Figure 4. Detection of OCT4-reactive CD8⁺ T cells. (A–C) MACS-sorted CD4⁺ cells were stimulated with autologous dendritic cells (DCs) loaded with an immunoreactive OCT4-derived peptide. CD4⁺ cells stimulated with anti-CD3/anti-CD28 beads and CD4⁺ cells exposed to unloaded DCs served as a positive and negative control condition, respectively. (A) Results from one representative donor showing no significant interferon γ (IFNγ) production (on day 3) and no antigen-dependent CD8⁺ T-cell proliferation (on day 6) (p > 0.05) are reported. (B) After 12 d of co-culture, a second stimulation was performed with -derived peptide-loaded DCs. Thereafter, peptide-specific CD8+ T cells were analyzed for their cytotoxic functions by intracellular cytokine staining and flow cytometry. The expression of activation markers by cells from one representative healthy donor is shown, together with the gating strategy. (C) Results from one representative donor showing the mobilization of CD107a and the expression of IFNγ and tumor necrosis factor α (TNFα) by the same cells [gating strategy as in (B)] are shown.