Antibody Mediated Immunity to SARS-CoV-2 and Human Coronaviruses: Multiplex Beads Assay and Volumetric Absorptive Microsampling to Generate Immune Repertoire Cartography

Abstract

The COVID-19 pandemic is caused by SARS-CoV-2, a novel zoonotic coronavirus. Emerging evidence indicates that preexisting humoral immunity against other seasonal human coronaviruses (HCoVs) plays a critical role in the specific antibody response to SARS-CoV-2. However, current work to assess the effects of preexisting and cross-reactive anti-HCoVs antibodies has been limited. To address this issue, we have adapted our previously reported multiplex assay to simultaneously and quantitatively measure anti-HCoV antibodies. The full mPlex-CoV panel covers the spike (S) and nucleocapsid (N) proteins of three highly pathogenic HCoVs (SARS-CoV-1, SARS-CoV-2, MERS) and four human seasonal strains (OC43, HKU1, NL63, 229E). Combining this assay with volumetric absorptive microsampling (VAMS), we measured the anti-HCoV IgG, IgA, and IgM antibodies in fingerstick blood samples. The results demonstrate that the mPlex-CoV assay has high specificity and sensitivity. It can detect strain-specific anti-HCoV antibodies down to 0.1 ng/ml with 4 log assay range and with low intra- and inter-assay coefficients of variation (%CV). We also estimate multiple strain HCoVs IgG, IgA and IgM concentration in VAMS samples in three categories of subjects: pre-COVID-19 (n=21), post-COVID-19 convalescents (n=19), and COVID-19 vaccine recipients (n=14). Using metric multidimensional scaling (MDS) analysis, HCoVs IgG concentrations in fingerstick blood samples were well separated between the pre-COVID-19, post-COVID-19 convalescents, and COVID-19 vaccine recipients. In addition, we demonstrate how multi-dimensional scaling analysis can be used to visualize IgG mediated antibody immunity against multiple human coronaviruses. We conclude that the combination of VAMS and the mPlex-Cov assay is well suited to performing remote study sample collection under pandemic conditions to monitor HCoVs antibody responses in population studies.

Boosting of Cross-Reactive Antibodies to Endemic Coronaviruses by SARS-CoV-2 Infection but not Vaccination with Stabilized Spike

“…To date, antibody cross-reactivity has been inferred from indirect evidence in the form of boosted responses to endemic CoV [23][24][25][26][27] , and more conclusively observed for select monoclonal antibodies that have been cloned and cross tested 33,34 . To better generalize the more definitive monoclonal studies, we sought to directly define the cross-reactivity of polyclonal antibodies raised following SARS-CoV-2 infection.…”

“…Numerous studies have observed elevated responses to endemic CoV following SARS-CoV-2 infection [23][24][25][26][27] , and more recent work has shown that the magnitude of this "back-boosting" effect is inversely correlated with the induction of IgG and IgM against to SARS-CoV-2 spike (S) protein 28 . Given differential degrees of homology between the receptor binding domain (RBD) in S1 that is the target of the majority of neutralizing antibodies, and the better conserved S2 domain that may be the target of antibodies with diverse effector functions but which are rarely neutralizing, the original antigenic sin hypothesis suggests that lower titers of neutralizing antibodies against SARS-CoV-2 may result from boosting of pre-existing cross-reactive lineages.…”

Pre-Existing Cross-Reactive Antibody Responses Do Not Significantly Impact Inactivated COVID-19 Vaccine-Induced Neutralization

“…Additionally, T cell-mediated anti-SARS-CoV-2 responses were found in unexposed human, indicating a cross-reaction between human CoVs and SARS-CoV-2 (10)(11)(12)(13)(14). Accumulating evidence suggests that pre-existing humoral immunity to sCoVs may play a role in the specific anti-SARS-CoV-2 antibody responses (15), but this possibility is controversial (16,17). Some studies showed that elevated levels of pre-existing antibodies against sCoVs, specifically OC43 and HKU1, were associated with a less severe course of COVID-19, suggesting a protective effect of prior exposure to sCoVs (18)(19)(20).…”

Formation and Expansion of Memory B Cells against Coronavirus in Acutely Infected COVID-19 Individuals

“…Levels of binding IgG Abs from serum, MPAbs, and PPAbs were measured against a broad range of coronavirus proteins (Table S2) by multiplex assay, as recently described [54]. Sera was diluted to 1:1000 and Abs secreted by PBs and post-stimulated MBCs were diluted to 1:2, all with PBS.…”