooled upper respiratory support its use high-throughput testing