生物体液差异蛋白质组学研究技术体系的建立

The establishment of a technique system for differential proteomics of body fluids

为探讨以2D-DIGE为核心的生物体液差异蛋白质组学技术体系,取2组不同生物学状态下的体液样本进行研究。每组包含一例人类多发性硬化样本和一例对照样本,共设3组平行实验。经冷丙酮沉淀法除盐提取蛋白并精确定量,分别用分CY5和CY2荧光染料按最小标记法对多发性硬化样本和对照组进行标记。另再取混合蛋白内用CY3标记。混合样品后分别在3块2D-gel上进行电泳,通过Typhoon 9400多功能荧光扫描仪及DeCyder 2-D差异分析软件进行DIGE分析。差异蛋白用MALDI-TOF/TOF进行鉴定,将所得结果录入Metarcore计算平台,进行蛋白质相关分析。结果表明,利用该方法研究不同生物学状态下的体液标本,结合网络图谱分析,可得到较多有意义的候选蛋白信息。

In order to establish a technique system for differential pmteomics of body fluids, cerebrespinal fluid samples in two different physiological states were studied, There was a Multile Sclerosis sample and a control for each group, which was done in triplicate. Samples were precipitated with ice-cold acetone and the protein concentration was accurately quantified. The control and abnormal groups were labeled with two different Cyanine Dyes （ Cy2 and Cy5） using the minimal labeling method while the internal standard group was labeled with Cy3. Then 2D-E was used and three gels were rim. Each gel contained a control group, an abnomml group and the internal standard group labeled with 3 different Cye Dyes. The information including differential express protein spots and their levels were obtained using a Typhoon 9400 scanner and DeCyder 2-D software. Protein spots were identified by MALDI-TOF/TOF. Metacore integrated software was used to analyze the interactions between proteins. The results show that a differential proteomics techniaue system is effective to discover valuable candidate proteins.