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Updated: May 9, 2026
Isolation, Purification, and Differentiation of Osteoclast Precursors from Rat Bone Marrow
Published on: May 19, 2019
[Comparison of the activity and yield rate of osteoblast obtained by different digestion methods]
Ling-hui Li1, Dao-Fang Ding, Guo-Qing Du
1Shi's Center of Orthopedics and Traumatology, Shuguang Hospital Affiliated to Shanghai University of TCM, Shanghai 201203, China.
Objective:
To compared the activity and yield rate of osteoblast obtained by different collagenase digestion methods, to find a better way to extract osteoblast for the experimental researches of osteoporosis.
Methods:
Ten 24-hour-old SD rats were were euthanized. The cranium of rats were removed and cuted into blocks of 1 mm x 1 mm size. After digested by trypsin for 15 min, all the cranium were divided into two equal parts, and randomly divided into two groups which would be digested by type I collagenase and type II collagenase separately for two times. The rat cells of the two groups were cultured in thermostat incubator with 5% CO2 under the condition of 37 degrees C. The primary culture osteoblasts were counted by using a haemacytometer after digestion and 72 hours later. The second generation osteoblasts cultured 48 h were dyed by NBT/BCIP staining solution, and were detected by quantitative measurement with PNPP.
Results:
The cells had irregular shapes. The results of cell counting showed that the cell number of type I group was larger than type 11 group. Alkaline phosphatase dyeing were positive. Detecting of alkaline phosphatase using the method of PNPP showed that the absorbance value in type I group were higher than type II group (P<0.05).
Conclusion:
Two types of collagenase are both suitable for the in vitro culture of rat osteoblasts. The activity and yield rate of osteoblasts in type I group are higher which could provide more stable seed cells for the treatment of osteoporosis.
