低浓度NaOH对小鼠精子DNA完整性的影响

›› 2010, Vol. 30 ›› Issue (2): 82-86.

低浓度NaOH对小鼠精子DNA完整性的影响

1.华东理工大学，上海 200237；2.上海市计划生育科学研究所国家人口计生委计划生育药具重点实验室，上海 200032

收稿日期:2009-08-12 出版日期:2010-02-28 发布日期:2010-02-28
基金资助:
国家自然科学基金（30872765）和上海市科技发展基金（071409004）

Impact of Low Concentration of NaOH on DNA Integrity of Mouse Sperm Before ICSI

1.East China University of Science & Technology, Shanghai 200237, China;2. Shanghai Institute of Planned Parenthood Research Laboratory of Contraceptives and Devices Research,National Population and Family Planning Commol/Lision of China, Shanghai 200032, China

Received:2009-08-12 Online:2010-02-28 Published:2010-02-28

摘要/Abstract

摘要： 目的用精子DNA完整性评价NaOH溶液对小鼠精子进行头尾分离的最优条件.方法取2～3月龄雄鼠附睾尾精子，稀释到合适浓度(5×10⁶～10×10⁶/ml)，0～37℃下，加入不同浓度NaOH溶液孵育，显微镜下观察精子的头尾分离率，流式细胞仪检测精子的质膜完整性及DNA完整性.结果 NaOH对精子头尾分离的效果，与碱浓度、作用时间和作用温度呈正相关，作用温度越高，时间越长，头尾分离效果越好，但同时对精子DNA的损伤程度也就越大.本研究证实精子在10 mmol/t，NaOH溶液，0℃下孵育2 h，精子头尾分离的效率可达55%，而精子DNA损伤与空白对照相比，则无显著性差异.结论在本研究条件下低浓度NaOH处理小鼠精子，方法简单易行，可有效去除小鼠尾巴，而又不使其DNA受损.

关键词: ICSI, DNA损伤, 流式细胞仪, NaOH

Abstract: Objective To optimize the method of separation of mouse sperm head and tail by treating with different concentration of NaOH so as to improve the efficiency of ICSI. Methods The sperm were collected from the epididymis of male mice at the age of 2-3 months and diluted to an appropriate density. The sperm head and tail separation rate was determined under the microscope after incubating with different concentration of NaOH at the range of 0-37°C. Flow cytometry was utilized to evaluate both of membrane and DNA integrity of the sperm cells. Results The sperm head and tail separation rate was positively correlated with NaOH concentration, incubation time and temperature. However，DNA integrity of the sperm cells was negatively correlated to NaOH concentration and incubation temperature and time. The lower NaOH concentration and temperature as well as the shorter incubation time led to less serious DNA damage. The results indicated that the condition of 10 mmol/L NaOH for 2 h at 0°C was the optimal treatment to achieve 55% of sperm head and tail separation rate without injury of their DNA. Conclusion The low concentration NaOH treatment of sperm with the optimized condition in our study is a simple and convenient methodwhich can effectively separated the tail from sperm while remain the high DNA integrity.

Key words: ICSI, DNA damage, Flow cytometry, NaOH

宋丽伟1，史庭燕2，林菊芳1，李翀2，袁瑶2，欧伶1，施惠娟2. 低浓度NaOH对小鼠精子DNA完整性的影响[J]. , 2010, 30(2): 82-86.

SONG Li-wei1，SHI Ting-yan2，LIN Ju-fang1，LI Chong2，YUAN Yao2，OU Ling1，SHI Hui-juan2. Impact of Low Concentration of NaOH on DNA Integrity of Mouse Sperm Before ICSI[J]. , 2010, 30(2): 82-86.

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