Tanca Alessandro, Uzzau Sergio, Addis Maria Filippa
Porto Conte Ricerche, Loc, S.P. 55 Porto Conte/Capo Caccia Km 8.400, Tramariglio, Alghero (SS), 07041, Italy.
Methods Mol Biol. 2015;1295:117-34. doi: 10.1007/978-1-4939-2550-6_11.
Archival formalin-fixed, paraffin-embedded (FFPE) tissue repositories and their associated clinical information can represent a valuable resource for tissue proteomics. In order to make these tissues available for protein biomarker discovery and validation studies, dedicated sample preparation procedures overcoming the intermolecular cross-links introduced by formalin need to be implemented. This chapter describes a full-length protein extraction protocol optimized for downstream gel-based proteomics applications. Using the procedures detailed here, SDS-PAGE, western immunoblotting, GeLC-MS/MS, 2D-PAGE, and 2D-DIGE can be carried out on FFPE tissues. Technical tips, critical aspects, and drawbacks of the method are presented and discussed.
存档的福尔马林固定石蜡包埋（FFPE）组织库及其相关临床信息可能是组织蛋白质组学的宝贵资源。为了使这些组织可用于蛋白质生物标志物的发现和验证研究，需要实施专门的样品制备程序，以克服福尔马林引入的分子间交联。本章描述了一种针对基于凝胶的蛋白质组学下游应用进行优化的全长蛋白质提取方案。使用此处详述的程序，可以对FFPE组织进行SDS-PAGE、western免疫印迹、GeLC-MS/MS、二维聚丙烯酰胺凝胶电泳（2D-PAGE）和二维差异凝胶电泳（2D-DIGE）。本文介绍并讨论了该方法的技术要点、关键方面和缺点。
