Most BM HSCs bind to Angptl3. (A-B) Freshly isolated adult BM Lin⁻ cells isolated by AutoMACS were incubated with 1500 ng/mL Angptl3 and then stained with goat anti-Angptl3 antibody, followed by anti–goat-APC, anti–Sca-1-PE/CY5.5, anti–c-Kit-FITC, and anti–CD150-PE (or anti–Flk-2-PE, and anti–CD34-PE) antibodies. (A) Flow cytometry plots demonstrating that 83% Lin⁻Sca-1⁺Kit⁺CD150⁺ cells bound to Angptl3 compared with controls. (B) Summary of the percentages of total BM, Lin⁻, Lin⁺, and LSKCD34⁻FLK-2⁻ cells (LSKFC) or LSKCD150⁺ cells (LSKCD150) that bound to Angptl3. (C-E) Freshly isolated adult CD45.2 BM cells were incubated with Angptl3 and then stained with goat anti-Angptl3 and anti–goat-Cy5 antibodies. Incubation without goat anti-Angptl3 and cells not treated with Angptl3 served as negative controls. (C) Thirty thousand positively stained cells and the same number of negatively stained cells were transplanted together with 1 × 10⁵ CD45.1 competitor cells into lethally irradiated (10 Gy) CD45.1 mice (n = 8-9). Peripheral blood engraftments are shown at 4, 12, and 24 weeks after transplantation. (*statistically significant difference between 2 groups, P < .05). (D) Representative FACS plots of peripheral blood mononuclear cells, demonstrating the multilineage contributions of Angptl3-binding HSCs in 1 mouse at 24 weeks after transplantation. (E) Multilineage contribution of Angptl3-binding cells at 12 weeks after transplantation (n = 9).