Depletion of CD4 T cells enhances lymphocyte activation and NK cell activity. Mice were treated as described in Figure 2, except that CD4 depletion was performed as described in “Materials and methods.” (A) Histograms of the activation markers of lymphocytes isolated from Peyer patches of pH60/Surv-vaccinated mice either with (dark lines) or without (shaded areas) CD4 depletion are shown. (Left) Gated on CD11c⁺I-A/I-E⁺ cells. (Middle) Gated on CD3⁺CD8⁺ cells. (Right) Gated on CD3^-DX5⁺ cells. ^*P < .01 compared with the nondepletion group. All increases were confirmed in a separate experiment using another 2 mice in each group. (B-C) Mice vaccinated with pH60/Surv were not depleted (▪) or were depleted of CD4 T cells before vaccination (▴) or after vaccination (▵). (B) Freshly isolated splenocytes were used in a standard ⁵¹Cr-release assay against Yac-1 NK target cells. ^*P < .05 compared with the nondepletion or postvaccination depletion groups. (Top panels) CD8 and DX5 distribution of splenocytes obtained from undepleted or CD4-depleted mice. (C) Splenocytes were stimulated in vitro for 5 days with irradiated CT-26 cells and were used in a standard ⁵¹Cr-release assay against these tumor cells. ^*P < .05 for the prevaccination and postvaccination depletion groups compared with the nondepletion group. All experiments were repeated at least once and yielded similar results.