Treg depletion or adoptive cell transfer to CD3^−/− mice is not sufficient to restore an Ova-specific CD8⁺ T cell response.

(A) B6 (n = 3) and SM-Ova (n = 3) mice were injected i.p. at day −4 and day −1 with 250 µg of anti-CD25 (PC-61) antibody to inactivate endogenous Treg, or with PBS, followed by the injection of rAAV-Ova at day 0. Splenocytes from treated mice were collected 14 days after immunization and analyzed as before by flow cytometry for the presence of Ova-specific CD8⁺ cells using H-2K^b/Ova_257–264 pentamer staining. Representative FACS profiles and bar graphs are shown and numbers correspond to percentages of cells positively stained with the H-2K^b/Ova_257–264 pentamer in the gated CD8⁺ population. (B) 2×10⁷ purified CD8⁺ T cells originating from B6 or SM-Ova donor mice were adoptively transferred into syngenic CD3^−/− recipient mice (n = 6) together with 3×10⁷ purified CD4⁺ T cells originating exclusively from B6 donors. Recipient CD3^−/− mice were immunized with rAAV-Ova one day later and splenocytes were collected, enumerated and analyzed 14 days after by flow cytometry to evaluate the percentages and absolute numbers of Ova-specific CD8⁺ cells. Representative profiles gated on the CD8⁺ population are shown in the left panel and the total numbers of Ova-specific CD8⁺ per spleen are shown in the right bar graph.