pDCs present MOG_35–55 during EAE. (A and B) LN cells were pooled from 8–10 WT and pIII+IV^−/− mice that had been immunized 2 d previously with MOG_35–55+CFA or CFA alone. The cells were then stained with antibodies against CD19, CD11c, and B220, and cDCs (CD19⁻CD11c^hiB220⁻ cells) and pDCs (CD19⁻CD11c^intB220⁺ cells) were purified by sorting. (A) Representative flow cytometry profiles are shown to illustrate the labeling and purification of pDCs and cDCs. (B) Variable numbers of purified cDCs and pDCs were incubated for 18 h with 2D2 T cells. The percentages of activated CD69^hi CD4⁺ 2D2 T cells were then quantified by flow cytometry. Results are representative of two experiments. The graph shows the means and SEM derived from two experiments. (C) MHCII (I-A^b), CD86, and CD40 expression was analyzed by flow cytometry on cDCs and pDCs isolated from the dLNs of WT mice that had been immunized 2 d previously with MOG_35–55+CFA. cDCs and pDCs isolated from the LNs of nonimmunized WT mice (day 0) were used as controls. The histograms represent the mean fluorescence intensity (mfi) and SEM derived from two experiments.