tDCs represent transendothelial cells. (A) Numbers of thymic moDCs and DCs (gated as in Fig. S4 A) in Ifnar1^−/− and Ifnlr1^−/− mice, shown as KO/WT ratio of cell numbers; n = 5–6 mice from two independent experiments. (B) Numbers of thymic moDC and DCs in Cd40l^−/− mice, shown as KO/WT ratio of cell numbers; n = 4–7 mice from two independent experiments. (C) Numbers of thymic moDC and DCs in Tcra^−/− mice, shown as KO/WT ratio of cell numbers; n = 4 mice from two independent experiments. (D) Frequency of eGFP⁺ thymic moDC and DCs in Mx1^eGFP mice; n = 5–7 mice from three independent experiments. (E) Representative flow cytometry plots showing thymic SIRPα⁺ DCs (gated as shown in Fig. S4 A) from birth (0 days) to 49-day-old mice. (F) Frequency of DC2 and tDC among total CD11c⁺ and CD11b⁺ cells in the thymus of C57BL/6 mice from birth (0 days old) to 49-day-old mice; n = 6 mice from two independent experiments. (G) Frequency of labeled thymic moDC and DCs by i.v. administration of anti–CD11c-PE antibody. Mice were euthanized 2 min after administration; n = 6 mice per three independent experiments. The cells were gated as shown in Fig. S4 F. (H) Representative confocal microscopy images comparing the localization of TCF4⁺ cells in the thymus of WT C57BL/6 and Zeb2^Δ1+2+3 mice. The association with thymic microvessels was assessed by colocalization of TCF4⁺ cells with CD31⁺ positivity. Medullary region was identified by Hoechst staining. Scale bars represent 100 μm. (I) Numbers of free and CD31-associated TCF4⁺ cells in the specific thymus area of WT C57BL/6 and Zeb2^Δ1+2+3 mice. Data are shown as mean ± SD. Statistical analysis was performed by a one-sample t test and Wilcoxon test with a theoretical mean of 1 (A–C), one-way ANOVA with multiple comparisons analysis (D and G), and a two-sided Fisher’s exact test (I); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, and n.s., not significant.