Figure 3. Inferred naïve mAbs from six lineages vary in antigen binding capability and ADCC function.

(A–B) Binding kinetics of the inferred naive antibody (ligand) from each lineage to indicated concentrations of monomeric C.ZA.1197MB gp41 ectodomain (A) or BL035.W6M.C1 gp120 (B) (analyte). Best fitting lines (red) to a 1:1 binding model of ligand:analyte binding are shown. Data are representative of two independent experiments. (C) Positive control-normalized RFADCC activity of inferred naive antibodies compared to their respective mature antibodies. Normalization is described in Methods. Asterisks indicate indeterminate activity, as defined in Methods. Data are represented as mean ± SEM and reflect at least five independent experiments; source data are available in Figure 3—source data 1. 0: naive; m: mature. See Figure 3—figure supplement 1 for functional assessment of alternative naïve mAbs.

Figure 3—figure supplement 1. Alternative naive mAb functionality.

(A, C) Functional RFADCC comparisons of each lineage’s ‘original’ (0_VH and 0_VL) and ‘alternative’ (A_VH and A_VL) inferred naive antibodies. In cases where the alternative approach did not infer a naive sequence that differed from the original naive sequence identified by partis, alternative chains were paired with original chains, as indicated. For the 157 lineage (C), original and alternative heavy and light chains were also cross-paired. Data are representative of at least two independent experiments. (B) Binding of indicated 157-lineage naive antibodies (ligand) to monomeric BL035 gp120 (analyte, 250 nM), measured by BLI without the ‘double reference’ approach described in Materials and methods. (D) Graphic summary of paired VH and VL antibody sequences relative to inferred naïve 157-0_VH0_VK. Shaded regions demarcate CDRs (gray). Vertical black lines indicate variable region amino acid substitutions. 0: original naive; A: alternative naive; m: mature.