Splenic pDCs capture and degrade Ags efficiently. (A) Spleen cells were incubated for 1 hour in the presence of labeled Ags at 37°C in culture medium (CM) or at 4°C in CM containing 0.1% azide. Cells were stained with anti-CD11c and anti-B220 mAbs and analyzed by flow cytometry. DC subsets were gated as indicated. (B) Top panel: fluorescent intensity of labeled Ags was analyzed for each gated DC subset incubated at 37°C (thick line histogram) or at 4°C in CM containing 0.1% azide (filled histogram). Bottom panel: the percentage of cells gated in M1 that have captured Ag is plotted for each subset. Results represent cumulative data from 4 independent experiments and are expressed as mean percentages plus or minus SD. *P < .05, ***P < .001. (C) Spleen cells were incubated with Lucifer Yellow (0.4 mg/mL) in the presence (dotted line histogram) or absence (thick line histogram) of inhibitor (dimethyl amiloride, 100 μM) at 37°C or at 4°C (filled histogram), and fluorescent intensity was analyzed for both DC subsets. Lucifer Yellow⁺ cells were gated as shown. (D) Percentage of Lucifer Yellow⁺ cells in each condition is plotted. Results represent cumulative data from 4 independent experiments and are expressed as mean percentages plus or minus SD.