Mesenchymal stem cells derived from porcine bone marrow and isolated by adherence.
Bone marrow (30 ± 5 mL) was harvested from the posterior iliac crest of healthy pigs. Mononuclear cells were separated from red cells by density gradient centrifugation with Ficoll Paque (density, 1.077 g/cm3). The buffy coat was collected and washed twice for 5 minutes at 1500 rpm. The cells are plated in α-MEM medium, containing HEPES, fetal bovine serum (10%), antibiotic solution (penicillin (10,000 U), streptomycin (10 mg)), amphotericin B (25 µg) /mL), L-glutamine (200 mM), and incubated at 37°C, and 5% carbon dioxide. Cells were initially plated in 25 cm2 vented culture flasks, at a density of 2 × 106 cells/mL. After 3 days, medium and nonadherent cells were discarded.