Figure 7.

15Rα/15‐stimulated antigen‐specific T cells efficiently lyse targets at lower effector : target (E : T) ratios. T cell cytotoxic capacity was measured in a standard ⁵¹Cr release assay, performed at 21–28 days after culture initiation using peptide‐loaded autologous BLCL as targets. BLCL not loaded with peptide were usedas control. (a) A fixed E : T ratio of 10 T cells to one target cell was used and the cytotoxic activity of T cells sensitized in all culture conditions was tested against targets loaded with serial dilutions of the NLVPMVATV (NLV) peptide (10 nM, 1 nM, 0·1nM, 10 pM and 0·1pM at 37ºC × 3 h in serum free medium). (b) The cytotoxic activity of T cells was evaluated at decreasing E : T ratios against targets loaded with a fixed concentration (10 nM) of peptide. (c) T cells in all culture conditions were evaluated for expression of intracellular granzyme B upon secondary restimulation with NLV peptide‐loaded autologous peripheral blood mononuclear cells (PBMC) 21–28 days after culture initiation. T cells co‐incubated with peptide‐loaded autologous PBMC were labelled with fluorescently labelled anti‐CD3, anti‐CD8 and anti‐CD4, followed by incubation with anti‐human granzyme B after cell permeabilization and analysed by fluorescence activated cell sorter (FACS). The proportion of granzyme B‐positive T cells CD8⁺ T cells was evaluated. T cells sensitized in the presence of 15Rα/15 complexes generated significantly higher proportions of granzyme B⁺ T cells compared to sensitization in the presence of soluble IL‐2 (P = 0·05).