Bone marrow-derived mesenchymal stromal cells (IBGM)

The bone marrow-derived mononuclear cell fraction is isolated by density-gradient centrifugation and then resuspended. The mesenchymal stem cells are separated from hematopoietic cells by adherence, culturing in Dulbecco's modified Eagle's low glucose medium, fetal calf serum (10%) and penicillin/streptomycin (1%) in a humidified incubator, at 37°C with 5% CO₂, in 175 cm² tissue culture flasks, with periodical washing to remove nonadherent cells. When cells reach 80% confluence, they are trypsinized and replated, and the process is repeated for two more passages.