scRNA-seq analysis of changes in immune cell composition in tumors of different treatment groups. Mice were challenged with B16-F10 tumors and were given indicated treatments; tumors were isolated and digested. TILs from tumors were isolated and stained with anti-CD45.2 antibody for sorting by FACS, and a 10X library was prepared and analyzed as described in the Materials and methods. (A) UMAP graph showing the clusters and annotation. (B) UMAP graph showing the clusters in each treatment group. (C) Heatmap displays selected marker expressions for each cluster. (D) Dot plots show the differential gene expression of cell-specific lineage markers in different clusters. (E) UMAP graphs showing the expression of selected markers. (F) Bar plot of the frequency of each cluster. Cluster names are indicated on the x axis, and frequencies of each cluster are on the y axis. Data are representative of two independent experiments (n = 5 mice per group). Mice within each group were polled for analysis. Irradiated parental B16 tumor cells (Vac) were used as a control for the IVAX group.