An Original Multiplex Method to Assess Five Different SARS-CoV-2 Antibodies

Objectives: Accurate SARS-CoV-2 serological assays are urgently needed to help diagnose infection, determine past exposure of populations and assess the response to future vaccines. The study aims at assessing the performance of the multiplex D-tek COVIDOT 5 IgG assay for the detection of SARS-CoV-2 IgG antibodies (N, S1+S2, S1, S2 and RBD).

Methods: Sensitivity and dynamic trend to seropositivity were evaluated in 218 samples obtained from 46 rRT-PCR confirmed COVID-19 patients. Non-SARS-CoV-2 sera (n=118) collected before the COVID-19 pandemic with a potential cross-reaction to the SARS-CoV-2 immunoassay were included in the specificity analysis.

Results: A gradual dynamic trend since symptom onset was observed for all IgG antibodies. Sensitivities before day 14 were suboptimal. At ≥21 days, sensitivities reached 100% (93.4-100%) for N, S1+S2, S2 and RBD-directed IgG and 96.3% (87.3-99.6%) for S1-directed IgG. In 42 out of 46 patients (91.3%), all five antibodies were detected at ≥14 days. The four remaining patients had between 2 and 4 positive antibodies at their respective maximal follow-up period. The specificity was 100 % for S1+S2, S2 and RBD, 98.3% for N and 92.4% (86.0-96.5%) for S1-directed IgG. The combined use of antigens increases the early sensitivity whilst enforcing high specificity.

Conclusions: Sensitivities at ≥21 days and specificities were excellent, especially for N, S1+S2, S2 and RBD-directed IgG. Caution is however required when interpreting single S1-directed reactivities. Using a multiplex assay complies with the orthogonal testing algorithm of the CDC and allows a better and critical interpretation of the serological status of a patient.