Fig. 7: Validation of R3DC23 binding contacts.
From: Ultrapotent SARS coronavirus-neutralizing single-domain antibodies that clamp the spike at its base
a Binding of R3DC23 to cells expressing the indicated spike mutants, measured by flow cytometry. The left graph shows the ratio of the MFI of R3DC23 binding at the indicated concentrations to transfected (GFP+) cells expressing spikes harboring the indicated point mutation and that of non-transfected (GFP-) cells. The right graph shows the mean EC50 of R3DC23 binding to cells expressing spikes harboring the indicated point mutations (n = 2). b Neutralization of VSV particles pseudotyped with SARS-CoV-2 spikes harboring the indicated point mutations. Mean neutralization IC50 (n = 2) is shown. c Fusogenicity of spike variants with point mutations introduced in the HR2 region, in the presence of palivizumab (negative control) and huR3DC23-Fc. Cells were treated with antibody (1 µg/mL), 2 h after co-transfection of a spike variant and a GFP expression vector, and syncytium formation was monitored with live-cell imaging. Shown in the graph is the mean ± SD (N = 3 samples) slope of the curve derived from the area of green objects at time points 24 h after transfection and 38 h after transfection as a measure for fusogenicity of the HR2 mutant. d Neutralization of VSV particles pseudotyped with BA.2.75 or BA2.75.2 spikes by huR3DC23-Fc, S309 (positive control) and palivizumab (negative control). Mean ± SD (N = 3 replicates of the same dilution series) of normalized GFP MFI is shown. e Binding to surface-expressed Khosta-2 spike by a dilution series of R3DC23 (lef graph) or huR3DC23-Fc, palivizumab or S309 at 10 µg/mL. The ratio of MFI of bound antibody to GFP+ cells expressing spikes or no spikes over that of non-transfected GFP- cells is shown. Source data are provided as a source data file.
