温度和Hoechst 33258浓度对人活精子标记效果的影响

目的研究孵育温度和荧光染料Hoechst 33258(H258)的浓度对活体标记人类精子效果的影响。方法随机选取14份精液标本,经洗涤处理后,在不同温度(0℃、室温和37℃)下分别用0(对照)、0.01、0.1 mg/mL的H258孵育1 h。记录精子前向运动百分率及H258着色率。以未予处理的精液标本作为空白对照组。结果在观察条件范围内,孵育温度与H258浓度对精子运动的影响有统计学意义。随着H258质量浓度的升高和孵育温度的降低,精子前向运动百分率下降(P<0.01)。精子着色率随孵育温度和H258质量浓度的升高而提高;当H258质量浓度为0.1 mg/mL时,不同孵育温度下的精子着色率比较差异有统计学意义(P<0.01)。结论 H258可用作精子活体荧光标记,37℃+0.1 mg/mL H258是本研究中最优的染色条件。但仍需要根据不同使用目的选择孵育条件。

Influence of temperature and Hoechst 33258 concentration on vital labeling of human sperm

Objective To investigate the influence of culture temperature and concentration of fluorescence dye Hoechst 33258(H258) on vital labeling of human sperm.Methods Fourteen semen samples were randomly selected,and were cultured with 0 mg/mL(control),0.01 mg/mL and 0.1 mg/mL H258 for 1 h under different temperatures(0 ℃,room temperature and 37 ℃) after washing.Progressive motility of human sperm was assessed,and H258 staining rates of the sperm were recorded.Semen samples without treatment were served as blank controls.Results Under the conditions of this research,H258 concentrations and culture temperatures had significant impact on sperm motility.The percents of progressive motility of sperm gradually decreased with the increase of H258 concentrations and decrease of culture temperatures(P<0.01).The staining rates of the sperm gradually increased with the culture temperatures and mass concentrations of H258,and there were significant differences among staining rates of the sperm cultured with different temperatures at the mass concentration of 0.1 mg/mL(P<0.01).Conclusion H258 can be used in vital labeling of human sperm.The optimal staining condition was 37 ℃+0.1 mg/mL H258 in this study.Culture condition should be determined by specific purpose.