Surveillance of Immunological Status after Vaccination by two Serological Assays based on SARS-CoV-2 Spike Protein

Purpose: Two serological assays, an Enzyme-Linked Immunosorbent Assay (ELISA) and a Lateral Flow Assay (LFA), have been developed based on the SARS-CoV-2 recombinant Receptor Binding Domain (RBD-ELISA) and the combination of Trimeric Spike (S) and Nucleoprotein (N), S-LFA and N-LFA, respectively, as candidate tools for both indirect measurement of virus circulation and assessment of infection and vaccine-induced immunity. Methods & Materials: A total of 1272 human serum samples collected from volunteers (SARS-CoV-2 infected, non-infected or vaccinated) were evaluated by the two assays. For the RBD-ELISA, plates were coated with RBD, sera were added at 1/5 dilution and bound antibodies were detected with RBD labelled with Horseradish Peroxidase. For the LFA, two parallel strips were used: one for detection of N-specific antibodies (Hoste A. el al, 2020); and another one for detection of S-specific antibodies, using S both as capture and detector reagent. Twenty microliters of blood or ten microliters of serum were applied to each cassette and results were interpreted after ten minutes.A seroneutralization assay was used as reference for the detection of neutralizing antibodies with RBD-ELISA and Reference sera (World Health Organization), for determination of the Limit of detection (LoD). MedCalc® 10 software was used for statistical analysis. Results: The potential diagnostic application with sera from naturally infected and non-infected volunteers showed sensitivity, specificity and agreement (kappa) values of 95.1%, 99.0% and 0.94 respectively for RBD-ELISA; while 97.2%, 99.3% and 0.967 respectively for N-LFA; or 93.2% 98.3 %, 0.923, respectively for S-LFA. Serum samples from vaccinated individuals were analyzed for the specific detection of antibodies to the S protein: for vaccinated but non-infected individuals, sensitivity reached 97.3% after 15 days post-second vaccination dose whereas for previously infected people reached 100% after only 15 days post-first dose. The performance of RBD-ELISA showed good agreement with seroneutralization and excellent agreement with S-LFA (kappa 0.979). Conclusion: The dual N/S LFA represents a valuable tool to detect SARS-CoV-2 infection due to its complementary information on N and S-specific antibody response. Furthermore, the S-LFA and RBD-ELISA are both proven to be able to determine the extent of antibody response after vaccination.