郑州大学学报（医学版）2017，Vol.52Issue(3)：370-375,6.DOI:10.13705/j.issn.1671-6825.2017.03.032
正常宫颈上皮细胞原代培养体系的改良
An improved culture system for the primary culture of normal cervical epithelial cells
摘要
Abstract
Aim: To explore an effective and repeatable method for the primary culture of normal cervical epithelial cells.Methods: A total of 60 normal human ectocervical tissue were collected and divided into two groups, primarily cultured using combined type Ⅱdispase and 0.25 g/mL trypsin-EDTA digestion method and type Ⅰ collagenase digestion method separately, and to compare the effectiveness of isolation, furthermore, to observe the detached effectiveness of combined dispase digestion method after digestion for different time.Cervical epithelial cells were cultured in 5% FBS dK-SFM and dK-SFM alone, respectively, to observe the growth of the primary cervical cells.The cultured cells were identified by the morphological analysis and pancytokeratin was identified by immunocytochemistry.Results: The total success rate of the primary culture in this study was 40%.The success rate, density, primary cell fusion time and purity of cells of the dispase digestion group were all higher than those of collagenase digestion group(P<0.05).The dispase digestion for 20 h was more effective than digestion for 16 and 24 h(P<0.05).The cell growth curve showed that the cells cultured in 5% FBS dK-SFM grew better and had a higher viability than cells cultured in dK-SFM alone(P<0.05).The primary cultured cells grew well and could be subcultured for five to six generations,and they also could be cryopreserved and resuscitated.Pancytokeratin positive staining was determined by immunocytochemistry.Conclusion: It is an effective and repeatable method for primary culture of normal cervical epithelial cells by using combined type Ⅱ dispase and 0.25 g/mL trypsin-EDTA digestion and 5% FBS dK-SFM medium.关键词
原代培养/宫颈上皮细胞/中性蛋白酶/角质细胞无血清培养基Key words
primary culture/cervical epithelial cell/dispase/dK-SFM分类
医药卫生引用本文复制引用
李雅,荣守华,智艳芳,樊婷婷,邱翠,李肖甫..正常宫颈上皮细胞原代培养体系的改良[J].郑州大学学报（医学版）,2017,52(3):370-375,6.基金项目
河南省医学科技攻关计划资助项目 201503112 （）
