UCSCs were obtained from Cyagen Biosciences Inc. (HUXUC-01001, Cyagen). The UCSCs were placed in a 6-well plate and cultured [31 (link)] in MEM (12571071, GIBCO) complete medium containing 10% fetal bovine serum (FBS; 10099141C, GIBCO) and 1% penicillin-streptomycin (15140122, GIBCO) with 5% CO2 at 37 °C. The media was changed every three days until the cells were confluent, and the cells were then digested and passaged using pancreatin (25200056, GIBCO).
Cells at passage 3 (P3) were collected to identify UCSCs surface markers including CD90-fitc (Invitrogen, Article No. 11-0909-41), CD29-pe-cy5 (BD PHOSFLOW, Article No. 559882), CD73-pe (Invitrogen, Article No. 12-0739-41), CD105-apc (Invitrogen, Article No. 17-1057-41), and HLA-apc-cy7 (Invitrogen, Article No. 47-9956-41). Cells were incubated in 100 μL staining solution at 4 °C for 30 min. We added 1 ml phosphate buffered saline (PBS) and centrifuged the cells at 400×g for 5 min. The supernatant was discarded, and the cells were suspended in 200 μL PBS for flow cytometry (FCM) analysis (Becton Dickinson, Aria II).
Cells at passage 3 (P3) were collected to identify UCSCs surface markers including CD90-fitc (Invitrogen, Article No. 11-0909-41), CD29-pe-cy5 (BD PHOSFLOW, Article No. 559882), CD73-pe (Invitrogen, Article No. 12-0739-41), CD105-apc (Invitrogen, Article No. 17-1057-41), and HLA-apc-cy7 (Invitrogen, Article No. 47-9956-41). Cells were incubated in 100 μL staining solution at 4 °C for 30 min. We added 1 ml phosphate buffered saline (PBS) and centrifuged the cells at 400×g for 5 min. The supernatant was discarded, and the cells were suspended in 200 μL PBS for flow cytometry (FCM) analysis (Becton Dickinson, Aria II).
Free full text: Click here
