人胆汁蛋白质组双向荧光差异凝胶电泳图谱的建立与差异分析  

Comparative proteomics analysis of human bile using two.dimensional differential in-gel electrophorisis

作　　者：陈波[1] 刘小方[2] 邹声泉[1] 鲁艳军[3] 汪昕[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院普外科,武汉430030 [2]烟台毓璜顶医院肝胆外科 [3]华中科技大学同济医学院蛋白质组学研究中心

出　　处：《中华实验外科杂志》2008年第4期466-468,共3页Chinese Journal of Experimental Surgery

基　　金：国家高新技术研究发展计划（863）项目（2002AA241061）

摘　　要：目的建立人胆汁蛋A质组双向荧光差异凝胶电泳分析流程用于胆汁比较蛋A质组学研究。方法实验组与对照组样本各6例，分别取自于胆管癌及胆总管结石病例。样本经纯化处理及定量后，分别标记不同的荧光染料后进行双向电泳与差异分析。结果两组样本均获得高分辨力的双向电泳图谱；各标记蛋白质点的荧光强度与蛋A表达量呈线性关系；实验组与对照组间共筛选出55个差异表达蛋白，其表达量两组间相差1．5倍以上，t检验有统计学意义（P〈0．05）。结论建立了基于双向荧光差异凝胶电泳基础上的胆汁比较蛋白质组学的分析流程。Objective To establish a two-dimensional differential in-gel electrophorisis process for comparative proteomic analysis of human bile. Methods Bile fluid was obtained from 12 patients with obstructive jaundice （each 6 cases of eholangiocarcinoma and choledocholith）. The unfractionated bile fluid was performed sonication and ultracentrifugation to remove the cellular debris and Nucleic Acid as a crude separation. Commercial kits Cleanascite and microcon were adopted to remove the abundant lipid and salt. After quantify by BCA Kit, concentrated and purified samples were labeled with three different CyDyes including one internal standard, and separated with 2-D DIGE in triplicate experiments. Biological variations of the level of protein expression were analyzed with DeCyder software and evaluated for reproducibility and statistical significance. Results Common gel artifacts can be reduced by above mentioned samples treatment method and high resolution bihary 2-D DIGE maps can be obtained ; the fluorescence intensity of separated protein spots have a line relationship with the level of their protein expression; 55 matched protein spots with statistical variance of two groups were detected （t-test, P 〈 0.05）. Conclusion An ideal, sample preparation strategy and optimized 2-D DIGE analysis process for human bile were established.

关 键 词：蛋白质组 胆汁 双向荧光差异凝胶电泳 

分 类 号：R781.5[医药卫生—口腔医学]