Figure 8

The number of MAGE-A4-specific Th1 cells increased in vitro culture and the expanded Th1 cells were activated in the presence of the cognate peptide without APC. (A) CD4⁺ T cells from donors 4 and 5 were stimulated with APC pulsed with MAGE-A4-helper epitope peptide weekly. At days 0, 7, 14, 21 and 28, the cultured T cells were collected and the total cell number was counted. The growth rates compared with number at day 0 were displayed. (B) In the absence of APC, MAGE-A4-speficific Th1 cells were cultured with irrelevant (open bar) or cognate (filled bar) peptide (10 μg ml⁻¹) for 24 h. The culture supernatants were collected and assessed cytokine levels in ELISA. (C) In the absence of APC, the Th1 cells were cultured with autologous CD8⁺ T cells in the presence of control (irrelevant) or cognate (M41) peptide (10 μg ml⁻¹). At day 5, IFN-γ-producing CD8⁺ T cells in stimulating with anti-CD3 mAb were assessed by intracellular staining using PE-conjugated anti-CD8 mAb and FITC-conjugated anti- IFN-γ mAb. These experiments were carried out independently at least three times. Data denote mean±s.e.m. and are representative of the experiments.