二维阵列色谱分离系统及其在血浆蛋白质组中的应用  

Array-Based Two Dimensional Liquid Chromatography System for Proteomic Analysis of Human Plasma

作　　者：黄志[1] 邓楠[2] 晏国全[1] 高明霞[1] 梁振[2] 张丽华[2] 张祥民[1] 张玉奎[2] 

机构地区：[1]复旦大学化学系,生物医学研究院,上海200433 [2]中国科学院大连化学物理研究所分离分析化学重点实验室,大连116023

出　　处：《分析化学》2015年第10期1472-1478,共7页Chinese Journal of Analytical Chemistry

基　　金：国家高技术研究发展计划(No.2012AA020202);科技部基础研究计划项目(No.2012CB910604);国家自然科学基金资助项目(Nos.21027002;21175026;21275034)资助~~

摘　　要：本研究发展了在线二维阵列式液相色谱系统,对血浆蛋白质进行分离分析。在线二维阵列式液相色谱系统进行血浆样品的蛋白质水平分离,整个分离过程仅需4 h,并能进行高丰度蛋白质的快速定位。相比于高丰度蛋白质馏分的直接鉴定,经蛋白质均衡器材料富集后,鉴定得到的低丰度蛋白质数量提高了10倍,低丰度蛋白质的损失大幅度减少。本研究将二维阵列色谱分离系统对血浆样品进行完整的分离分析,共鉴定到了1474种蛋白质,蛋白质浓度动态范围达到了7个数量级。高丰度蛋白质馏分中共鉴定到252种蛋白质,其中有61种为高丰度蛋白质。结果表明,在线二维阵列式液相色谱系统实现了血浆样品中高丰度蛋白质的快速去除和高丰度蛋白质馏分中低丰度蛋白质的富集,显著提高了血浆蛋白质的鉴定能力,在其它复杂样品的分离分析中具有一定的应用前景。Human plasma proteome has the characteristics of complexity in component and large dynamic range of protein concentrations. Herein, we developed an array-based online two dimensional liquid chromatography system combined with protein equalizer technology for the large-scale depletion of high abundance proteins and enrichment of low abundance proteins in human plasma. The array-based online two dimensional liquid chromatography system could separate the plasma at the intact protein level with the merits of good reproducibility and high throughput. The total separation time was only 4 h and the fast location of high abundance proteins was also achieved. After the high abundance protein fractions was treated by ampholine@PM polymer microsphere,the number of the identified low abundance proteins increased tenfold,which significantly decreased the loss of low abundance proteins in high abundance protein fractions. The techniques were then applied to perform the proteomic analysis of human plasma sample. The total number of identified proteins was 1474 and the dynamic range of protein concentration was 7. Meanwhile,252 proteins were identified in high abundance protein fractions,among which 61 proteins belong to high abundance proteins.These results demonstrated that an array-based online two dimensional liquid chromatography system combined with protein equalizer technology could efficiently achieve the large-scale depletion of high abundance proteins and enrichment of low abundance proteins in human plasma,with a remarkable improvement in protein identification and a great prospect in the proteomic research of other complex samples.

关 键 词：多维色谱 阵列色谱 高丰度蛋白质 蛋白质均衡器 血浆 

分 类 号：R446.1[医药卫生—诊断学] O652.63[医药卫生—临床医学]