(A) Schematics of single-cell T cell cloning. Post-vaccine T cells from HD2 and HD3 were stained with 1501-YF45 tetramers; sorted based on T_MN, T_SCM, or T_CM phenotypes; and expanded for 2 to 3 weeks in culture. (B) In vitro–expanded T cell clones were restained with tetramers and cultured with vehicle- (DMSO) or peptide-treated monocyte-derived DCs. Representative plots show tetramer staining and cytokine production by intracellular cytokine staining. (C and D) T cell clones were stimulated with decreasing concentrations of YFV peptides. The response was measured by TNF-α production (C) and quantified by EC₅₀ values after subtracting the background signal from vehicle-treated control (D). (E) Peptide dose response of T cell clones by IFN-γ, IL-2, and IL-2⁺TNF-α⁺IFN-γ⁺ production. (F) Representative histograms show CTV dilution in response to 10 μg/mL of peptide stimulation. (G) Plot summarizes the frequency of the CTV^lo population after a 5-day culture for clones in each phenotypic group. All experiments were repeated at least twice with n = 5 in each group. (C and E) RM 2-way ANOVA was performed and corrected with Tukey’s multiple comparison test. (D and G) Kruskal-Wallis test and Dunn’s multiple comparison test were used. PHA, phytohemagglutinin.