Anti human cd44 fitc

Colon cancer cell lines Caco, HT-29, SW480, SW620, LoVo, and HCT-116 were purchased from Shanghai Institute of Biochemistry and Cell Biology (Chinese Academy of Sciences, China). To analyze the expression of CD44 and CD133 in different lines, cells were routinely cultured, harvested, trypsin-digested and re-suspended in stain buffer (1 × 10⁶ cells in 80 μl). Cells were then treated with 20 μl FcR Blocking Reagent for 15 min, and incubated with antibodies (human anti-CD44-FITC and human anti-CD133-PE, Miltenyi Biotec, San Diego, CA, USA) for 30 min. After staining, cells were subjected to flow cytometry for analysis using BD Accuri C6 (CA, USA).
To isolate CD44^high/CD133^high and CD44^low/CD133^low populations in the HCT-116 line, cells were trypsinized and blocked with FcR Blocking Reagent. Propidium iodide staining was applied to exclude the dead cells. Live cells were incubated with antibodies (human anti-CD44-FITC and human anti-CD133-APC, Miltenyi Biotec, Auburn, USA) for 30 min. CD44^low/CD133^low and CD44^high/CD133^high cells were sorted by a cell sorter (BD FACSARIA III, CA, USA).
For cell cycle analysis, CD44^high/CD133^high and CD44^low/CD133^low cells were stained with propidium iodide and analyzed using a flow cytometer (BD Calibur, CA, USA) and ModFit software (Verity, Maine, USA).