Splenic CD8α⁺ and CD8α^– DCs do not expand after Flt3-L treatment in Cl 13–infected mice. (A) Dot plots show double staining for CD11c and CD8α molecules on splenocytes from mice treated with Flt3-L or PBS, which was initiated at day 15 after infection. Boxes labeled a denote CD11c⁺CD8α^– (myeloid) DCs, and boxes labeled b denote CD11c⁺CD8α⁺ (lymphoid) DCs. (B) The fold expansion of CD11c⁺CD8α⁺ (boxes labeled b) and CD11c⁺CD8α^– (boxes labeled a) DCs is plotted for naive, Cl 13–infected, and ARM-infected mice, calculated as indicated in Methods. (C) DCs were isolated from spleens of mice that were treated with either PBS (filled squares) or Flt3-L (open triangles) for 15 days. The treatment began 5 days prior to infection and continued for an additional 10 days. Mice were sacrificed at days 0, 5, 10, and 15 after infection. Day 0 after infection corresponds to day 5 of Flt3-L treatment, and day 10 after infection represents the end of Flt3-L treatment. Flow cytometric analyses were performed to determine the number of splenic CD11c⁺CD8α^– (myeloid) and CD11c⁺CD8α⁺ (lymphoid) DCs. The populations were gated as shown in Figure 3A. The absolute number of CD11c⁺CD8α^– and CD11c⁺CD8α⁺ DCs are plotted for naive, ARM-infected, and Cl 13–infected mice. The data are representative of two independent experiments using nine mice per group. (D) The percentage of infected BMCs was calculated at days 3, 5, 7, and 15 after infection in Cl 13–infected (white bars) and ARM-infected (gray bars) mice. BMCs were harvested at the indicated timepoints, stained with an LCMV NP–specific antibody directly conjugated to Alexa 488, and analyzed by flow cytometry.