Figure 4.

Optimization and sensitivity evaluation of the enhanced universal RT-qPCR assay to detect SARS-CoV-2 and SARS-CoV ORF3ab-E genes. Ten-fold serial dilution (1 × 10⁰ to 1 × 10⁸ per 3 μL) of RNA transcripts of SARS-CoV-2 (a), SARS-CoV (b), and Bat/SARSr-CoV (c) were processed in triplicates for RT-qPCR optimization. Each dilution was labeled with different colors to identify differences in the amplification curve, as shown in the figure. (d) Limit of detection in 20 repetitions using diluted RNAs (100, 20, 10, and 1 RNA copies per 3 μL). (e) RT-qPCR-positive amplification was determined from the mean cycle threshold value for each RNA dilution point. N.C: negative control; Ct: cycle threshold; SD: standard deviation; “-”: not determined.