Assessment of separation methods for extracellular vesicles from human and mouse brain tissues and human cerebrospinal fluids.
Assessment of separation methods for extracellular vesicles from human and mouse brain tissues and human cerebrospinal fluids.
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DOI：
10.1016/j.ymeth.2020.02.002
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发表时间：
2020-05-01
期刊：
影响因子：
--
通讯作者：
Ikezu T
中科院分区：
文献类型：
--
作者：
Muraoka S;Lin W;Chen M;Hersh SW;Emili A;Xia W;Ikezu T
细胞外囊泡（EV）是一个统一的术语，膜封闭的囊泡物种普遍分泌的几乎每一种细胞类型，并存在于所有的体液。它们携带脂质、代谢物、核酸和蛋白质的货物，用于它们从细胞中清除以及用于细胞与细胞的通信。由于分离方案的不发达，特别是来自中枢神经系统（包括动物和人脑组织以及脑脊液）的分离方案，以及分离的EV中蛋白质的低产量，EV的确切组成及其特定功能还没有很好地理解。为了了解它们的确切分子组成及其功能作用，开发可靠的EV分离方案是必要的。在这里，我们报告的方法，从人和小鼠的未固定的冷冻脑组织的蔗糖步骤梯度超离心法，和从人的脑脊液的亲和捕获法分离EV。通过纳米颗粒跟踪分析、透射电子显微镜和标记和无标记质谱法对分离的EV的形态学、生物物理学和蛋白质组学特性进行评估，以进行蛋白质谱分析，并对每次评估进行分步方案。
Extracellular vesicle (EV) is a unified terminology of membrane-enclosed vesicular species ubiquitously secreted by almost every cell type and present in all body fluids. They carry a cargo of lipids, metabolites, nucleic acids and proteins for their clearance from cells as well as for cell-to-cell communications. The exact composition of EVs and their specific functions are not well understood due to the underdevelopment of the separation protocols, especially those from the central nervous system including animal and human brain tissues as well as cerebrospinal fluids, and the low yield of proteins in the separated EVs. To understand their exact molecular composition and their functional roles, development of the reliable protocols for EV separation is necessary. Here we report the methods for EV separation from human and mouse unfixed frozen brain tissues by a sucrose step gradient ultracentrifugation method, and from human cerebrospinal fluids by an affinity capture method. The separated EVs were assessed for morphological, biophysical and proteomic properties of separated EVs by nanoparticle tracking analysis, transmission electron microscopy, and labeled and label-free mass-spectroscopy for protein profiling with step-by-step protocols for each assessment.
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DOI：
10.1007/s00018-018-2773-4
发表时间：
2018-08
期刊：
Cellular and molecular life sciences : CMLS
影响因子：
--
作者：
Karimi N;Cvjetkovic A;Jang SC;Crescitelli R;Hosseinpour Feizi MA;Nieuwland R;Lötvall J;Lässer C
通讯作者：
Lässer C
影响因子：
16
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DOI：
10.1073/pnas.1521230113
发表时间：
2016-02-23
影响因子：
11.1
作者：
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影响因子：
4.8
作者：
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Simons, Mikael
DOI：
10.2215/cjn.12191018
发表时间：
2019-06-07
影响因子：
9.8
作者：
Bruschi, Maurizio;Granata, Simona;Zaza, Gianluigi
通讯作者：
Zaza, Gianluigi