外泌体的不同染色方法效果比较  

Comparison of different staining methods for exosomes

作　　者：朱艳 张雷 周晨明 闫静 孟丽 ZHU Yan;ZHANG Lei;ZHOU Chen-ming;YAN Jing;MENG Li(Electron Microscopy Center,Hebei Medical University,Shijiazhuang 050017,China;Department of Histology and Embryology,the School of Basic Medicine Sciences,Hebei Medical University, Shijiazhuang 050017,China)

机构地区：[1]河北医科大学电镜实验中心,河北石家庄050017 [2]河北医科大学基础医学院组织胚胎学教研室,河北石家庄050017

出　　处：《河北医科大学学报》2019年第9期1068-1071,共4页Journal of Hebei Medical University

基　　金：河北医科大学科研发展基金项目(KYFZ045)

摘　　要：目的通过用四氧化锇、磷钨酸、钼酸铵和醋酸双氧铀4种染液分别对外泌体负染色,并对染色效果进行观察和对比分析,以期选出对外泌体染色效果最好的负染液。方法用差速离心法提取外泌体后,采用漂浮法分别用四氧化锇、磷钨酸、钼酸铵、醋酸双氧铀4种不同染液对外泌体进行负染色,透射电镜观察,从外泌体的杯托状或圆盘状结构清晰度、膜边缘锐利与否、染色背景干净程度、反差效果等方面对比4种染液的效果。结果四氧化锇的染色背景出现大量泡状结构,外泌体结构不明显。磷钨酸对外泌体的染色效果不如醋酸双氧铀和钼酸铵,外泌体的胞膜边缘不清晰,染色背景有杂质。钼酸铵对外泌体的染色效果也较好,其囊泡的膜结构较清楚,外泌体的杯托状结构可较清楚见到,但是反差较弱。醋酸双氧铀染色下,外泌体典型的杯托状或圆盘状结构清晰可见,膜边缘锐利,染色背景干净,反差效果佳。结论醋酸双氧铀对外泌体的染色效果最好,是最合适的负染染液。Objective To compare and analyze the staining effect and select the best negative dyeing solution for exosomes after negatively staining the exosomes with four kinds of dyes,such as osmium tetroxide,phosphotungstic acid,ammonium molybdate and uranyl acetate.Methods After exosomes were extracted by differential centrifugation,the exosomes were negatively stained with four different dyeing solutions of osmium tetroxide,phosphotungstic acid,ammonium molybdate and uranyl acetate by means of floating method,and observed by transmission electron microscopy.The clarity of the cup-shaped or disc-shaped structure of the exosomes,the sharpness of the film edge,the cleanness of the staining background,and the contrast effect werecompared according the effects of the four dyeing solutions.Results Osmium tetroxide staining background appeared a large number of vesicles,exosome structurewas not obvious.The dyeing effect of phosphotungstic acid on exosomes was not as good as that of uranium dioxide acetate and ammonium molybdate.The cell membrane edge of exosomes was not clear and the dyeing background was impurities.Ammonium molybdate had a better staining effect on exosomes.The membrane structure of the vesicles was clear,and the cup-supporting structure of the exosomes can be clearly seen,but the contrast was weak.Under uranium acetate hydrogen peroxide staining,the typical cup-shaped or discoid structure of exosome were clearly visible,the edge of membrane was sharp,the dyeing background was clean,and the contrast effect were good.Conclusion The uranyl acetate has the best staining effect on exosomes,and it is the most suitable negative dyeing solution.

关 键 词：外泌体 负染色 透射电镜 

分 类 号：R331[医药卫生—人体生理学]