Flow cytometric analysis of MHCII expression on thymic, splenic, and LN DCs in MHCII-deficient mouse strains. (A) Cells from collagenase-digested thymi (top row) and spleens (bottom row) were stained for CD11c and I-A^b. Shown are histograms of I-Ab expression on CD11c⁺ cells of the indicated mouse strains (bold line) or the Aα^-/- control (dotted line). The percentage of MHCII-expressing cells in the indicated gates is given as bold numbers (those for the Aα^-/- controls are given in parenthesis in the diagram on the left). (B) Mean frequencies ± SD of MHCII⁺ cells among thymic CD11c⁺ DCs of 3 mice analyzed in each group. To control for background differences in the analysis of the RFX5/CIITA double-deficient (CR^-/-) mice, we also examined CIITA^-/- RFX5^+/- mice, which were derived from the same breeding stock as CR^-/- mice. (C) Cells from collagenase-digested brachial and inguinal LNs were magnetically enriched for CD11c⁺ cells and subsequently stained for CD11c, DEC205, and I-A^b. Top: The histograms show I-A^b expression of CD11c⁺ cells (bold line, bold number). The dotted line indicates I-Ab staining on control cells from Aα^-/- animals. Bottom: CD11c and DEC205 expression on I-A^b-expressing LN cells. (D) The expression of I-A^b α- and β-chain in magnetically purified DCs from collagenase-digested spleens and thymi of the indicated mouse strains was analyzed by qRT-PCR. Analysis of β-actin expression was used for standardization. Shown are expression levels relative to the ones in DCs from C57BL/6 mice. The error bars indicate the standard deviation.