从非洲爪蟾中进行蛋白质复合物的免疫分离。
Immunoisolation of protein complexes from Xenopus.
作者信息
Conlon Frank L, Miteva Yana, Kaltenbrun Erin, Waldron Lauren, Greco Todd M, Cristea Ileana M
机构信息
Department of Genetics, University of North Carolina, Chapel Hill, NC, USA.
出版信息
Methods Mol Biol. 2012;917:369-90. doi: 10.1007/978-1-61779-992-1_21.
Abstract
The immunoaffinity isolation of protein complexes is an essential technique for the purification and -concentration of protein complexes from cells and tissues. In this chapter we present the methodologies for the purification of proteins and protein complexes from Xenopus laevis and Xenopus tropicalis. Specific to this protocol are the techniques for the cryolysis of Xenopus cells and tissues, a procedure that limits contamination from yolk proteins while preserving endogenous protein complexes, the methodologies for immunoaffinity purification of proteins using magnetic beads, and the protocols for western blot analysis. In addition, the procedures in this chapter can be extended to use with proteomic analysis of protein complexes as presented in the following chapter.
摘要
蛋白质复合物的免疫亲和分离是从细胞和组织中纯化和浓缩蛋白质复合物的一项重要技术。在本章中，我们介绍了从非洲爪蟾和热带爪蟾中纯化蛋白质和蛋白质复合物的方法。本方案特有的技术包括非洲爪蟾细胞和组织的冷冻溶解技术，该技术在保留内源性蛋白质复合物的同时限制了卵黄蛋白的污染；使用磁珠进行蛋白质免疫亲和纯化的方法；以及蛋白质印迹分析的方案。此外，本章中的程序可以扩展用于下一章介绍的蛋白质复合物的蛋白质组学分析。
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