Figure 2. FLDCs migrate to the dLN and induce Ag‐specific responses following adoptive transfer.

Following Ag stimulation, dsRed⁺ FLDCs were injected s.c. into naïve WT mice, and 48 h later, dLNs were harvested and the presence of dsRed⁺ CD11c⁺ cells was assessed by flow cytometry (A, B) or confocal microscopy (E).

• A
  dsRed⁺ CD11c⁺ FLDCs were gated as pDCs (CD45R⁺), cDC1s (CD45R⁻ CD24⁺), or cDC2s (CD45R⁻ CD11b⁺).
• B
  Absolute numbers of dsRed⁺ CD11c⁺ FLDCs in the dLN calculated from flow analysis and cell counts.
• C
  Percentage of transferred dsRed⁺ CD11c⁺ cells that were cDC1s or cDC2s.
• D
  CD86 expression on transferred dsRed⁺ CD11c⁺ FLDCs.
• E
  Confocal microscopy of dLN sections after FLDC transfer: Upper row depicts overlay of CD3 (green), CD45R (gray) and dsRed (red); bottom row depicts dsRed (red) alone. White dashed line represents division between T cell (CD3⁺) and B cell zones (CD45R⁺). Scale bars represent 38 μm.
• F, G
  Seven days after transfer, dLN cells were restimulated for 72 h with 15 μg/ml SEA (F), 1 μg/ml St (G), or medium alone (M) and cytokine production assessed by ELISA.

Data information: Results are least squares mean ± SEM (B–D) or mean ± SEM (F, G). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (analyzed using a three‐way full‐factorial fit model, with contrast analysis used to test differences between experimental groups (B–D) or one‐way ANOVA (F, G)). Data from one of three or more experiments (A, E–G) or three experiments pooled (B–D) (n = 3–9 animals per group). gMFI, geometric mean fluorescence intensity.