To generate a growth curve, USCs and ADSCs (cell passage 3, 5, and 7) were seeded at a density of 500 cells/cm2. The cell proliferation rates were analyzed using the cell counting kit-8 (Dojindo, Kumamoto, Japan) according to the manufacturer's instructions at 1, 3, 5, 7, and 9 days after plating, and doubling time was also measured.
For colony forming analysis, USCs and ADSCs were seeded at a density of 1×103 in 24-well plate. After incubation for 14 days, the cells were fixed with 10% paraformaldehyde and stained with 0.5% crystal violet (Sigma-Aldrich, St. Louis, MO, USA) for 15 min. Cells were washed with distilled water and scanned plate after dry. Stained colony was dissolved in 10% acetic acid and absorbance was measured with ELISA reader (Molecular Devices, Sunnyvale, CA, USA) at 562 nm.
Flow cytometric evaluation was performed with phycoerythrin (PE)-conjugated mouse monoclonal antibodies specific for embryonic/mesenchymal stem cell marker (SSEA4), mesenchymal stem cell markers (CD44, CD73, CD90, CD105), hematopoietic stem cell markers (CD34, CD45, c-kit), and immunologic marker (HLA-DR) (BD Biosciences, San Jose, CA, USA), according to the manufacturer's instructions. Approximately 1×104 cells were measured using fluorescence-activated cell sorter (BD Biosciences) system equipped with the CellQuest program. Antibody information was listed atTable 2 .
For colony forming analysis, USCs and ADSCs were seeded at a density of 1×103 in 24-well plate. After incubation for 14 days, the cells were fixed with 10% paraformaldehyde and stained with 0.5% crystal violet (Sigma-Aldrich, St. Louis, MO, USA) for 15 min. Cells were washed with distilled water and scanned plate after dry. Stained colony was dissolved in 10% acetic acid and absorbance was measured with ELISA reader (Molecular Devices, Sunnyvale, CA, USA) at 562 nm.
Flow cytometric evaluation was performed with phycoerythrin (PE)-conjugated mouse monoclonal antibodies specific for embryonic/mesenchymal stem cell marker (SSEA4), mesenchymal stem cell markers (CD44, CD73, CD90, CD105), hematopoietic stem cell markers (CD34, CD45, c-kit), and immunologic marker (HLA-DR) (BD Biosciences, San Jose, CA, USA), according to the manufacturer's instructions. Approximately 1×104 cells were measured using fluorescence-activated cell sorter (BD Biosciences) system equipped with the CellQuest program. Antibody information was listed at