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. 2020 Nov 12;183(4):1058–1069.e19. doi: 10.1016/j.cell.2020.09.049

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© 2020 The Authors

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

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Figure S5 — Binding Epitope Characterization of Selected mAbs, Related to Figures 1, 3, and 4 and Table S3

(A) Competition for RBD binding between top 18 mAbs and ACE2. ELISA-based measurements of human ACE2 binding to SARS-CoV-2 RBD after pre-incubation with the indicated neutralizing mAbs. Values are shown relative to antibody-free condition as mean + SD from three independent measurements.

(B) Competition for RBD binding between combinations of potent neutralizing mAbs is illustrated as a heatmap. Shades of green indicate the degree of competition for RBD binding of detection mAb in presence of 100-fold excess of competing mAb relative to non-competition conditions. Green squares indicate no competition. Values are shown as mean of two independent experiments.

(C) Representative immunofluorescence staining on VeroB4 cells overexpressing spike protein of indicated coronavirus with SARS-CoV-2 mAb CV07-209 at 5 μg/ml. For all other 17 of the selected 18 mAbs (top 18, Table S3), similar results were obtained.

(D) Binding of indicated mAbs to fusion proteins containing the RBD of indicated coronaviruses and the constant region of rabbit IgG revealed by ELISA. For all other top 18 mAbs, similar results were obtained as for CV07-209. Values indicate mean + SD from two wells of one experiment.

(E) Representative HEp-2 cell staining with a commercial anti-nuclear antibody as positive control revealed nuclear binding (top). S1-reactive non-neutralizing mAb CV38-148 exhibited cytoplasmatic binding (middle). Neutralizing mAb CV07-209 showed no binding (bottom). All mAbs selected for detailed characterization (top 18, Table S3) revealed similar results like CV07-209 when used at 50 μg/ml. Representative scale bar: 25 μm.

(F) Structural comparison of CV07-270/RBD and P2B-2F6/RBD complexes. Structure of CV07-270 (pink, left) and structure of P2B-2F6 (PDB 7BWJ) (Ju et al., 2020) (blue, middle) in complex with RBD (white), as well as superimposition of the structures of CV07-270/RBD and P2B-2F6/RBD based on the RBD (right).