Proteomics Core

蛋白质组学核心（核心B）为PPG的研究人员提供最先进的仪器、方法和蛋白质组学专业知识。蛋白质组学核心的一个独特功能是尖端的定量蛋白质组学分析技术的可用性，允许高灵敏度测量胃癌模型中的蛋白质组变化。Proteomics Core提供1）复杂样品中蛋白质的鉴定，2）体外和体内样品差异蛋白质表达的定量分析，以及3）蛋白质修饰的全局分析。 蛋白质组学核心人员提供实验设计咨询以及样品制备，质谱分析和主要数据分析。多台用于LC-MS/MS实验的高性能质谱仪和MALDI TOF-TOF仪器可用于蛋白质组学分析。对于复杂样品中的蛋白质鉴定，分析涉及多维LC-MS/MS，然后使用在高级计算研究和教育中心（ACCRE）的多处理器Linux集群上运行的Sequest软件进行自动数据库搜索。使用定制IDPicker或商业Scaffold软件评估、过滤和比较蛋白质鉴定。TagRecon的额外数据分析能够从MS/MS数据中识别出非预期的修饰或序列变体。Core还使用稳定同位素差异标记策略iTRAQ和SILAC分别对来自动物模型或细胞培养物的样本进行相对定量。使用多反应监测LC-MS/MS方法完成蛋白表达变化的验证。 项目1（Peek）将利用iTRAQ方法测量感染致癌H.幽门螺杆菌或突变体喂养铁充足和铁耗尽的饮食。项目2（Wilson）将利用相同的方法确定EGFR激活后胃上皮细胞蛋白质组的变化。此外，本项目将利用SILAC方法测量条件永生化细胞中EGFR激活后磷酸化蛋白质组的变化。项目3（封面）将采用iTRAQ方法测量H的差异。幽门螺杆菌输出菌株从沙鼠喂养高盐或正常饮食。在所有研究中将采用靶向MRM验证，以确认特定蛋白质组学变化

The Proteomics Core (Core B) provides state-of-the-art instrumentation, methodology, and expertise in proteomics to investigators in this PPG. A unique feature of the Proteomics Core is the availability of cutting¿ edge quantitative proteomics analysis technologies that allows high sensitivity measurements of proteome changes in models of gastric cancer. The Proteomics Core provides 1) identification of proteins in complex samples, 2) quantitative analyses of differential protein expression from in vitro and in vivo samples, and 3) global analyses of protein modifications. Proteomics Core staff provides consultation on experimentaldesign as well as hands on sample preparation, mass spectrometry analysis, and primary data analysis. Multiple high performance mass spectrometers for LC-MS/MS experiments and MALDI TOF-TOF instruments are available for proteomic analyses. For protein identification from complex samples, analysis involves multidimensional LC-MS/MS followed by automated database searching with Sequest software running on a multiprocessor Linux cluster in the Advanced Computing Center for Research and Education (ACCRE). Protein identifications are evaluated, filtered, and compared using custom lDPicker or commercial Scaffold software. Additional data analysis by TagRecon enables identification of unanticipated modifications or sequence variants from MS/MS data. The Core also provides relative quantitation using stable isotope differential labeling strategies, iTRAQ and SILAC, for samples derived from animal models or from cell culture, respectively. Validation of protein expression changes is accomplished using multiple reaction monitoring LC-MS/MS methods. Project 1 (Peek) will utilize the iTRAQ method to measure proteomic changes in gastric epithelial cells from gerbils infected with carcinogenic H. pylori or mutants fed iron-replete and iron-deplete diets. Project 2 (Wilson) will utilize the same approach to determine changes in gastric epithelial cell proteomes upon EGFR activation. In addition, this Project will utilize the SILAC method to measure changes in the phosphoproteome upon EGFR activation in conditionally immortalized cells. Project 3 (Cover) will employ the iTRAQ method to measure differences in H. pylori output strains from gerbils fed high salt or regular diets. Targeted MRM validation will be employed in all studies to confirm specific proteomic changes