2.5. Osteogenic Differentiation: General Culture Setting

To evaluate the osteogenic potential, 35,000 MSCs per well were transferred into 24-well plates (Nunc, Rosklide, Denmark) and cultured in osteogenic differentiation medium (86% DMEM high-glucose, 10% FCS, 100 µg/mL penicillin/streptomycin, 2.5 µg/mL amphotericin B, 0.1 µM dexamethasone (Sigma Aldrich, Steinheim, Germany), 2.5 µg/mL ascorbic acid-2-phosphate (Sigma-Aldrich, Steinheim, Germany), 10 mM beta glycerophosphate (Merck, Darmstadt, Germany).

For the individual setting, MSCs of each donor were seeded in duplicates. In the pooled setting 10 replicates were cultured. Quantification of osteogenic differentiation was performed on day 1 (D1), 7 (D7), 14 (D14) and 21 (D21). Media were changed twice per week.