Figure 4. Immunogenicity analysis of mRNA vaccines in K18-hACE2 transgenic mice.

Seven-week-old female K18-hACE2 mice were immunized and boosted with 5 or 0.25 μg of mRNA vaccines (control (black symbols), mRNA-1273 (red symbols), or mRNA-1273.351 (blue symbols). A. Scheme of immunizations, spleen harvest, blood draw, and virus challenge. B. Serum anti-spike IgG responses at three weeks after booster immunization with mRNA vaccines (control (black symbols), mRNA-1273 (red symbols), mRNA-1273.351 (blue symbols) against indicated spike proteins (Wuhan-1, B.1.1.7, B.1.351, or B.1.617.2). (n = 3 (control vaccine) or 8 (spike vaccines), two independent experiments, boxes illustrate mean value, dotted line shows the LOD; two-way ANOVA with Tukey’s post-test: *, P < 0.05; **, P < 0.01). C-L. Serum neutralizing antibody responses three weeks after boosting as judged by FRNT (half-maximal reduction, FRNT₅₀ values) with WA1/2020 D614G (C, H), WA1/2020 D614G/N501Y (D, I), B.1.1.7/E484K (E, J), B.1.351 (F, K), or B.1.617.2 (G, L) in mice immunized with 5 (C-G) or 0.25 (H-L) μg of control (n = 5–10), mRNA-1273 (n = 20–24), and mRNA-1273.351 (n = 21–24) vaccines (two independent experiments, boxes illustrate geometric mean values, dotted line shows LOD; one-way Kruskal-Wallis ANOVA with Dunn’s post-test: *, P < 0.05; **, P < 0.01; ***, P < 0.001; **** P < 0.0001). M-N. Antigenic map of sera from K18-hACE2 mice titrated against WA1/2020 D614G, WA1/2020 N501Y/D614G, B.1.1.7/E484K, B.1.351, and B.1.617.2. The maps show sera from immunized mice that received 5 μg (M) or 0.25 μg (N) doses, respectively with symbol details described in Figure 1. O-R. CD8⁺ (O, P) and CD4⁺ (Q, R) T cell responses in K18-hACE2 mice at day 35, two weeks after booster immunization with mRNA vaccines (n= 10 for each group, two independent experiments, boxes illustrate median values, one-way ANOVA with Tukey’s post-test: ns, not significant; *, P < 0.05; **, P > 0.01; ***, P < 0.001; **** P < 0.0001).