Exosomes from Cell Culture-Conditioned Medium: Isolation by Ultracentrifugation and Characterization
Exosomes from Cell Culture-Conditioned Medium: Isolation by Ultracentrifugation and Characterization
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DOI：
10.1007/978-1-4939-9133-4_19
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发表时间：
2019-01-01
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影响因子：
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通讯作者：
Purushothaman, Anurag
中科院分区：
文献类型：
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作者：
Purushothaman, Anurag
外泌体是由大多数细胞类型分泌的内体来源的小囊泡。最近的研究已经确定外泌体是细胞间通讯的重要介质，也是许多临床应用的重要来源材料，包括微创疾病诊断。已经通过许多不同的方法从体外细胞培养上清液中纯化外泌体;然而，最简单和可靠的方法涉及通过超离心纯化。本章描述了一个详细的协议，用于分离外泌体从细胞培养条件培养基中使用ultracentbridgation和他们的表征的基础上的大小，数量和蛋白质表达的几个互补的方法，如透射电子显微镜，纳米粒子跟踪分析，蛋白质印迹和流式细胞术。
Exosomes are small vesicles of endosomal origin secreted by most cell types. Recent studies have identified exosomes as important mediators of intercellular communication and as important source materials for many clinical applications, including minimal invasive disease diagnosis. Exosomes have been purified from in vitro cell culture supernatants by many different methods; however the most simple and reliable method involves purification by ultracentrifugation. This chapter describes a detailed protocol for isolating exosomes from cell culture-conditioned medium using ultracentrifugation and their characterization based upon size, number, and protein expression by several complementary methods such as transmission electron microscopy, nanoparticle tracking analysis, western blotting, and flow cytometry.