2D-DIGE System for Proteomics

描述(申请人提供)：为蛋白质组学的二维差示凝胶电泳(2D-DGE)样品制备系统申请资金，该系统包括：分离、荧光成像、计算机化数据分析、机器人挑选斑点、蛋白质分解和传送到适合于质谱学分析的目标的设备。由于使用了不同发射光谱的荧光染料来标记样品，该系统提供了对差异蛋白质表达的半定量评估，并提高了样品对之间的重复性。因此，增加了统计可靠性，并更有效地利用了宝贵的样本。这一功能对于使用细胞分选机制备的样本或激光捕获的细胞群体的蛋白质组学分析更加关键，如一些项目中所述。该成像设备还允许将蛋白质模式与使用荧光染料检测到的潜在磷酸化蛋白质和免疫活性蛋白质进行比较。参与这项提议的研究人员将蛋白质组学应用于信号转导、细胞通讯、肥胖、糖尿病和癌症研究以及生物发现。这一仪器系统还将有助于测试本提案中未包括的新项目的可行性。这些仪器将由经验丰富的蛋白质化学家和质谱学家在现有的共享蛋白质核心设施内操作和监督，并有生产力和协作互动的记录。

DESCRIPTION (provided by applicant): Funds are requested for a two dimensional difference gel electrophoresis (2D-DIGE) sample preparation system for proteomics, consisting of devices for: separation, fluorescence imaging, computerized data analysis, robotic spot-picking, proteolytic digestion, and delivery to targets suitable for mass spectrometry analysis. This system provides a semi-quantitative assessment of differential protein expression with enhanced reproducibility between pairs of samples, since fluorescent dyes of different emission spectra are utilized to label the samples. As a result, statistical reliability is increased, and precious samples are more efficiently used. This feature is even more critical for proteomics analysis of samples prepared using the cell sorter or of laser captured cell populations, as described in some projects. The imaging device also permits comparison of protein patterns with potentially phosphorylated proteins detected using a fluorescent dye and with immunoreactive proteins. Investigators participating in this proposal have proteomics applications in signal transduction, cell communications, obesity, diabetes and cancer research and biological discovery. This instrument system will also be useful for testing feasibility of new projects not included in this proposal. The instruments will be operated and supervised by experienced protein chemists and mass spectrometrists within an existing shared protein core facility with a record of productivity and collaborative interactions.