Fig. 2. APNs target Ag-specific T cells and induce cell uptake in vitro.

(A) Illustration of the interaction of P14 CD8⁺ T cells with its cognate APNs (GP33/D^b), in contrast to the lack of binding to the noncognate control (GP100/D^b APN). (B) Representative flow plots of noncognate APNs and cognate APNs binding to CD8⁺ T cells in splenocytes from P14 TCR transgenic mice. Frequencies depicted are based on gating on CD8⁺ cells. (C) Ag-specific binding of cognate and noncognate APNs to CD8⁺ T cells isolated from TCR transgenic P14 or OT-1 mice. ****P < 0.0001, one-way analysis of variance (ANOVA) and Tukey post-test and correction. All data are means ± SD; n = 3 biologically independent wells. (D and E) OT-1 CD8⁺ splenocytes stained with OVA/K^b APNs at 4° or 37°C and analyzed by flow cytometry before and after treatment with an acetate buffer to strip cell surface proteins. GP100/D^b APNs served as a noncognate control. ****P < 0.0001 between OVA/K^b APN treatment with and without acid treatment at 4°C; n.s., not significant, where P = 0.61 between OVA/K^b APN with and without acid treatment at 37°C; two-way ANOVA and Sidak post-test and correction. All data are means ± SD; n = 3 biologically independent wells. (F) eGFP mRNA expression in P14 CD8⁺ T cells after coincubation with free-form eGFP mRNA or eGFP mRNA loaded in GP33/D^b APNs for 24 hours.