Figure 4. BFP-CXCL10⁺ myeloid cell clusters enriched for CD11c⁺MHC-H⁺ cells.

(A) Representative dot plots of BFP-CXCL10 and RFP-CXCL9 expression by CD11b⁺ cells within the day-5 OVA/CFA-immunized REX3 dermis.

(B) Representative histograms of MHC-II expression by BFP-CXCL10⁺ (blue) and BFP-CXCL9⁺ (red) CD45⁺ cells from (A). Gray: RFP-CXCL9⁻ BFP-CXCL10⁻ CD45⁺ cells.

(C) tSNE analysis on singlet live CD45⁺ cells using concatenated multi-color flow cytometry data.

(D) Immune subsets gated by manual flow cytometry analysis displayed on tSNE plots (left). Overlay of BFP-CXCL10⁺ cells (right).

(E) Frequency of BFP-CXCL10⁺ cells (left), BFP-CXCL10⁺RFP-CXCL9⁺ cells (middle). and BFP-CXCL10⁻RFP-CXCL9⁺ cells (right) within each of the immune subsets in (D). Bars represent means ± SEM of three independent experiments, three to four mice/experiment.

(F) Density plots for BFP-CXCL10, RFP-CXCL9, and MHC-II within each subtype in (E).

(G–I) MHC-II⁺ (orange) staining relative to the BFP-CXCL10⁺ cluster (G), cluster analysis of BFP-CXCL10⁺ and MHC-II⁺ cells (H), MHC-II MFI of cells (I) relative to the BFP-CXCL10⁺ clusters in (H). Representative maximal z projection images by IV-MPM from two independent experiments, two to three mice/imaging session, >10 imaging volumes; scale bar, 50 μm.

(J) Colocalization of MHC-II⁺, BFP-CXCL10⁺ and Th1 cells. Right panel (processed image), cluster analysis of MHC-II and BFP-CXCL10 cells using the DBSCAN-based algorithm. Representative maximal z projection images by IV-MPM from two independent experiments, two to three mice/imaging session, >10 imaging volumes; scale bar, 50 μm.

(K) CD11c⁺ (yellow) cells relative to BFP-CXCL10⁺ clusters (blue); day-5 OVA/CFA-immunized REX3 ears.