Wang Weijie, Guo Tong, Rudnick Paul A, Song Tao, Li Jie, Zhuang Zhengping, Zheng Wenxin, Devoe Don L, Lee Cheng S, Balgley Brian M
Calibrant Biosystems, 910 Clopper Road, Suite 220N, Gaithersburg, Maryland 20878, USA.
Anal Chem. 2007 Feb 1;79(3):1002-9. doi: 10.1021/ac061613i.
This work expands our tissue proteome capabilities from the analysis of soluble proteins in previous studies to the examination of membrane proteins within the pellets of enriched and selectively isolated tumor cells procured from microdissected tissue specimens. The pellets of targeted ovarian tumor cells are treated by two different membrane protein extraction methods, including the use of detergent and organic solvent. The detergent-based membrane protein preparation protocol not only extracts proteins effectively from cell pellets but also is compatible with subsequent proteome analysis using combined capillary isoelctric focusing/nano reversed-phase liquid chromatography separations coupled with nano electrospray ionization mass spectrometry. Among proteins identified from an amount of pellet equivalent to 20 000 cells, 773 proteins are predicted to contain one or more transmembrane domains, corresponding to 22% membrane proteome coverage within the SwissProt Human protein sequence entries.
这项工作将我们的组织蛋白质组分析能力从之前研究中对可溶性蛋白质的分析扩展到对从显微切割组织标本中富集并选择性分离的肿瘤细胞沉淀中的膜蛋白进行检测。靶向卵巢肿瘤细胞的沉淀通过两种不同的膜蛋白提取方法进行处理，包括使用去污剂和有机溶剂。基于去污剂的膜蛋白制备方案不仅能有效地从细胞沉淀中提取蛋白质，还与随后使用毛细管等电聚焦/纳米反相液相色谱联用纳米电喷雾电离质谱的蛋白质组分析兼容。在相当于20000个细胞数量的沉淀中鉴定出的蛋白质中，有773种蛋白质预计含有一个或多个跨膜结构域，相当于SwissProt人类蛋白质序列条目中膜蛋白质组覆盖率的22%。
