Anti cd48 hm48 1

Peripheral blood was extracted by retroorbital bleeding and blood was collected in EDTA-coated tubes. Red blood cells were lysed with ACK buffer (Quality Biological 118-156-101) for 5 min on ice. Bone marrow was isolated the tibia, femur, and pelvic bones by spinning cleaned bones for 2 min at 8,000 rpm. Single cell suspensions of spleen and thymus were prepared in FACS buffer (3% FBS in PBS) by crushing the organ through a 70-µm strainer with a plunger. Cells were blocked with rat IgG (I8015; Sigma-Aldrich) for 20 min on ice. Staining for cell surface antigens was performed by adding fluorochrome- or biotin-conjugated antibodies on ice between 20 min and overnight on ice in the dark. The following antibodies were used in this study: anti-CD11b (M1/70), anti–Gr-1 (RB6-8C5), anti-B220 (RA3-6B2), and anti–Sca-1 (D7) all from BD; anti-CD71 (RI7217), anti–Ter-119 (116228), anti–c-Kit (CD117, 2B8), anti-CD150 (TC15-12F12.2), and anti-CD48 (HM481), all from BioLegend; and anti-CD3 (145-2C11) from Thermo Fisher Scientific.