Integrated Instrumentation for Proteomics

描述（由申请人提供）：本赠款的目的是购买 一个集成的仪器系统，包括高分辨率和 高通量双向电泳和专用质谱 光谱分析相关的蛋白质表达模式（蛋白质组学） 人类疾病所要求的仪器将提供最先进的 为NIH资助的大型生物医学研究提供蛋白质组学分析能力 在伯明翰的亚拉巴马大学的社区。分析单 蛋白质的改变和修饰一直是大规模利用 光谱设施，主要是通过使用矩阵辅助 激光解吸电离飞行时间质谱仪（MALDI-TOF MS）， 还有Q-TOF在这个后基因组时代，一旦多肽凝胶“点” 切除、胰蛋白酶消化并通过MALDI-TOF分析， 序列数据使得能够通过以下方式快速鉴定未知多肽 各种软件。所要求的仪器将解决主要因素 限制蛋白质组学分析：（1）增强相关蛋白质的检测， 在2-D凝胶“展示”中的多肽，（2）允许更高分辨率的多肽， 显示给定样品，和（3）能够高通量处理凝胶 “斑点”使得该步骤不是速率限制。随机软件确保 各种仪器之间的无缝集成，并允许从 开始步骤，等电聚焦，到最后步骤，MALDI-TOF 基于自动肽质量指纹图谱的蛋白质鉴定。在 去年，首席研究员指导了一项初步的发展计划， 二维电泳与质谱联用的阶段 设施，利用微型凝胶和考马斯或银染色的微型凝胶， 第二维度。结论是，必须采用更大的格式 凝胶，荧光检测，机器人切除和凝胶内数字化， 凝胶“斑点”，以及点样到MALDI-TOF板上，以自动 MALDI-TOF质谱法要求的集成系统（BioRad-Micromass） 将为我们的蛋白质组学分析能力 以维持高质量的生物医学研究的水平， UAB的标志。

DESCRIPTION (provided by applicant): The objective of this grant is to purchase an integrated instrumentation system that includes high-resolution and high-throughput, two-dimensional electrophoresis and dedicated mass spectrometry to analyze patterns of protein expression (proteomics) relevant to human disease. The requested instrumentation will provide state-of-the-art proteomics analytical capabilities for a large NIH-funded biomedical research community at the University of Alabama at Birmingham. Analysis of single protein changes and modifications has been a major utilization of the mass spectrometry facility, principally through use of the matrix-assisted laser-desorption ionization-time-of-flight mass spectrometer (MALDI-TOF MS), and the Q-TOF. In this postgenome era, once a polypeptide gel "spot" is excised, trypsin-digested and analyzed by MALDI-TOF, the availability of genome sequence data enables the rapid identification of unknown polypeptides through various software. The requested instrumentation will address the major factors that limit proteomic analysis: (1) enhance the detection of a relevant polypeptide(s) in a 2-D gel "display," (2) allow greater resolution of a display for a given sample, and (3) enable high-throughput processing of gel "spots" such that this step is not rate limiting. Accompanying software ensure seamless integration among various instruments, and allow sample tracking from beginning step, isoelectric focusing, to the last step, MALDI-TOF identification of protein based on automated peptide mass fingerprinting. In the last year, the principal investigator directed an initial developmental phase of 2-D electrophoresis in conjunction with the mass spectrometry facility, utilizing mini gels and Coomassie-or silver stained mini gels for the second dimension. The conclusion is that it is essential to go to larger-format gels, with fluorescence detection, robotic excision and in-gel digestions of gel "spots," as well as spotting onto MALDI-TOF plates, ending with automated MALDI-TOF mass spectrometry The requested integrated system (BioRad-Micromass) will provide the instrumentation to take our proteomics analytical capabilities to the level that will sustain the high quality of biomedical research that is a hallmark of UAB.