The frequency of in vivo expanded Ag-specific T cells and their functional activity. Three million naive CD4 OT-II (A) or CD8 OT-I (C) T cells (CD45.2⁺) were administrated intravenously into SJL mice (CD45.1⁺). The next day, recipient mice were injected with 2.5 μg anti–CD19-OVA conjugates or 50 μg isotype mAb-OVA. Splenocytes or lymph node cells were harvested at day 7 or day 15, and the percentage of CD45.2⁺ cells was enumerated by flow cytometry. Cells were gated on the CD4⁺ or CD8⁺ population. Splenocytes from mice receiving CD4 OT-II cells were harvested at day 7 or day 15 (B) and restimulated with various amounts of anti–CD19-OVA conjugates for 60 hours. Proliferation was measured by ³H-thymidine incorporation. Splenocytes from mice receiving CD8 OT-I cells were harvested at day 7 and restimulated with 10 μg/mL OVA overnight (D). The cells were stained for intracellular IFN-γ. Cells were gated on the CD8⁺CD45.2⁺ population. Mean plus or minus SE is shown (n = 3).