Anti human cd90 apc

Flow cytometry was performed using anti-human CD90 APC (BD Pharmin-gen™), anti-human CD105 FITC (BioRad), anti-human CD73 PE (BD Pharmingen™), anti-human CD34 PE (BD Pharmingen™), anti-human CD31 APC (R&D Systems), anti-human CD45 FTIC (BD Pharmingen™). Experiments were performed using hASCs in passages between 5 and 6, before the seeding and after 4 days in culture on the unmodified and modified surfaces.
Cells were trypsinized, counted and re-suspended in PBS with 2% (w/v) bovine serum albumin (BSA) (Sigma) with a concentration of 2 Â 10 5 cells/100 mL and incubated with the antibodies at the concentration advised by the manufacturers. After incubation for 20 min at room temperature, protected from light, cells were washed with PBS/BSA, re-suspended in PBS with 1% formaldehyde (Sigma) and analyzed in a BD FACSCalibur™ flow cytometer (BD Biosciences). Cells of interest were gated in a forward vs. side scatter dot plot with a linear scale. Isotype controls were made to discern non-specific from specific staining. A minimum of 10,000 gated events were acquired and labeled cells were quantified. CD's variations were calculated relatively to their initial content in the hASCs before seeding, for each sample. The calculation consisted in the difference between the percentages of each CD before and after 4 days in culture, for both donors.