DL1-expressing Ccl19⁺ splenic stromal cells regulate Notch-mediated differentiation of Esam⁺ DCs. (A) Representative FACS profile of splenic DCs (gated as CD11c⁺B220⁻) of control, DL1^+/ΔCcl19 and DL1^Δ/ΔCcl19 mice. (B) Bar diagrams show relative (left) and absolute numbers (right) of total splenic DCs (means of 4–5 mice per group, of two independent experiments). (C) Representative FACS profiles of Esam expression on total splenic CD11c⁺ DCs (top) and the different DC subsets (bottom), classified with CD8α and CD11b expression. Percentages represent mean ± SD of 5 animals per group of two independent experiments. (D) Bar diagrams show relative (top) and absolute numbers (bottom) of Esam⁺, CD11b⁺, and CD8α⁺ DCs from spleens of control, DL1^+/ΔCcl19 and DL1^Δ/ΔCcl19 mice (mean ± SD of 4–5 animals per group of two independent). (E) In vivo priming of CD4⁺ T cells in absence of Esam⁺ DCs. OT-II CD4⁺ T cells were purified, labeled with CFSE, and injected i.v. into control and DL1^Δ/ΔCcl19 mice that were subsequently immunized with OVA and LPS as adjuvant. 3 d later, splenic cells were analyzed for the proliferation of the CFSE-labeled OT-II T cells. Representative data of one experiment are depicted and percentages of each cell division are shown as mean ± SD of two independent experiments with at least three animals per group. *, P < 0.05; **, P < 0.01. Unpaired Student’s t test was used to determine significance.