Development And Performance Evaluation of A Rapid In-House ELISA for Retrospective Serosurveillance of SARS-CoV-2

Background In the ongoing pandemic situation of COVID-19, serological tests can complement the molecular diagnostic methods, and can be one of the important tools of sero-surveillance and vaccine evaluation. Aim To develop and evaluate a rapid SARS-CoV-2 specific IgG ELISA with increased sensitivity and specificity. Methods In order to develop the ELISA, three panels of samples (n=184) have been used: panel 1 (n=19) and panel 2 (n=60) were collected from RT-PCR positive patients within 14 and after 14 days respectively following the onset of clinical signs of disease whereas panel 3 consisted of negative samples (n=105) collected either from healthy donors during pre-pandemic era, pandemic era or from RT-PCR negative healthy individuals. As a capturing agent full-length SARS-CoV-2 specific recombinant nucleocapsid was immobilized and blocked using blocking agent. In total of 30 samples from the panels have been tested with Elecsys Anti-SARS-CoV-2 for selecting positive and negative controls, as well as comparator assay. The threshold cut-off point, inter-assay and intra assay variations were determined. Results The assay time was set at a total of 30 minutes with the sensitivity of 84% (95% confidence interval, CI, 60.4%, 96.6%) and 98% (95% CI, 91.1%, 100.0%), for panel 1 and 2 respectively, with overall 94.9% sensitivity (95% CI 87.5%, 98.6%). Moreover, the clinical specificity is 97.1% (95% CI, 91.9%, 99.4%) with no cross-reaction with dengue sample. The overall positive and negative predictive values are 96.2% (95% CI 89.2%, 99.2%) and 96.2% (95% CI, 90.6% 99.0%) respectively. In-house ELISA demonstrated 100% positive and negative percent agreement with ROCHE (Elecsys; Anti-SARS-CoV-2), with a Cohens kappa value of 1.00 (very strong agreement), while comparing 13 positive and 17 negative confirmed cases. Conclusion The assay is rapid and can be applied as one of the early and retrospective sero-monitoring tools in all over the affected areas.