Fig. 4

Ex vivo peptide stimulation assays of T cells. T cells were pulsed with peptide, expanded and restimulated in short-term in vitro culture. Cells were gated on live-cells, lymphocytic fraction, CD3⁺ cells. a At IM1, IFN-γ secretion of T cells was measured after stimulation with a peptide cocktail containing all four peptides 4. b At IM2, T cells were stimulated with single peptides, including a no-peptide control, respectively. c IFN-γ-secretion and phenotype of a T-cell clone (#33) responsive to peptide 1 obtained from peripheral T cells at IM3. d Percentage of cells from clonal populations of defined specificity responding to their peptide with IFN-γ⁺ production. e mRNA expression of granzymeB, perforin and CRTAM in peptide-specific clones from IM3 measured by RT-PCR. Expression of the target gene was normalized to expression of human GAPDH. *P < 0.05, **P < 0.01