CMML peripheral blood mononuclear cells. (A-B) Peripheral blood mononuclear cells were selected from a healthy donor (control) and a CMML patient before AutoMacs negative selection for monocyte enrichment. (A) Side/forward scatters were analyzed by flow cytometry. (B) Flow cytometric analysis of CD14 and CD24 identifying CD14⁺/CD24⁻ in the healthy donor and an additional population of CD14⁻/CD24⁺ cells in the CMML sample. (C) CD14⁺/CD24⁻ and CD14⁻/CD24⁺ CMML cells were further enriched through positive selection based CD14 expression. (D) RQ-PCR analysis of CD14 and CD24 gene expression in cells sorted as in panels B and C (mean ± SD of triplicates). Each panel shows 1 representative of 20 samples. (E) RQ-PCR analysis of indicated genes in CD14⁺/CD24⁻ and CD14⁻/CD24⁺ cells sorted from 10 patients. Expression normalized to 1 in CD14⁺/CD24⁻; the ratio of expression in CD14⁻/CD24⁺ cells is shown. Bar represents median. (F) TET2 gene sequence was analyzed in CD14⁺/CD24⁻ and CD14⁻/CD24⁺ cells sorted from 19 patients. Indicated abnormalities were found in all cases in the 2 cell populations.