Metadata about: High throughput serological tests that can establish the presence and functional activity of anti-SARS-COV2 antibodies are urgently needed. Here we present microsphere-based Flow Cytometry assays that quantify both anti-spike IgGs in plasma, and the ability of plasma to inhibit the binding of spike protein to angiotensin converting enzyme 2 (ACE2). First, we detected anti-trimer IgGs in 22/24 and anti-RBD IgGs in 21/24 COVID+ subjects at a median of 36 (range 14-73) days following documented SARS-CoV-2 RNA (+) secretions. Next, we find that plasma from all 22/24 subjects with anti-trimer IgGs inhibited ACE2-trimer binding to a greater degree than controls, and that the degree of inhibition correlated with anti-trimer IgG levels. Depletion of trimer-reactive Igs from plasma reduced ACE2-trimer inhibitory capacity to a greater degree than depletion of RBD-reactive Igs, suggesting that inhibitory antibodies act by binding both within and outside of the RBD. Amongst the 24 subjects, presence of fever was associated with higher levels of anti-trimer IgG and inhibition of binding to human ACE2. This inhibition assay may be broadly useful to quantify the functional antibody response of recovered COVID19 patients or vaccine recipients in a cell-free assay system.

About: High throughput serological tests that can establish the presence and functional activity of anti-SARS-COV2 antibodies are urgently needed. Here we present microsphere-based Flow Cytometry assays that quantify both anti-spike IgGs in plasma, and the ability of plasma to inhibit the binding of spike protein to angiotensin converting enzyme 2 (ACE2). First, we detected anti-trimer IgGs in 22/24 and anti-RBD IgGs in 21/24 COVID+ subjects at a median of 36 (range 14-73) days following documented SARS-CoV-2 RNA (+) secretions. Next, we find that plasma from all 22/24 subjects with anti-trimer IgGs inhibited ACE2-trimer binding to a greater degree than controls, and that the degree of inhibition correlated with anti-trimer IgG levels. Depletion of trimer-reactive Igs from plasma reduced ACE2-trimer inhibitory capacity to a greater degree than depletion of RBD-reactive Igs, suggesting that inhibitory antibodies act by binding both within and outside of the RBD. Amongst the 24 subjects, presence of fever was associated with higher levels of anti-trimer IgG and inhibition of binding to human ACE2. This inhibition assay may be broadly useful to quantify the functional antibody response of recovered COVID19 patients or vaccine recipients in a cell-free assay system.

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