奶牛乳腺上皮细胞体外培养条件优化的初步研究

doi:10.7668/hbnxb.2010.S1.019

摘要/Abstract

摘要： 从培养基、细胞接种浓度、冻存液三方面对奶牛乳腺上皮细胞体外培养条件进行了优化。采用台盼兰染色计数方法绘制细胞生长曲线,评定细胞体外生长增殖。结果表明,使用DMEM/F12培养基细胞生长状况最好,DMEM培养的细胞较DMEM/F12生长缓慢,F12其次,RPMI1640培养细胞生长最慢;以1×10⁴的接种浓度于24孔板,细胞生长状态最好,细胞的生长周期经历了潜伏期、对数生长期、稳定期、衰亡期4个生长阶段,生长曲线符合"S"型生长规律;采用胎牛血清(90%FBS+10%DMSO)和培养基(70%培养基+20%FBS+10%DMSO)两种冻存液冻存奶牛乳腺上皮细胞,细胞复苏培养后,胎牛血清冻存的细胞生长状态和增殖速度都明显优于培养基冻存的奶牛乳腺上皮细胞,从培养的第3天开始,2组细胞数量差异显著(P ＜;0.05)。本研究结果为建立体外培养体系提供了一定参考。

关键词: 奶牛乳腺上皮细胞, 体外培养, 生长曲线, 培养基, 接种浓度, 冻存

Abstract: The objective of this study was to optimize the culture conditions in vitro for bovine mammary epithelial cells(BMECs). The medium,inoculation concentration,and the freeze preservation of BMECs were studied in culture condition by cell proliferation,which was evaluated by cell count trypan blue dying. The results show that the growth of cell cultured in Media Dulbecco's modified Eagle's medium(DMEM) was slower than in DMEM/F12,followed by F12 and then RPMI1640. The inoculation concentration of the epithelial cells indicated that 1 × 10⁴ cells status in each well was the best mode. The cell growth cycle experienced latent phase,logarithmic growth phase,stable phase and decline growth stages,and the growth curve accorded with the " S" -type growth law. The serum-freezing solution(90% FBS + 10% dimethyl sulfoxide) and medium-freezing solution(70% medium + 20% FBS +10% dimethyl sulfoxide) were used to preserve the bovine mammary epithelial cells. The cell proliferation and morphology of serum-freezing state were better than the medium-freezing state. The number of serum-freezing state was significant higher than medium state(P ＜ 0. 05) from the third day. The present study supplied some date for the method of culturing BMECs in vitro.

Key words: Bovine mammary epithelial cells, Culture in vitro, Growth curve, Medium, Inoculation concentration, Frozen preservation

中图分类号:

S858.23

李喜艳, 王加启, 魏宏阳, 卜登攀, 胡菡, 周凌云. 奶牛乳腺上皮细胞体外培养条件优化的初步研究[J]. 华北农学报, 2010, 25(S1): 79-85. doi: 10.7668/hbnxb.2010.S1.019.

LI Xi-yan, WANG Jia-qi, WEI Hong-yang, BU Deng-pan, HU Han, ZHOU Ling-yun. The Optimal Culture Conditions in vitro for Bovine Mammary Epithelial Cells[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2010, 25(S1): 79-85. doi: 10.7668/hbnxb.2010.S1.019.

http://www.hbnxb.net/CN/Y2010/V25/IS1/79

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