Figure 4.

More Complex acDC Cytokine Cocktails Induce Ag-Reactive CD8⁺ TC in Similar Numbers but Produce More Cytokines

(A) PBMCs and CBMCs (3 donors each, same donors as in Figure 3; 2 × 10⁶ CBMCs per well in 48-well plates) were cultured using the indicated acDC cytokine cocktails followed by TNF-α, PGE2, IL-1β, and low-dose IL-7 along with MelanA_26–35 peptide after 24 h and by IL-2, IL-15, and IL-7 from day 2 of culture. Absolute numbers of MelanA_26–-35-reactive CD8⁺ T cells per million PBMCs (green) or CBMCs (black) obtained at the end of the 10-day culture are plotted, as determined with MelanA_26–35-loaded HLA-A2 MMrs and CountBright beads. (B) The same cultures were tested at day 13 during a 6-h recall assay in the presence of HLA-A2⁺ LCL cells pulsed with MelanA_26–35 or no peptide. The graph displays absolute numbers of CD8⁺ T cells from PBMCs (green) or CBMCs (black) producing at least one cytokine among IFN-γ, TNF-α, IL-2, and MIP-1β in response to MelanA_26–35-pulsed LCL cells after background subtraction, i.e., the number of cytokine-positive CD8⁺ T cells detected in response to unpulsed LCL cells. Results are expressed as mean ± SD (*p < 0.05).