Phenotypic and functional analysis of T cell responses generated by p55^gag and p24CE DNA vaccination.

<p><b>(A)</b> Mice (N = 5/group) were vaccinated 3 times (week 0, 3 and 6) with 20 µg of a plasmid expressing HXB2 p55^gag (clade B) or 20 µg of a mixture of plasmids expressing SP-p24CE1 and SP-p24CE2. The mice were sacrificed 2 weeks after the last immunization. Three independent experiments were performed and a representative experiment is shown. (<b>B</b>) Pooled splenocytes were stimulated with Clade A, B or C peptide pools (15-mers) spanning the p24^gag region (left panel) and the Group M consensus peptide pool (right panel). The frequency of the CD4⁺ (open bars) and CD8⁺ (filled bars) p24^gag-specific IFN-γ producing T cells was determined. (<b>C</b>) The splenocytes from the SP-p24CE (left panel) and p55^gag (right panel) DNA vaccinated mice were stimulated with peptide pools specific for the individual CEs. The frequency of the CD4⁺ (open bars) and CD8⁺ (filled bars) CE-specific IFN-γ producing T cells was determined. (<b>D</b>) Plot overlays show the phenotypic and functional characterization of the antigen-specific T cells induced by SP-p24CE (left panels) and p55^gag (right panels) DNA vaccines upon stimulation with p24^gag–specific peptide pool. Total T cells recovered from the spleen are shown as grey contours, and the antigen-specific IFN-γ⁺ T cells are overlaid as red (CD4⁺ T cells) or black (CD8⁺ T cells) dots. The plots show the CD4/CD8 distribution (top panel), memory phenotype as determined by CD44/CD62L staining (middle panel) and TNF-α/CD107a expression (bottom panel) among the T cells from vaccinated mice. The frequency of CD4 (red) and CD8 (black) IFN-γ T lymphocytes is shown.