Fig. 1

Generation of Ad2-E, Ad2-prME, and Ad2-prME-NS1 and evaluation of their immunogenicity in mice. a Schematic diagram of the Ad2-ZIKV vaccines in this study. SP signal peptide; 2A the coding sequence for self-cleaving 2A peptide. b, c Expression of ZIKV antigens by Vero cells infected with Ad2-E, Ad2-prME, and Ad2-prME-NS1, as well as an empty Ad2 vector (Ad2-empty) at 100 viral particles (vp) per cell. Forty-eight hours after infection, the expression of E (b) and NS1 (c) in the culture supernatants (upper panel) and cell lysates (bottom panel) were assessed by Western-blot analysis. The full, uncropped graphs can be referred to Supplementary Figs. S8–S13. d Timeline of immunization and immune analysis. Three or 12 weeks after the second immunization, mice were sacrificed and the serum samples were collected. e, f The binding antibodies to E (e) and NS1 (f) at 3 weeks post immunization were assessed by ELISA. The titers were calculated as the reciprocal of the sera dilution at which the optical density value at 450 nm (O.D. 450) was higher than the cut-off. g, h The neutralizing antibodies (g) and the inhibitory antibodies (h) at 3 weeks post immunization were assessed by the Fluorescence-based neutralization assay and inhibition assay, respectively. The titers were calculated as the reciprocal of the sera dilution at which the number of infected cells was reduced by 50%. i, j The binding antibodies to E (i) and NS1 (j) at 12 weeks post immunization. k, l The neutralizing antibodies (k) and the inhibitory antibodies (l) at 12 weeks post immunization. The data were representative of two independent experiments and presented as mean ± standard error (SEM). Comparison between different groups were performed by one-way analysis of variance (ANOVA, n = 5/group). *p < 0.05; **p < 0.01; ***p < 0.001