Chinese Journal of Tissue Engineering Research ›› 2013, Vol. 17 ›› Issue (41): 7199-7204.doi: 10.3969/j.issn.2095-4344.2013.41.003

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Primary culture and identification of neonatal rat osteoblasts

Cheng Hao1, Zhang Yan-fang1, Xu Wei2   

  1. 1Department of Physics, School of Information Engineering, Guangdong Medical College, Guangzhou  523808, Guangdong Province, China; 2Guangxi Wuzhou Pharmaceutical (Group) Co., Ltd., Wuzhou  543002, Guangxi Zhuang Autonomous Region, China
  • Received:2013-04-05 Revised:2013-07-16 Online:2013-10-08 Published:2013-11-01
  • Contact: Zhang Yan-fang, M.D., Associate chief physician, Department of Physics, School of Information Engineering, Guangdong Medical College, Guangzhou 523808, Guangdong Province, China zjzyf2006@163.com
  • About author:Cheng Hao★, Master, Department of Physics, School of Information Engineering, Guangdong Medical College, Guangzhou 523808, Guangdong Province, China chenghao712@163.com
  • Supported by:

    Key Medical and Health Project of Dongguan, No. 2012105102006*; General Medical and Health Project of Dongguan Ctiy, No. 201210515200001*

Abstract:

BACKGROUND: Tissue engineering requires a lot of seed cells. Osteoblasts have become important seed cells in bone tissue engineering. However, it is difficult to culture the osteoblasts, and cell number, purity, proliferation and differentiation activity are different obtained by different culture methods.
OBJECTIVE: To identify and compare three common primary osteoblat culture methods, and to explore a method for the primary culture of osteoblasts which is easy to operate, economical and effective, in order to provide basis for the further experimental research.  
METHODS: Calvarias were dissected from newborn Sprague Dawley rats in 72 hours, and osteoblasts were isolated with collagenase digestion method, sequential digestion method and bone tissue method respectively. The morphological observation and cytochemical staining were performed, the growth curve of the cells was drawn with Cell Counting Kit-8 method, and the rate of living osteobalsts was counted with trypan blue staining.
RESULTS AND CONCLUSION: The proliferation of the insolated and cultured osteoblasts was well with typical characteristics of osteoblasts, cytochemical staining results were positive. Compared with the sequential collagenase digestion method, the collagenase digestion method presented higher production of osteoblasts and higher cell survival rate (P < 0.05), and the collagenase digestion method was easier than the sequential collagenase digestion method and cost less than sequential collagenase digestion method. Bone tissue method was the easiest method with less damage to cells, but bone tissue method presented lower production of osteoblasts and cost much more time, which cannot be used in large-scale osteoblast culture. The collagenase digestion method is a simple, efficient and ideal method for isolation and culture of primary osteoblasts.

Key words: rats, skull, esteoblasts, cell culture techniques, cell proliferation, growth charts

CLC Number: