Serological Test to Determine Exposure to SARS-CoV-2: ELISA Based on the Receptor-Binding Domain of the Spike Protein (S-RBD_N318-V510) Expressed in <i>Escherichia coli</i>

Abstract

Massive worldwide serological testing for SARS-CoV-2 is needed to determine the extent of virus exposure in a particular region, the ratio of symptomatic to asymptomatic infected persons, and the duration and extent of immunity after infection. To achieve this, the development and production of reliable and cost-effective SARS-CoV-2 antigens is critical. We report the bacterial production of the peptide S-RBDN318-V510, which contains the receptor-binding domain of the SARS-CoV-2 spike protein (region of 193 amino acid residues from asparagine-318 to valine-510) of the SARS-CoV-2 spike protein. We purified this peptide using a straightforward approach involving bacterial lysis, his-tag-mediated affinity chromatography, and imidazole-assisted refolding. The antigen performances of S-RBDN318-V510 and a commercial full-length spike protein were compared in ELISAs. In direct ELISAs, where the antigen was directly bound to the ELISA surface, both antigens discriminated sera from non-exposed and exposed individuals. However, the discriminating resolution was better in ELISAs that used the full-spike antigen than the S-RBDN318-V510. Attachment of the antigens to the ELISA surface using a layer of anti-histidine antibodies gave equivalent resolution for both S-RBDN318-V510 and the full-length spike protein. Results demonstrate that ELISA-functional SARS-CoV-2 antigens can be produced in bacterial cultures, and that S-RBDN318-V510 may represent a cost-effective alternative to the use of structurally more complex antigens in serological COVID-19 testing.

Keywords

• SARS-CoV-2
• COVID-19
• ELISA
• serological testing
• spike
• receptor binding domain