NanoUPLC-nano-ESI-MS/MS鉴定二维凝胶银染后蛋白质点的方法  

Identification of two-dimensional gel protein spots after silver staining by NanoUPLC-nano-ESI-MS/MS

作　　者：王鸿丽[1] 刘锋[1] 李卫华[1] 李萍[1] 杨松成[1] 何昆[1] 

机构地区：[1]国家生物医学分析中心,北京100850

出　　处：《军事医学》2011年第7期541-545,共5页Military Medical Sciences

摘　　要：目的提高银染色后二维凝胶电泳图谱中蛋白质斑点的鉴定率。方法采用欧洲分子生物学实验室(EMBL)银染色方法结合改进后的质谱兼容胶内酶切方法并应用超高效纳升液相色谱-电喷雾串联质谱(NanoUPLC-nano-ESI-MS/MS)对二维凝胶电泳上的蛋白质斑点进行分析。结果二维凝胶上50个蛋白质斑点中46个点得到鉴定,共鉴定出74个有意义的蛋白质,其中一个蛋白斑点鉴定出6个有意义的蛋白质。结论该方法获得的样品经质谱检测和网上数据库比对,蛋白鉴定率高达90%以上,与基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)和串联飞行时间质谱(MALDI-TOF/TOF-MS)方法相比,蛋白质的鉴定率明显提高,表明该方法适合鉴定复合蛋白质斑点,是鉴定银染后二维凝胶蛋白质的一种强有力的分析手段。Objective To improve the identification rate of protein spots on two-dimensional gel electrophoresis after silver staining.Methods and Results Samples were prepared using the technology of European Molecular Biology Laboratory（EMBL） silver staining with in-gel digestion.The protein spots on the 2D gel were directly analyzed by NanoUPLC-nano-ESI-MS/MS.Forty-six of the 50 protein spots on the 2D gel were identified.Seventy-four significant proteins were distinguished and 6 proteins were identified from one of the 46 protein spots.Conclusion Over 90% processed proteins are identified by our procedures as confirmed by database search.The identification rate is significantly increased compared with the MALDI-TOF-MS and MALDI-TOF/TOF-MS methods.It is a powerful analytical method for proteomics research by NanoUPLC-nano-ESI-MS/MS.

关 键 词：蛋白质组学 凝胶 电泳 双向 银染色法 胶内酶切 NanoUPLC-nano-ESI-MS/MS 

分 类 号：Q51-33[生物学—生物化学]