Adoptive Transfer of Antigen-Specific T Cells

All mice were used in accordance with the guidelines of the Institutional Animal Care and Use Committees at the University of Minnesota. C57BL/6J mice were purchased from The Jackson Laboratory. Thy1.1⁺ P14, CD45.1⁺ OT-I, and CD45.1⁺ mice were fully backcrossed to C57BL/6J mice and maintained in our animal colony. OT-I.Ifng^−/− mice were generously provided by M. Mescher (University of Minnesota), and were generated as follows: B6.129S7-Ifng^tm1Ts/J mice, deficient for IFN-γ (Ifng), were obtained from The Jackson Laboratory and bred with OT-I mice. P14 immune chimeras were generated by transferring 5 × 10⁴ naive transgenic Thy1.1⁺ P14 T cells into naive C57BL/6J mice and infecting with 2 × 10⁵ p.f.u. LCMV Armstrong the next day. Memory OT-I cells were generated by transferring 5 × 10⁴ naïve transgenic CD45.1⁺ OT-I T cells into naive C57BL/6J mice or into memory P14 chimeras. The next day, recipients were infected with either 1 × 10⁶ p.f.u. VSV-OVA or 2 × 10⁶ p.f.u. VV-OVA i.v. For local re-challenge experiments, 50 μg of the indicated peptides or 4 × 10⁵ p.f.u. VV-gp33 or VV-OVA was delivered t.c. as described^{13 (link),23 (link)} in a volume of 35 μl delivered by modified gel loading pipet. For depleting circulating memory P14 CD8⁺ T cells, mice were injected with 3 μg anti-Thy1.1 (clone HIS51) into the peritoneal cavity. Depletion was confirmed by Thy1.1 (clone OX-7) and H-2D^b/gp33 MHC I tetramer staining.