Schematic for macaque trials. DNA/HIV and MVA/HIV are recombinant DNA and MVA vectors expressing HIV-1 vaccine inserts, respectively. i.v, intravenous; i.m, intramuscular; pfu, plaque forming units.

VV-specific neutralizing antibody elicited by HIV DMM vaccine. (A) Neutralizing antibody response against intracellular mature virion form of vaccinia virus measured using a GFP/flow based assay. Each symbol represents an individual macaque. The shaded area on the graph represents the background level of neutralizing activity observed in the sera of unvaccinated macaques. (B) Neutralizing antibody response against the extracellular form of vaccinia virus using a comet assay. VIG, vaccinia immunoglobulin.

Cellular and humoral immunity, and correlates for protection following MPXV challenge. (A) MPXV-specific neutralizing antibody. Neutralization of monkeypox Zaire strain was performed using a plaque reduction assay. (B) VV-specific CD8 T cell response. VV-specific IFN-γ producing CD8 T cells were measured using an ICS assay. (C) Correlation between the peak viremia and immune responses on the day of challenge. (D) Correlation between the peak viremia and immune responses at day 8, post challenge.

Temporal VV-specific cellular immunity elicited by the DMM and Dryvax vaccines. VV-specific IFN-γ producing CD4 and CD8 T cells were measured using an ICS assay. (A) A representative ICS assay. PBMC were stimulated with vaccinia as described in materials and methods and stained for CD3, CD8 and IFN-γ. Cells were gated on lymphocytes based on the scatter pattern, followed by CD3 expression and analyzed for the expression of CD8 and IFN-γ. Cells in the right quadrants represent CD8 cells and left quadrants represent CD4 cells (CD3 positive, CD8 negative). The frequencies in the upper quadrants are IFN-γ producing cells as a % of total CD4 cells (left quadrants) or total CD8 cells (right quadrants). (B) VV-specific CD4 response in DMM vaccinated macaques. (C) VV-specific CD4 response in Dryvax vaccinated macaques. (D) VV-specific CD8 response in DMM vaccinated macaques. (E) VV-specific CD8 response in Dryvax vaccinated macaques. Each symbol represents an individual macaque. There are 15 animals in the DMM group. For week 9, data are for 5 macaques and for week 174, data are for 10 macaques. The numbers on the graphs represent the geometric means for the groups at different time points. The shaded area on the graph represents the level of detection.

Comparison of VV-specific cellular and humoral responses elicited by Dryvax, and DMM vaccines in macaques. (A) VV-specific CD4 T cell response. (B) VV-specific CD8 T cell response. VV-specific IFN-γ producing CD4 and CD8 T cells were measured using an ICS assay. (C) Vaccinia virus MV-specific neutralizing antibody response. MV-specific neutralizing antibody response was measured using GFP based assay. Each symbol represents an individual macaque. The numbers on the graph represent the geometric means for the group at different time points. The shaded area on the graph represents the level of detection. ‘n’ represents the number of animals in the respective group. Peak CD4 and CD8 responses for Dryvax and DMM represent weeks 2 and 33 of the respective trials. Memory CD4 and CD8 responses for Dryvax and DMM represent weeks 12 and 40 of the respective trials. Peak neutralizing antibody response for Dryvax and DMM represent weeks 2 and 35 of the respective trials. Memory neutralizing antibody response for Dryvax and DMM represent weeks 84 and 167 of the respective trials. DMM, DNA/MVA/MVA.