2.5. Flow Cytometry Analysis of Hematopoietic Stem and Progenitor Cells (HSPCs)

For analysis of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs), bone marrow cells were isolated from mouse femurs with sterile PBS and filtered. Lineage⁻Sca1⁺c-kit⁺ (LSK) was employed to define HSCs as suggested in earlier studies [3, 18, 19]. Bone marrow cells (5 × 10⁶) were first labeled with biotin-conjugated antibodies recognizing Gr1, CD4, CD8, CD11b, B220, and Ter119 (mixed lineage antibodies, BioLegend, CA, USA) and then incubated with streptavidin-PerCP (BioLegend), sca1-PE, and c-kit-APC antibodies (eBioscience, CA, USA). The frequencies of HPCs and HSCs were determined via flow cytometry and analyzed by using BD Accuri C6 software (BD Biosciences, USA).