Fig. 3: Optimisation of heat-inactivation conditions for SARS-CoV-2 hid-RT-PCR.

a C_T values for aliquots of nasopharyngeal swab samples inactivated at different temperature and time conditions, shown on absolute scale (left) and C_T change relative to the 95 °C 5 min condition (right). P-values calculated as two-tailed Wilcoxon signed rank tests. *FDR corrected for multiple testing. Median values shown in red. b Heatmaps of ranks based on absolute C_T values for nasopharyngeal swab samples (n = 20) with or without addition of polyvinylsulfonic acid (PVSA) and/or ethylenediaminetetraacetic acid (EDTA) before performing heat inactivation at 95 °C for 5 min. For EDTA-containing samples, equimolar supplement of MgCl₂ was added to the RT-PCR reaction as indicated. The same samples were evaluated by RT-qPCR repeatedly at different number of days stored at 4 °C after the heat activation. All conditions are listed in Table 2. c Boxplots of C_T values relative to the 95 °C condition without additives, ordered by the rank in (b). Center lines denote the median, hinges denote IQR and whiskers denote outlier points at maximum 1.5 × IQR. d Change in rank over days of storage at 4 °C. Colors and order same as in (b, c). Annotations refer to samples stored for 0 days. e Change in C_T values over days stored at 4 °C across different conditions. Thick line and ribbon indicate fitted LOESS curve and ±95% confidence interval, respectively. The dashed line indicates the 95 °C (without additive) condition.