FIG. 6.

Adjuvant effect of recombinant MVA expressing murine RANKL. (A) BHK-21 cells were infected with either MVA-RANKL or MVA-p3 and then were stained with biotinylated anti-RANKL monoclonal antibody followed by PE-labeled streptavidin. The bold histograms indicate the staining with anti-RANKL monoclonal antibody and the thin histograms indicate the staining with isotype-matched control rat immunoglobulin G. (B) C57BL/6 mice (n = 3) were first primed with 5 × 10⁸ PFU of Ad-MANY and then boosted 11 days later with 5 × 10⁸ PFU of MVA-MANY or MVA-p3 and 5 × 10⁷ PFU of MVA-RANKL or MVA-p3. The mice were sacrificed 10 days after the boost immunization, and their spleens were removed. The freshly isolated splenocytes were subjected to the ELISPOT assay for IFN-γ-producing cells in response to ANYNFTLV peptide-pulsed EL-4 cells. The number of IFN-γ-secreting cells/10⁶ cells was counted 24 h later. The number of IFN-γ-secreting cells that appeared against peptide-unpulsed EL-4 was subtracted from the number of IFN-γ-secreting cells that appeared against peptide-pulsed EL-4. Data represent the mean ± standard deviation of three mice in each group. *, P < 0.05 by the Dunnett's two-tailed t test. NS, not significantly different. The data are representative one of two independent experiments.