Isolation of osteoblasts from human bone.

Isolation of osteoblasts from human bone.

Bone samples were cleaned of adherent soft tissue and osteoblasts isolated by two methods.

a. The bone was cut into small pieces (2 mm × 2 mm), washed in Ca 2+ - and Mg 2+ -free digestion's solution (10 min), and then sequentially digested (1 mg/ml trypsin, 10 min; 2 mg/ml Dispase, 20 min; and 3 mg/ml collagenase, 2 × 30 min) in digestion's solution at 37°C. Cells released by the collagenase digestions were washed and grown to confluence in 25 cm culture flasks (Falcon) in Primary cell culture system supplemented with 10% fetal calf serum (FCS). Incubations were carried out at 37 ° C in a humidified atmosphere of 5% CO 2 / 95% air; the medium was changed every 2-3 days.

b. Bone samples were cut into small pieces and plated into 5 cm Petri dishes containing 4 ml primary cell culture system supplemented with 10% FCS. Bone cells were allowed to grow out from the explants until confluent; the medium was changed every 2-3 Days.

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