Fitc anti mouse sca 1

EPC mobilization was performed with a Calibur flow cytometer (Becton-Dickinson, San Jose, CA, USA), fluorescence-activated cell sorting (FACS). Briefly, peripheral blood was incubated with fluorescein isothiocyanate (FITC) anti-mouse Sca-1 (eBioscience, San Diego, CA, USA) and phycoerythrin (PE) anti-mouse Flk-1 antibodies (VEFGR-2; eBioscience). Isotype-identical antibodies served as controls (Becton-Dickinson, Franklin Lakes, NJ, USA). Circulating EPCs were double-positive gated for Sca-1 and Flk-1 [13 (link)], each analysis included 100,000 events. In cell study, hyperglycemia condition was around 25 mM glucose (glucose 20 mM + medium 5 mM).

We wanted to understand EPC mobilization in a hyperuricemia model in response to tissue ischemia (72 hours after surgery). We collected peripheral blood MNCs and examined them with a fluorescence-activated cell sorter (FACS Calibur, Beckman Cytomics FC 500). We used fluorescein isothiocyanate (FITC) anti-mouse Sca-1 (eBioscience, San Diego, CA, USA) and phycoerythrin (PE) anti-mouse Flk-1 (VEGFR-2, eBioscience) antibodies with fluorescein isothiocyanate (FITC). Circulating EPCs were quantified by enumerating Sca-1⁺/Flk-1⁺ cells, compared to total cell number.

The mononuclear cells were incubated with fluorescein isothiocyanate (FITC) anti-mouse Sca-1 (eBioscience) and phycoerythrin anti-mouse Flk-1 (VEGFR-2, eBioscience) antibodies at 4°C for 30 minutes. The expression of Sca-1⁺/ Flk-1⁺ cells (EPC-like cells) in mononuclear cells were analyzed by flow cytometry with a Cytomic FC 500 (Beckman-Coulter, Miami, FL). For analysis, 10⁵ circulating cells were quantified by enumerating Sca-1⁺/ Flk-1⁺ cells and were scored using a CXP software (Beckman-Coulter).