IgG3 and IgM Identified as Key to SARS-CoV-2 Neutralization in Convalescent Plasma Pools

Abstract

Analysis of convalescent plasma derived from individuals has shown that IgG3 has the most important role in binding to SARS-CoV-2 antigens; however, this has not yet been confirmed in large studies, and the link between binding and neutralization has not been confirmed. By analyzing plasma pools consisting of 247-567 individual convalescent donors, we demonstrated the binding of IgG3 and IgM to Spike-1 protein and the receptor-binding domain correlates strongly with viral neutralization in vitro. Furthermore, despite accounting for only approximately 12% of total immunoglobulin mass, collectively IgG3 and IgM account for approximately 80% of the total neutralization. This may have important implications for the development of potent therapies for COVID-19, as it indicates that hyperimmune globulins or convalescent plasma donations with high IgG3 concentrations may be a highly efficacious therapy.

Fig 1. Experimental depletion workflow of convalescent plasma pool.

Δ indicates removal of specific Ig class.

Fig 2. Serological characterization of investigated plasma pools.

(A) Neutralization titer of convalescent (n = 7) and pre-pandemic (n = 3) plasma pools. Data are shown as mean ± SD. (B) EUROIMMUN testing results (S1-IgG ELISA) for convalescent plasma pools (n = 7). (C) Ig classes of convalescent and pre-pandemic plasma pools against S1 ELISA (modified EUROIMMUN ELISA). Data are shown as mean ± SD. Asterisks indicate the level of statistical significance calculated using the Student’s t-test: *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001. (D) Representative plot for AUC calculation in a single pool sample showing transformed optical density values for different dilutions. AUC, area under the curve; Ig, immunoglobulin; LDL, lower detection limit; NT, neutralization titer; SD, standard deviation.

Fig 3. Binding capability of SARS-CoV-2 specific antibody Ig classes utilizing ELISAs to different viral antigens in selected individual convalescent donors and pools to SARS-CoV-2 viral antigens and correlation analysis.

(A–C) Ig classes of seven convalescent and three pre-pandemic plasma pools against: A) NC; B) S1; C) S1-RBD. Data are shown as mean ± SD. Asterisks indicate the level of statistical significance calculated using the Student’s t-test: *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001. AUC, area under the curve. (D-F) Overlay of specific binding capacity of individual donors, plasma pools, and pre-pandemic plasma pools against D) NC, E) S1, and F) S1-RBD respectively. (G) Neutralizing potency (AU/ml) of pre-pandemic pools, convalescent pools, and individual donors. AU, arbitrary unit;

Fig 4. Comparison of neutralization capacity against total Ig mass.

(A) Contribution of Ig classes to the neutralization of SARS-CoV-2 (percentage contribution) based on experimental data depicted in Table 1. (B) The abundance of Ig classes as percentage of total Ig in convalescent plasma pool are based on quantitative nephelometric results.