Research Paper Volume 15, Issue 9 pp 3356—3380

Figure 1. CyTOF immunophenotyping of the aging mouse prostate. (A) Workflow for prostate immunophenotyping using CyTOF. Mouse prostates of different ages were isolated, dissociated to single cells, and stained with a panel of metal-tagged antibodies before data acquisition via mass cytometry. Bivariate plots show gating for single, live CD45⁺ immune cells before analysis using clustering algorithms (t-SNE and X-Shift). Percentages represent the fraction of events that are within the gate in each bivariate plot. ¹⁹¹Ir/¹⁹³Ir labels DNA of all cells to distinguish singlets from doublets. ¹⁰³Rh labels DNA of dead cells. This flowchart was adapted from a previous publication from our group [12]. (B) t-SNE plot generated from the immune cells from mouse prostate, bladder, and kidney. Left: Heat maps showing expression of selected lineage markers by immune cells clustered using t-SNE. See Supplementary Figure 2A for the full set of markers. Right: t-SNE plot separated into 3 broad groups of immune cells (T cells, B cells, and myeloid/NK cells). (C–E) Quantification of changes to the mouse prostate immune cell composition during adult aging for CD3⁺ T cells (C), B220⁺ B cells (D), and myeloid/NK cells (E). Spearman correlation coefficient (ρ) and associated p-value (P_ρ) represent the correlation between % immune cell type and age. Data represents mean ± SD of 4 biological replicates at each age. Kruskal-Wallis, p < 0.01 (T cells, B cells, and Myeloid/NK cells). Dunn’s multiple comparisons test against 3-months-old, ^*p < 0.05, ^**p < 0.01.