基于PEG6000富集精液来源外泌体的提取及鉴定

目的介绍PEG6000富集精液来源外泌体的提取和鉴定方法，为精液外泌体相关研究奠定基础。方法选自南方医科大学 南方医院检验中心6名正常志愿者精液，经8% PEG6000富集并多步离心结合超速离心方法获取提取物，然后采用透射电子显 微镜观察其形态，纳米颗粒跟踪分析仪分析其大小，蛋白印迹法检测其蛋白组成成分。结果提取物呈类圆形、椭圆形囊泡，质 膜完整，大小约为30~150 nm，内部含有低电子密度物质，均阳性表达CD63、ALIX、TSG101 蛋白，且不含精子细胞内质网 Calnexin蛋白。结论经过对提取物的形态、大小以及其蛋白成分分析证实所提取到的物质为精液来源的外泌体，为后续开展 精液外泌体临床及基础研究提供了方法学基础。

Extraction and identification of semen-derived exosomes using PEG6000

Objective To extract and identify semen-derived exosome using PEG6000. Methods Exosomes were extracted from semen specimens from 6 healthy volunteers with step-by-step centrifugations and ultracentrifugation prior to 8% PEG6000 enrichment. The extracted exosomes were characterized by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) and Western blotting. Results The pellets obtained were round or elliptic membrane vesicles 30 to 150 nm in diameter with intact double membranes and contained low electron density material. The pellets expressed CD63, ALIX and TSG101 molecules but not calnexin that was expressed in sperm cells. Conclusion The PEG6000-based method for extraction of exosomes from semen samples facilitates future studies of seminal exosomes.