Figure 3. Cooperation between HEL_105–120 and OVA_323–339-specific CD4 T cells requires simultaneous presentation of both peptides by the same APC.

BALB/c mice were seeded with 10⁵ MACS purified CD4+ DO11.10 T cells one day prior to being injected with 10⁷ splenocytes pulsed with either HEL_105–120 alone (H), OVA_323–339 alone (O), both HEL_105–120 and OVA_323–339 (H+O), or with 10⁷ H and 10⁷ O splenocytes (H, O). The peptide-specific cytokine producing cells in the spleen and draining lymph nodes of these mice were enumerated by ELISpot on day nine after the first injection. Each data point represents the number of specific ELISpot-forming cells in the lymphoid organ of a single mouse. The probability that the mean number of ELISpot-forming cells between the H, H+O and H, O groups is the same was assessed by one-way ANOVA (P-values; top left corner of all plots). Post-hoc comparisons revealed significant differences between the means of the groups indicated. These are pooled results from two to three independent experiments.