Pushing beyond specifications: Evaluation of linearity and clinical performance of a fully automated SARS-CoV-2 RT-PCR assay for reliable quantification in blood and other materials outside recommendations

1 Background The ongoing SARS-CoV-2 pandemic presents a unique challenge to diagnostic laboratories. There are preliminary studies correlating qRT-PCR results from different materials to clinical outcomes, yet, comparability is limited due to the plethora of different assays used for diagnostics. In this study we evaluate clinical performance and linear range for the SARS-CoV-2 IVD (cobas6800/8800 system, a fully automated sample-to-result platform) in different clinically relevant matrix materials outside official specifications. Methods Assay performance was assessed in human plasma, BAL/BL and transport medium following chemical inactivation. For analytical evaluation, respective matrix materials were spiked with SARS-CoV-2 RNA in ten-fold dilution series. The efficacy of chemical inactivation by guanidine hydrochloride solution was confirmed in cell culture infectivity experiments. For correlation, a total of 235 predetermined clinical samples including respiratory swabs, plasma and BAL/BL were subjected to the SARS-CoV-2 IVD test and results were compared. Results The SARS-CoV-2 IVD showed excellent linearity over five to seven log steps depending on matrix material. Chemical inactivation resulted in a reduction in plaque forming units of at least 3.5 log steps, while having no significant impact on assay performance. Inter-run consistency from three different testing sites demonstrated excellent comparability of RT-PCR results (maximum deviation was 1.53 CT). Clinical evaluation for respiratory swabs showed very good agreement with the comparator assay (Positive agreement 95.7%, negative agreement 98.9%). Conclusion The SARS-CoV-2 IVD test for the cobas6800/8800 systems offers excellent linear range and inter-run consistency for quantification of SARS-CoV-2 RNA in different matrices outside official specifications. Highlights Effective reduction of SARS-CoV-2 infectivity by chemical inactivation without affecting assay performance. SARS-CoV-2 IVD for the cobas 6800/8800 is linear over up to seven log steps in different materials including human plasma. Minimal variance of CT values between testing sites indicates high comparability of quantification results.