Chronic Ag-stimulated virus-specific CD8+ T cells recovered function after Ag rest. A total of 1 × 105 OT-I TCR Tg T cells were transferred to naive C57BL/6 mice. Mice were single or chronic stimulated with 1 × 106 TCID50 WSN-OVA, by i.p. injection, for 30 days. On day 30, chronic animals were Ag rested for 45 days, and an equal number of single or chronic Ag-rested virus-specific CD8+ T cells were transferred to naive C57BL/6 mice, which were then i.n. challenged with WSN-OVA and harvested on day 7. A, Representative flow cytometry (gated on CD8+ T cells) from lungs of single or chronic rested infected mice is shown. As indicated by Vα2+Vβ5.1+ cells, chronic Ag-rested CD8+ T cells expanded equivalently to single prime animals. B, Pooled absolute numbers indicate that Ag rest recovers not only frequency, but also the number of chronic rested virus-specific CD8+ T cells (n = 3 mice/group). C, Representative flow cytometry histogram (gated on CD8+ T cells) shows equivalent MFI of IFN-γ production after ex vivo stimulation with OVA254–267-specific peptide.