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CD20 antibody depletion of B lymphocytes effectively ameliorates multiple T cell-mediated autoimmune diseases through mechanisms that remain unclear. To address this, a mouse CD20 antibody that depletes >95% of mature B cells in mice with otherwise intact immune systems was used to assess the role of B cells in CD4⁺ and CD8⁺ T cell activation and expansion in vivo. B cell depletion had no direct effect on T cell subsets or the activation status of CD4⁺ and CD8⁺ T cells in naive mice. However, B cell depletion impaired CD4⁺ T cell activation and clonal expansion in response to protein antigens and pathogen challenge, whereas CD8⁺ T cell activation was not affected. In vivo dendritic cell ablation, along with CD20 immunotherapy, revealed that optimal antigen-specific CD4⁺ T cell priming required both B cells and dendritic cells. Most importantly, B cell depletion inhibited antigen-specific CD4⁺ T cell …