Affinity of Anti-Spike Antibodies in SARS-CoV-2 Patient Plasma and Its Effect on COVID-19 Antibody Assays

Abstract

Background: Measuring anti-spike protein antibodies in human plasma or serum is commonly used to determine prior exposure to SARS-CoV-2 infection and to assess the anti-viral protection capacity. According to the mass-action law, a lesser concentration of tightly binding antibody can produce the same quantity of antibody-antigen complexes as higher concentrations of lower affinity antibody. Thus, measurements of antibody levels reflect both affinity and concentration. These two fundamental parameters cannot be disentangled in clinical immunoassays, and so produce a bias which depends on the assay format. Methods: To determine the apparent affinity of anti-spike protein antibodies, a small number of antigen-coated magnetic microparticles were imaged by fluorescence microscopy after probing antigen-antibody equilibria directly in patient plasma. Direct and indirect anti-SARS-CoV-2 immunoassays were used to measure antibody levels in the blood of infected and immunized individuals. Findings: We observed affinity maturation of antibodies in convalescent and vaccinated individuals, showing that higher affinities are achieved much faster by vaccination. We demonstrate that direct and indirect immunoassays for measuring anti-spike protein antibodies depend differently on antibody affinity which, in turn, affects accurate interpretation of the results. Interpretation: Direct immunoassays show substantial antibody affinity dependence. This makes them useful for identifying past SARS-CoV-2 exposure. Indirect immunoassays provide more accurate quantifications of anti-viral antibody levels. Funding: No external funding sources were used in this study.Declaration of Interest: All authors are employed by Abbott LabsEthical Approval: Patient plasma/serum samples used for this study were purchased from New York Biologics (Southampton, NY), Access Biologicals, LLC (Vista, CA) and New York Blood Center (New York, NY). The samples purchased from Access Biologicals LLC were collected under an IRB protocol approved by Ballad Health System IRB. The samples purchased from New York Biologics were collected under an IRB approved by Ethical and Independent Review Services (E&I). The patient plasma samples purchased from New York Blood Center (New York, NY), were collected from volunteer blood donors who consented to the use of their samples for research purposes at the time of collection.