CD25 expression marks distinct subsets of cells in the CML LIC population. (A) Screening strategy for antigens differentially expressed on CML LSK cells. LSK cells were transduced with BCR-ABL retroviral vector and transplanted into recipient mice. Bone marrow cells were collected 10 to 12 days after transplantation, and staining patterns of candidate antigens in 3 types of cells (BCR-ABL⁺LSK, LKS⁻, and BCR-ABL⁻LSK) were analyzed by flow cytometry. (B) CD25⁺ cells in the 3 cell types in panel A (%) (means ± standard deviation [SD], n = 3). (C) Flow cytometric analysis of CD25 expression in normal bone marrow fractions, including LSK, GMP (Lin⁻c-Kit⁺Sca-1⁻CD34⁺FcγRII/III⁺), and CD4⁺ cells (orange histograms). Gray histograms indicate isotype control staining. Numbers indicate the CD25-positive fraction (means ± SD, n = 3). (D) Flow cytometric analysis of BCR-ABL⁺LSK cells from CML model mice costained with CD25 and FcεRIα. Giemsa-stained cytospin specimens of the 3 fractions (CD25⁺F⁺LSK, CD25⁺F⁻LSK, and CD25⁻F⁻LSK cells) are shown. Images were obtained and analyzed using a microscope (IX70; Olympus). UPlanApo 40×/0.85 objective lens (Olympus) and DP Controller (Olympus). (E) qPCR analysis of mast cell–related genes in the indicated fractions from CML model mice (means ± SD, n = 4). *P < .05; **P < .01.