Monoclonal antibodies cloned from donors CoV01 and CoV02.

(A) A representative flow cytometry gating strategy for staining SARS-CoV-2 RBD-specific memory B cells. (B) The frequencies of anti-SARS-CoV-2 RBD-specific memory B cells in COVID-19 donors CoV01-CoV17. No PBMCs were obtained for donor CoV18, therefore this donor was not included. Symbol code is given on the right. (C) Pie charts representing the total number of RBD-specific memory B cell sequences (heavy chains) obtained from donors CoV01 and CoV02. The numbers in the middle of the pies represent the total number of sequences, and the pink shaded slices represent B cell clonal families. The white slices represent single sequences that were not clonal. (D) Number of nucleotide substitutions in VH of the B cell sequences of CoV01 (red) and CoV02 (magenta). Statistical analysis was calculated using one-way ANOVA. (E) Amino acid length of CDRH3 from B cell sequences isolated from CoV01 (red) and CoV02 (magenta). (F) AUC in ELISA for each one of the 22 cloned mAbs against SARS-CoV-2 RBD (left) and Spike (right) proteins. Color code is given on the right of (H). (G) and (H) Degree of mAbs competition with mAb CR3022, and ACE2, as assessed by ELISA. mGO53 was used as isotype control. (I) Frequency of RBD-PE stained hACE2-expressing cells identified by flow cytometry in the presence of the 22 mAbs isolated from CoV01 and CoV02. mAbs that reduced RBD binding to hACE2 are marked with black arrows. Unlabeled RBD was used as a positive control.