Analytical Performance Evaluation of Three Commercial Rapid Nucleic Acid Assays for SARS-CoV-2

Purpose: To cope with the SARS-CoV-2 epidemic, several rapid nucleic acid assays have been approved for use, but the analytical performance has not been well evaluated. In this report, two key performance parameters, analytical sensitivity (limit of detection) and reproducibility, of three approved rapid nucleic acid assays were assessed using heat-inactivated SARS-CoV-2 culture supernatants quantified by digital PCR.

Methods: The LOD (limit of detection) and reproducibility of three approved rapid nucleic acid assays using their own instruments were assessed, while the LOD and reproducibility of two assays on a 7500 Real-Time instrument were assessed at the same time.

Results: Using their own instruments, 100% of samples with 1150 copies/mL viral RNA could be detected by the Da An and Coyote assays, while 90% of samples could be detected by the Ustar assay; yet, for 525 copies/mL and 287.5 copies/mL viral RNA, the detection rate of the Ustar assay was higher than that of either the Da An or Coyote assays. However, the three assays did not produce statistically significant results with the three different concentrations of viral RNA (P=0.46, 0.46 and 0.46). Using a 7500 Real-Time instrument, Da An and Coyote assays did not produce statistically significant results with the 1150, 525 and 287.5 copies/mL viral RNA (P>0.99, >0.99 and >0.99). The positive and negative detection rates of the three assays in the intra- and inter-assay stages were 100% on both their own instruments and the 7500 real-time PCR instrument.

Conclusion: Positive or strongly positive samples can be detected by the rapid nucleic acid assay, but the analytical performance should be optimized, and comprehensive evaluations are also required.