Fc500 cxp cytometer software

CDCs were harvested as single-cell suspensions using TrypLE™ Express (Thermo Fisher Scientific). Cells were then incubated with PE-conjugated mouse anti-human CD105 antibody (# 12-1057-42, eBioscience) or FITC-conjugated mouse anti-human CD90 antibody (# 11-0909-42, eBioscience) for 30 min. The percentages of CD105 and CD90 were quantitatively measured using a Cytomics FC500 instrument with FC500 CXP Cytometer software (Beckman Coulter Co.).

BM cells were collected from femurs by flushing with PBS using a syringe, filtering through a 40-μm cell strainer, and treating with ammonium chloride for hemolysis. Bone marrow cells were incubated with FITC Rat anti-Mouse CD34 (#553733, BD Biosciences, San Jose, California, USA) or FITC Rat IgG2a κ Isotype Control (#553929, BD Biosciences) on ice for 60 min. Analyses were then performed using a Cytomics FC500 with FC500 CXP Cytometer software (Beckman Coulter Co., Miami, FL, USA). CD34-positive and CD34–negative bone marrow cells were separated using a MACS Cell Separation instrument (Miltenyi Biotec, Bergisch Gladbach, Germany) with Purified Rat Anti-Mouse CD34 (#553731,BD Biosciences) and Anti-Rat IgG MicroBeads (Miltenyi Biotec).