CXCR4 defines heterogeneity among BM Ly6C^hi monocytes. (A) BM Ly6C^hi monocytes were gated accordingly (left) and subjected to t-SNE dimensional reduction (middle). Automatic clustering was performed using k-means, and clusters plotted into the t-SNE map (right). (B) Indicated markers are color mapped from blue (low expression) to red (high expression) into the t-SNE map. (C) Overlayed histograms of indicated markers constructed from data obtained from the k-means of cluster 1 (blue) and cluster 2 (red) that was generated from automated clustering. (A–C) Data are representative of one out of three experiments with at least n = 3 samples. (D–E) BM Ly6C^hi monocytes in mice divided into CXCR4^hi (blue) and CXCR4^lo (red) subsets (D) with scanning electron microscopy images of mouse CXCR4^hi (top) and CXCR4^lo (bottom) subsets (E). Bars, 1 µm. Data are representative of one out of three experiments. (F) Gating strategy and division of human BM classical monocytes into CXCR4^hi (blue) and CXCR4^lo (red) subsets. Data are representative of one out of three experiments conducted with seven independent donors.