RM autologous barcoded HSPC transplantation and detection of GFP⁺ cells in tissues. (A) RM HSPCs were transduced with a barcoded lentiviral library and transplanted into the autologous macaque after conditioning with 1000 rads TBI. The blood and tissue were sampled periodically to assess the clonal composition of HSPC-derived cells. (B) Representative flow cytometric gating strategy to measure GFP expression among leukocyte subsets. (C) The expression of GFP was determined by flow cytometry in tissue mononuclear cell suspensions after gating for CD45⁺ cells and lymphocytes and then lineage-defining markers. Representative liver samples from an unmanipulated control animal (top) and a sample from a barcoded animal ZJ31 collected 76 months after transplantation (bottom). (D) The percent GFP⁺ hematopoietic cells in the liver, gastrointestinal tract, or lymphoid tissues for the listed cell lineages is shown. Each individual animal is designated with a different symbol, including those that match animals reported in panel A, and a line connects samples obtained at the same time point. Tissue-resident myeloid cell percentage GFP positivity was not significantly different compared with B or natural killer (NK) cells in the same tissue and was significantly higher than in T cells in the tissue (using 2-way analysis of variance [ANOVA] with Dunnett multiple comparisons test, lines compare tissue-resident myeloid cells with other leukocyte subsets); ∗P < .05; ∗∗P < .01. GI, gastrointestinal; LN, lymph node.