Combination therapy decreased the frequency of myeloid clusters associated with immunosuppressive phenotypes. CyTOF proteomic analysis using a myeloid cell antibody panel for TILs from mice challenged with B16-F10 cells and given indicated treatments as described in the Materials and methods. (A) t-SNE plot of an equal number of CD11b⁺ CD3⁻ TILs from each group and overlaid with color-coded clusters. (B–D) t-SNE plots with clusters of intratumoral CD11b⁺ CD3⁻ cells in each treatment group. Expression of (C) inhibitory receptors and (D) M1/M2 macrophage markers projected onto t-SNE plot in A. (E) Bar plot of the frequency of each cluster. Cluster names are indicated on the y axis, and frequencies of each cluster are on the x axis. Data represent three independent experiments (n = 4–7 mice per group; t test, *P < 0.05), and significance is shown between αCTLA-4 and αCTLA-4 + IVAX treatment groups. Error bars represent means ± SEM. Irradiated parental B16 tumor cells (Vac) were used as a control for the IVAX group.