Fig. 2: Isolation and characterization of anti-CD4 Nbs.
a Summary of bacterial supernatant binding with CD4 protein tested by phage ELISA. The y-axis represents the ratio of the CD4 binding readout to the blank binding readout. Each dot corresponds to the bacterial supernatant from a single clone among the 1920 clones tested. b Positive binding VHHs inhibiting HIV-1 infection. Each dot represents one VHH clone supernatant. The dashed line indicates 50% relative inhibition. c Phylogenetic analysis of the 47 VHHs using the Neighbor-Joining method. The tree is drawn to scale, and branch lengths correspond to the evolutionary distances measured in amino acid substitutions per site. Evolutionary analyses were conducted using the Poisson correction method, with gaps and missing data excluded. MEGA6 software was utilized for the analyses. d Schematic diagram illustrating the structures of Nb-Fc (VHH fused with the Fc region), where the VHH is linked to the human Fc4 region (CH2–3). e Graph depicting the relative inhibition of cell supernatant from 47 various Nb-Fcs tested for neutralization against pseudovirus HIV-1JR-FL infection. The dashed line represents 50% inhibition. f Binding curves of 28 Nbs exhibiting interactions with the CD4 protein, as identified by BLI (Biolayer Interferometry), among the 47 various Nbs. g Neutralizing curve of the five selected Nbs against reference pseudoviruses of HIV-1JR-FL. Data for (g) are depicted as mean ± SEM. Source data are provided as a Source Data file.
