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 Escherichia coli

Márquez-Ipiña, Alan Roberto; González-González, Everardo; Rodríguez-Sánchez, Iram Pablo; Lara-Mayorga, Itzel Montserrat; Mejía-Manzano, Luis Alberto; Sánchez-Salazar, Mónica Gabriela; González-Valdez, José Guillermo; Ortiz-López, Rocio; Rojas-Martínez, Augusto; Trujillo-de Santiago, Grissel; Alvarez, Mario Moisés

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-RBD(N318-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-RBD(N318-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-RBD(N318-V510). Attachment of the antigens to the ELISA surface using a layer of anti-histidine antibodies gave equivalent resolution for both S-RBD(N318-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-RBD(N318-V510) may represent a cost-effective alternative to the use of structurally more complex antigens in serological COVID-19 testing.