(A) Various NKT cell subsets from the skin and dLNs were identified by flow cytometry using CD1d tetramer staining and additional markers, including iNKT (NK1.1⁺CD3⁺CD1d-tetramer⁺), type II NKT cells (NK1.1⁺CD3⁺CD1d-tetramer^–CD4⁺, which have a phenotype similar to that of NK1.1⁺CD4⁺ T cells), NKT-like NKT cells (NK1.1⁺CD3⁺CD1d-tetramer^–CD8⁺), and CD1d-tetramer^– CD4 CD8 DN NKT cells (NK1.1⁺CD3⁺CD1d-tetramer^–CD4^–CD8^–). FSC, forward scatter; SSC, side scatter. (B) The unbiased clustering tSNE algorithm was used to verify flow cytometry data in the FP skin and dLNs. The algorithm was run on concatenated total samples (infected and uninfected). iNKT cells were absent in CD1d-KO mice. (C) Total NKT cells were quantified in the skin in WT and CD1d-KO mice after infection with 2 × 10⁵ PFU of DENV2 s.c. by FP injection. DENV infection increased iNKT cells in WT but not in CD1d-KO mice in the (D) skin and (E) dLNs on days 3 and 5 after infection. (F) Quantification of CD1d^ind NKT cell subsets in the FP showed increased NKT-like cells at days 3 and 5 after infection in WT but not CD1d-KO mice, while other NKT cell subsets measured were not influenced. (G) Tracking of local versus recruited NKT-like cells in the skin, performed by CFSE labeling prior to DENV2 infection. CFSE⁺ or CFSE^– NKT-like cell (NK1.1⁺CD3⁺CD1d-tetramer^–CD8⁺) numbers in the FP skin were determined on day 3 after infection and are presented as stacked bars. iNKT and NKT-like cells were recruited into the skin during DENV infection, and both subsets were absent from the skin of CD1d-KO mice. Data are represented as means ± SEM and are for n = 5 mice combined from 2 independent experiments for each time point. *P < 0.05; **P < 0.01; ***P < 0.001, 1- or 2-way ANOVA with Holm-Šidák post test.