Quantification of indirect alloAg presentation using TEa Tg T cells. (A) Lethally irradiated CD45.1⁺ BALB/c or B6 recipient mice were transplanted at day 0 with 1-2 × 10⁶ CFSE-labeled CD45.2⁺ TEa T cells, with or without CD45.1⁺ B6 BM. Proliferation of TEa T cells (gated on CD45.2⁺Vα2⁺Vβ6⁺) was assessed by CFSE dilution 72 hours after transplantation, as shown. (B) Animals received primary grafts (day 0) as labeled. Fourteen days after transplant, 2-5 × 10⁶ TEa Tg cells were adoptively transferred and proliferation (gated on CD4⁺Vα2⁺Vβ6⁺) was assessed by CFSE dilution 72 hours later. Data shown are representative of a minimum of 3 replicate experiments. (C) TEa Tg cells were adoptively transferred at the time points indicated and CFSE dilution was analyzed using Modfit software. Indices of proliferation are shown (combined data from 3-6 replicate experiments; n = 5-9 for each group). The dotted line represents the proliferation index in syngeneic recipients where there is no alloantigen presentation. Error bars represent SEM. All indices of proliferation are significantly different from each other (P ≤ .03), except day 7 versus day 14, where P = .07.