Figure 4.

pMos1 and pMos2 promote antigen-specific cross-reactive B cell responses. C57BL/6 mice were inoculated with three doses of DNA plasmid intramuscularly every 2 weeks. Mice were sacrificed 2 weeks after the third immunization. Splenocytes were stained with fluorescence conjugated DENV1 and DENV2 Env proteins plus antibodies specific for different B cell surface markers. (A) Representative flow cytometry plots showing gating strategies to identify antigen-specific B cells that are class-switched, IgM⁺ or IgG⁺. The CD90.2, F4/80, CD11c, CD4, CD8, Ly6G, and NK1.1 antibodies were used for dump gating. The MBC refer to memory B cells, the GC refer to germinal center, and AP refer to activated precursors. Comparison of percentages of DENV1 Env-reactive (B), DENV2 Env-reactive (C), and cross-reactive (D) B cells that are class-switched (top row), IgM⁺ (middle row) and IgG1⁺ (bottom row). (E) Comparison of percentages of the class-switched, IgM⁺ and IgG1⁺ B cells that express Bcl-6 in splenocytes of mice immunized with pDen1, pMos1, pDen2, or pMos2. The data are mean ± SEM (n = 10 per group). The statistical comparison was assessed by Mann-Whitney U test.