(A) Immunohistochemistry of spleens and lymph nodes from control mice and mice that received Tvax expressing GFP. T cells were injected i.v., and 48 hours later, spleens and lymph nodes were harvested and stained for GFP by immunohistochemistry. Original magnification, ×20. (B) Tvax cells were created from WT (no antigen) or full-length OVA-expressing donor mice and labeled with CellTrace Violet (CTV), and 2 × 10⁶ cells were transferred i.v. into mice. The percentage Tvax cells as a fraction of CD3⁺ T cells in spleen and lymph nodes was measured by flow cytometry in mice sacrificed at the indicated time points (n = 4 per group). Error bars represent the SEM. (C) Staining of tCD19 and SIINFEKL-H-2Kb on T cells from WT and B2m^–/– donor mice transduced with tCD19-OVA-LLO190. (D) Tvax cells from WT and B2m^–/– donors were injected into mice, and the percentage of antigen-specific cells in the blood was measured by tetramer staining. (n = 10). (E) Schematic of dye transfer experiment to determine DC uptake of Tvax cells. (F) Splenic DCs were enriched by negative immunomagnetic selection 48 hours after injection of DiI-labeled Tvax cells, and DiI uptake by CD11c^hiPDCA^– cells was measured in CD8a⁺ and CD8a^– cell subsets by flow cytometry. (G) Naive OT-I CD8⁺ T cells were labeled with CFSE and incubated with CD11c^hiDiI⁺ and DiI^– cells sorted by FACS from mice 48 hours after vaccination. CFSE dilution (left panel) and CD44 expression (right panel) in OT-I T cells was measured 4 days later by flow cytometry. Results are representative of 3 biological replicates from 2 independent experiments. *P < 0.0001, by Mann-Whitney U test.