(A) Native (nonreducing) 3%–12% gel showed the effect of the reduction of MECA79 mAb with various MECA79/TCEP ratios on its molecular weight. (B) Corresponding trafficking of MECA79-NPs to LNs, as assessed by fluorescence imaging (mean ± SEM, Student’s t test, ***P < 0.001, n = 3–5 mice per group, 2–4 LNs from each mouse). (C) Hydrodynamic diameter of anti-CD3-NPs versus MECA79-anti-CD3-NPs (mean ± SEM, Student’s t test, **P < 0.01, n = 3 per group). (D) Sustained release profile of anti-CD3 from NPs over a duration of 14 days (n = 3 per group). (E) Western blot analysis of DLNs showed a significant increase in anti-CD3 expression in the MECA79-anti-CD3-NP–treated group (5 μg of anti-CD3) in comparison with the no-treatment and free anti-CD3 groups (5 μg of anti-CD3). β-Actin was used as a loading control. The fold change over the syngeneic sample was calculated after densitometric analysis and correction for β actin (control vs. free anti-CD3 vs. MECA79-anti-CD3-NP: 0 vs. 0.87 ± 0.09 vs. 1.43 ± 0.14, mean ± SEM, Student’s t test, ***P < 0.001, n = 4 mice per group). (F) *anti-CD3-NPs with or without MECA79 antibody conjugation were injected into skin-transplanted mice. The DLNs from MECA79-*anti-CD3-NP–treated mice contained a significantly greater percentage of *anti-CD3–labeled T cells and DCs than controls. No differences were seen in *anti-CD3–labeled B cells (mean ± SEM, Student’s t test, ***P < 0.001, ****P < 0.0001, n = 4 mice per group). (G) DCs were cultured in vitro with *anti-CD3 or *anti-CD3-NPs, washed, and reincubated with CD4⁺ and CD8⁺ T cells, and the degree of T cell labeling was analyzed. Significantly greater labeling with *anti-CD3 was observed in both CD4⁺ and CD8⁺ T cells incubated with *anti-CD3 NP cultured DCs as compared with those cultured with *anti-CD3 alone (*anti-CD3 vs. *anti-CD3-NP: 2.08 ± 0.35 vs. 39.5 ± 2.01 for CD4⁺, 2.52 ± 0.23 vs. 26.2 ± 1.97 for CD8⁺, mean ± SEM, Student’s t test, ****P < 0.0001, n = 4 per group).