FIGURE 3.

Emergence of a functional hierarchy of neu-specific nondominant CD8⁺ T cell responses isolated from vaccinated neu-N and FVB/N mice. One week following vaccination, splenocytes and vaccine draining lymph nodes were collected from five neu-N (A) or FVB/N (B) mice and incubated with 2.5 μg/ml peptide for 7 d. Stimulated lymphocytes were collected and incubated for T2-D^q cells pulsed with the corresponding peptide or NP_118–126 at varying concentrations. Cells were stained for CD8 and IFN-γ and analyzed by intracellular cytokine staining. A, CD8⁺ T cell responses directed against 24F1, 131E and 134C are the most prevalent in vaccinated neu-N mice as determined by peptide titration studies. B, Vaccinated FVB/N mice mount CD8⁺ T cell responses specific for nondominant epitopes that are significantly weaker than responses to RNEU_420–429. Shown for both are the mean percentage of CD8⁺ T cells that are IFN-γ ⁺ after subtraction of background NP_118–126 responses. These experiments were repeated at least twice with similar results.