水牛犊牛血浆外泌体的分离与鉴定

研究旨在从水牛犊牛血浆中分离外泌体，并对分离的外泌体进行分子生物学特征分析和鉴定。采用差速离心法分离3头水牛犊牛的血浆外泌体，并在透射电子显微镜下观察其形态，使用纳米粒度及Zeta电位仪对提取的外泌体进行粒径大小分析，应用Western blotting方法检测外泌体标记蛋白钙联蛋白（Calnexin）、肿瘤易感基因101（TSG101）、CD9、CD81在3头水牛犊牛血浆外泌体中的表达情况。结果显示，从水牛犊牛血浆中分离的外泌体形态多为圆形和椭圆形，直径在30~150 nm；Western blotting检测结果表明，在水牛血浆外泌体表面，特异性蛋白Calnexin、TSG101和CD81阳性表达，CD9不表达。本研究从形态学和分子生物学特征等方面证实研究获得的提取物为外泌体，由此说明，差速离心法可以成功分离提取水牛犊牛血浆中的外泌体，为水牛外泌体的后续研究提供了重要技术基础和参考。

Isolation and Identification of Plasma Exosomes in Buffalo Calves

The aim of this study was to separate exosomes from the plasma of buffalo calves, analysis and identify molecular biological characteristics of the isolated exosomes. The differential centrifugation was used to separate exosomes from plasma of three buffalo calves, and the morphology of extracted exosomes was observed under a transmission electron microscope. The particle size of the extracted exosomes was analyzed using nanoparticle size and Zeta potentiometer, and the expression of exosome marked proteins, including Calnexin, tumor susceptibility gene 101 (TSG101), CD9 and CD81, were detected by Western blotting method. The results showed that plasma exosomes of buffalo calves were mostly round and elliptical, with a diameter between 30 to 150 nm. The Western blotting results showed that the specific proteins Calnexin, TSG101 and CD81 were positive expressed in the extracted plasma exosomes, but protein CD9 was negative. Based on morphology and molecular characteristics, it was confirmed that the obtained extract were exosomes, indicating that the differential centrifugation could be used to successfully isolate plasma exosomes of buffalo calves. The results provided an important technical basis and reference for further research on buffalo exosome.