Fig 4.

Stronger secondary GC responses resulted in higher antibody titers. (A) Scheme of experiments. Two sets of mouse experiments were performed simultaneously. Mice (n = 6) were immunized in prime-only regimens (i.m., i.n., respectively, at 4 weeks) or prime-boost regimens (i.m. + i.n., i.n. + i.m., respectively, with an interval of 4 weeks) with 2 × 10⁷ IFU of AdC68-vST-vtRBM. About 100 µL PBS intramuscularly vaccinated mice served as control. The i.m. dLNs (pooled popliteal, inguinal, and iliac LNs, the dLNs for i.m. vaccination and also served as the ndLNs for i.n. vaccination) and i.n. dLNs (pooled cervical and mediastinal LNs, the dLNs for i.n. vaccination and also served as the ndLNs for i.m. vaccination) were harvested at 7 and 14 days post-last vaccination. (B and C) Absolute number of total or RBD⁺ GC B cells. (D) Spearman correlations of S-binding antibody titers and primary RBD⁺ GC B cell counts. The primary RBD⁺ GC B counts refer to RBD⁺ GC B cells induced by i.m. or i.n. priming in dLNs. BAbs here refer to endpoint titers of i.m. + i.m., i.n. + i.n., i.m. + i.n., i.n. + i.m. regimens after boosting. Data of control groups were also contained in all Spearman correlation analysis. (E) Spearman correlations of pVNT₅₀ titers and primary RBD⁺ GC B cell counts. pVNT₅₀ titers here refer to those of i.m. + i.m., i.n. + i.n., i.m. + i.n., and i.n. + i.m. regimens after boosting. (F) Spearman correlations of S-binding antibody titers and secondary RBD⁺ GC B cell counts. BAbs here refer to S-binding antibody titers of i.m. + i.n. and i.n. + i.m. regimens after boosting. Secondary RBD⁺ GC B counts refer to RBD⁺ GC B cells induced by i.m. + i.n. and i.n. + i.m. regimens in dLNs. (G) Spearman correlations of pVNT₅₀ titers and secondary RBD⁺ GC B cell counts. pVNT₅₀ titers here refer to those of i.m. + i.n. and i.n. + i.m. regimens after boosting. All data are represented as mean ± SEM and analyzed by one-way ANOVA with Tukey correction.*P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, and ****P ≤ 0.0001.