The production of LTα3 protein by donor T cells can be detected after T-cell receptor ligation. (A) Real-time PCR data showing mRNA expression of TNF, LTα, and LTβ in whole spleen, and FACS-sorted CD3⁺CD4⁺ and CD3⁺CD8⁺ donor T cells taken 7 days after allogeneic BMT. Syngeneic samples represent spleen from B6 recipients of B6 grafts. (B) Standard curve from murine LTα3 enzyme-linked immunosorbent assay, demonstrating the specificity of the assay for recombinant LTα3. Both TNF and murine LTα3 were serially diluted from a starting concentration of 10 000 pg/mL. (C) CD4⁺ T cells were isolated from naive B6.WT or B6.Lta^−/− mice, or at day 7 after BMT from B6D2F₁ recipients of B6.WT or B6.Lta^−/− BM plus T-cell grafts. Cells were stimulated with CD3/CD28 beads for 72 hours, and tissue culture supernatants were assessed for LTα3 protein. Data are mean ± SEM, from at least 6 separate experiments. *P < .05.