鼠胚成骨细胞的原代培养与鉴定

Primary culture and identification of fetal mouse osteoblasts

目的探讨经济、高效的原代成骨细胞培养方法,为进一步的实验研究奠定基础。方法孕龄19天的昆明孕鼠(KM),无菌条件下剖腹取出胎鼠,然后取胎鼠的颅盖骨,剔净、漂洗、剪碎。用0.25%胰蛋白酶消化10分钟,终止消化后,将碎骨块接种于培养瓶中培养,通过相差显微镜、透射电镜观察,并用碱性磷酸酶染色、骨钙素免疫组化染色和钙结节茜素红染色等方法对所获得的细胞进行鉴定。结果所培养的细胞具有典型的成骨细胞形态特征,碱性磷酸酶染色、骨钙素染色和钙结节染色均呈阳性。结论改良的组织块培养法可在较短时间内获得大量成骨细胞,所培养的细胞具有典型的成骨细胞形态和功能。

Objective To investigate an economical and efficient method of primary osteoblasts culture,so as to establish a foundation for further experimental study. Methods The calvaria harvested from 19 day fetal mouse was stripped off all soft tissues including the periosteam, rinsed and cut to trivial bone block under sterile condition. Then,the bone block was subcuhured in culture flask after digested with 0.25% trypsin for 10min, the acquired cells were identified with invert microscope and transmission electron microscope,alkaline phosphorase stain,immunohistochemistry stain of osteocalcin and alizarin bordeaux stain of calcified nodules. Results The cells obtained from fetal mouse calvaria were identified to be osteoblasts by invert mieroseope and transmission electron microscope,alkaline phosphorase stain , immunohistochemistry stain of osteocalcin and alizarin bordeaux stain of calcified nodule. Conclusion The modified tissue mess method is an ideal method to obtain and culture osteoblasts having typical characteristics.