大鼠胎鼠成骨细胞原代培养方法研究

Primary culture of osteoblasts from fetal rat

目的探索一种纯度高、数量大、操作简便的大鼠原代成骨细胞培养方法,为后续的实验研究奠定基础。方法 2009年3—9月在中国科学院金属研究所生物实验室,选择出生24h内的大鼠胎鼠4只,摘取颅骨,采用胰蛋白酶和Ⅰ型胶原酶交替消化分离成骨细胞并进行传代培养。相差显微镜下观察原代培养的成骨细胞形态;并采用碱性磷酸酶(ALP)染色、Ⅰ型胶原免疫荧光染色及茜素红染色对培养的细胞进行鉴定。结果该方法培养出的细胞具有典型的成骨细胞形态特征,ALP染色、Ⅰ型胶原免疫荧光染色及茜素红染色均表现为阳性。结论酶交替消化法原代培养胎鼠成骨细胞,获得的成骨细胞纯度高、数量大,可作为一种相对可靠、有效的原代成骨细胞培养方法。

Objective To establish a simple, quantities, purity proposal for osteoblast primary culture, and to lay the foundation for subsequent experimental studies. Methods From March to September 2009, at the biological laboratory of Institute of Metal Research, Chinese Academy of Science, the skull was taken from 4 fetal rats which were born within 24 hours. Use trypsin and collagenase I in turn to digest osteoblasts. Using ALP staining, collagen I immunofluorescence staining and alizarin red staining to identify the cultured cells. Results The cultured cells had typical morphological characteristics of osteoblasts; ALP staining, collagen I immunofluorescence staining and alizarin red staining were positive. Conclusion Osteoblasts achieved by this method are pure with large quantities, so this method can be used as a reliable and efficient way of primary culture.