Figure 4.

Potent immunogenicity of the parental RBD-I53-50 nanoparticle immunogen is maintained with addition of Rpk mutations. (A) Female BALB/c mice (six per group) were immunized at weeks 0 and 3. Each group received equimolar amounts of RBD antigen adjuvanted with AddaVax, which in total antigen equates to 5 μg per dose for HexaPro-foldon and 0.9 μg per dose for all other immunogens. Serum collection was performed at weeks 2 and 5. The RBD-I53-50 immunogen was prepared in two different buffer conditions, with one group including CHAPS as an excipient to bridge to previous studies. (B) Binding titers against HexaPro-foldon at weeks 2 and 5, as assessed by AUC from ELISA measurements of serial dilutions of serum. Each circle represents the AUC measurement from an individual mouse and horizontal lines show the geometric mean of each group. One mouse with a near-zero AUC at week 2 for group four was not plotted but still included in the geometric mean calculation. Midpoint titers are shown in Supplementary Figure 5A . (C) Autologous (D614G) pseudovirus neutralization using a lentivirus backbone. Each circle represents the neutralizing antibody titer at 50% inhibition (IC₅₀) for an individual mouse and horizontal lines show the geometric mean of each group. Pseudovirus neutralization titers using an MLV backbone are shown in Supplementary Figure 5B . Statistical analysis was performed using one-sided nonparametric Kruskal–Wallis test with Dunn’s multiple comparisons. *p < 0.05; **p < 0.01; ***p < 0.001.