Abstract
ObjectiveThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic challenges national health systems and the global economy. Monitoring of infection rates and seroprevalence can guide public health measures to combat the pandemic. This depends on reliable tests on active and former infections. Here, we set out to develop and validate a specific and sensitive enzyme linked immunosorbent assay (ELISA) for detection of anti-SARS-CoV-2 antibody levels.MethodsIn our ELISA, we used SARS-CoV-2 receptor-binding domain (RBD) and a stabilized version of the spike (S) ectodomain as antigens. We assessed sera from patients infected with seasonal coronaviruses, SARS-CoV-2 and controls. We determined and monitored IgM-, IgA- and IgG-antibody responses towards these antigens. In addition, for a panel of 22 sera, virus neutralization and ELISA parameters were measured and correlated.ResultsThe RBD-based ELISA detected SARS-CoV-2-directed antibodies, did not cross-react with seasonal coronavirus antibodies and correlated with virus neutralization (R2 = 0.89). Seroconversion started at 5 days after symptom onset and led to robust antibody levels at 10 days after symptom onset. We demonstrate high specificity (99.3%; N = 1000) and sensitivity (92% for IgA, 96% for IgG and 98% for IgM; > 10 days after PCR-proven infection; N = 53) in serum.ConclusionsWith the described RBD-based ELISA protocol, we provide a reliable test for seroepidemiological surveys. Due to high specificity and strong correlation with virus neutralization, the RBD ELISA holds great potential to become a preferred tool to assess thresholds of protective immunity after infection and vaccination.
Highlights
Coronaviruses (CoV) are known to cause respiratory diseases in humans
Virus neutralization assay using SARS-CoV-2 from respiratory specimen is described in the supplementary materials and methods
To promote robust production of the two recombinant proteins, coding sequences were codon-optimized and the autologous S-protein signal peptide was replaced by a minimal version of the tissue plasminogen activator signal peptide developed in our lab
Summary
The alphacoronaviruses HCoV-229E and HCoV-NL63 as well as the betacoronaviruses HCoV-OC43 and HCoV-HKU1 circulate seasonally in humans and cause common colds [1]. The immune response to the seasonal CoVs and SARSCoV and MERS-CoV has been studied intensively [3, 4]. They are able to trigger a humoral immune response that correlates with disease severity [3]. Mild infections resulted in short-lived and very low antibody titers near the detection limit. This loss of humoral immunity has been linked to the occurrence of (re-)infections with seasonal CoVs [3]
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