Fig 3. Characterization of Anti-MAYV Antibody Responses.
(A) Isotype specific ELISAs were performed to characterize and measure the MAYVBeAr binding antibodies in sera from naïve, AdV-MAYV prime vaccinated, AdV-MAYV prime + boost vaccinated, and MAYV infected mice. Preparations of heat inactivated whole MAYV stocks were bound to high affinity 96-well plates. Serial dilutions of mouse sera were plated in order to calculate binding dilution titer. Binding antibodies were detected by secondary antibodies specific for mouse IgG1, IgG2a, and IgG3 as well as a pan IgG/IgM. Error bars represent SD representative of quadruplicate biological replicates. Statistical analysis was performed on log transformed data by a one-way ANOVA (*P < 0.05, **P < 0.005, ***P = 0.0001, P < 0.0001). (B) Western blot analysis was used to determine antigen specificity of antibodies in serum from vaccinated mice. Protein lysates containing purified MAYVTrVl were separated by SDS-PAGE and transferred to immunoassay membranes for western blotting. Cross-reactive anti-CHIKV E1 (panel 1) and E2 (panel 2) monoclonal antibodies were used to identify MAYV envelope proteins. Serum derived from naïve mice (panel 3) and AdV-MAYV prime + boost vaccinated mice (panel 4) demonstrate the presence of envelope and capsid specific antibodies to MAYVTrVl following vaccination. The blots shown are representative images of 3 independent experiments.