Flow Cytometric Characterization of Stem Cells

A total of 1 million passage 3 cells were resuspended in 200 µL of PBS containing fluorescein isothiocyanate– or phycoerythrin-coupled antibodies against CD34, CD44, CD45, CD90, CD147, CD271 (BD Biosciences), CD117 (eBioscience), CD105, CD166 (Ancell), and vascular endothelial growth factor receptor–2 (VEGFR-2; R&D Systems) and incubated for 30 minutes at 4°C. The cells were then washed with PBS and resuspended in 350 µL of PBS for analysis. Fluorescein isothiocyanate– or phycoerythrin-coupled nonspecific mouse Immunoglobulin G (BD Biosciences) was used as an isotype control. Cell fluorescence was evaluated via flow cytometry using an EC800 instrument (SONY), and data were also analyzed using the EC800 instrument.^{9 (link),23 (link),24 (link),32 (link)}