B220 ^high , but not B220 ^low B cells are depleted by anti-CD20 therapy in EAE mice. (A) Clinical scores were tracked in mice in which EAE was induced by adoptive transfer of 10 million Th17-skewed encephalitogenic T cells into young SJL/J mice randomized to i.v. treatment with either anti-CD20 monoclonal antibody (n = 20) or isotype control antibody (n = 17) on days −1, 4, 9, 14, and 19 of EAE. (B) Mice were then euthanized at the postacute phase (day 24 of EAE), and the absolute number of IgM⁺IgD⁺B220^high, class-switched B220^high, and B220^low B cells in the femur/tibia BM, skull BM, dura, LM, and brain were analyzed by flow cytometry. (C) The ratio of IgM⁺IgD⁺B220^high to B220^low B cells and class-switched B220^high to B220^low B cells is also reported. The results are data from one experiment. Results are expressed as absolute numbers of whole tissue. Error bars indicate mean ± SEM. A Mann–Whitney test was conducted to assess statistical significance (* = P < 0.05, ** = P < 0.005, *** = P < 0.0005, **** = P < 0.0005, and no label = not significant).