Hybrid Triple Quadrupole Mass Spectrometer for Quantitative Mass Spectrometric Ap

用于定量质谱应用的混合三重四极杆质谱仪

• 批准号: 7794609
• 负责人: HARRY ISCHIROPOULOS
• 金额: $ 50万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7794609
• 关键词: Animals; Antibodies; Biochemical; Biological Markers; Biological Models; Biological Process; Cell model; Childhood; Collaborations; Development; Diagnosis; Disease; Enzyme-Linked Immunosorbent Assay; Funding; Generations; Genomics; Goals; Human; Hybrids; Ions; Knowledge; Liquid Chromatography; Methodology; Molecular; Pathway interactions; Pediatric Hospitals; Peptides; Philadelphia; Physiological; Physiological Processes; Post-Translational Protein Processing; Proteins; Proteolytic Processing; Quantitative Evaluations; Research; Research Institute; Research Personnel; Research Project Grants; Sampling; Sensitivity and Specificity; Stable Isotope Labeling; Staging; Techniques; Translational Research; United States National Institutes of Health; Validation; Western Blotting; base; insight; instrument; instrumentation; interest; mass spectrometer; multiple reaction monitoring; novel; outcome forecast; programs; public health relevance; research study; tandem mass spectrometry

DESCRIPTION (provided by applicant): The purpose of this application is to acquire a hybrid triple quadrupole-ion trap mass spectrometer, specifically the QTRAP 5500 from Applied Biosystems, with unique capabilities to support actively ongoing studies that aim to uncover critical physiological pathways and causes of pediatric diseases. This instrument will be used exclusively for quantitative mass spectrometric applications by 9 NIH funded investigators at the Joseph Stokes, Jr. Research Institute of the Children's Hospital of Philadelphia. Ongoing projects initiated by these investigators have employed genomic discovery platforms, biochemical, molecular, different mass spectrometric and orthogonal experiments to discover proteins of interest, posttranslational modifications, proteolytic processing and networks of proteins that execute biological functions. Therefore all these projects are well advanced past the stages of discovery and specific and practical protein and peptide targets have been identified. The immediate challenge for these projects is to move beyond qualitative descriptions to explore these discoveries in depth by quantifying changes in proteins and peptides. This goal cannot be accomplished with typical antibody based western blot, ELISA or even with our current linear ion trap mass spectrometers since these approaches are often neither feasible nor practical due to lack of sensitivity, specificity, and/or throughput. Targeted peptide quantification is becoming the clear choice as a highly selective and sensitive methodological approach for quantitative evaluation of dynamic changes of proteins, protein networks, posttranslational modifications, validation and utilization of biological markers. Towards this goal the PI and co-Investigator in collaboration with the users of the proposed instrument are actively developing methodologies that employ stable isotope labeled peptides and multiple reaction monitoring (MRM) workflows for the quantification of proteins in human samples as well as in animal and cellular model systems. Multiple reaction monitoring is emerging as the standard technique for quantitative liquid chromatography tandem mass spectrometry (LC/MS/MS) experiments; however generation and application of these MRM workflows with our current ion trap platforms is limited. Therefore the installation and use of the proposed mass spectrometer is vital for enabling the development and implementation of methodologies for targeted peptide quantification to generate sizeable new knowledge and insights. Overall, the proposed projects although biologically diverse are united by the need to develop, validate and implement quantitative hypothesis-driven mass spectrometric approaches. Thus, while each research project stands alone in scientific value and integrity the added value of this combined proposal is substantial. PUBLIC HEALTH RELEVANCE: The instrument will have a major and immediate impact on the research programs of 9 NIH-funded investigators that are engaged in both basic and translational research aimed to uncover fundamental physiological processes and mechanisms of disease at the Children's Hospital of Philadelphia. The requested instrumentation will facilitate the quantification of potentially novel biological markers for the prognosis and diagnosis of pediatric diseases.

描述（由申请人提供）：本申请的目的是获得一个混合式三重四极离子阱质谱仪，特别是应用生物系统公司的QTRAP 5500，具有独特的功能，以支持正在进行的旨在揭示儿科疾病的关键生理途径和原因的研究。该仪器将专门用于美国国立卫生研究院资助的费城儿童医院Joseph Stokes， Jr.研究所的9名研究人员的定量质谱应用。由这些研究人员发起的正在进行的项目利用基因组发现平台、生化、分子、不同质谱和正交实验来发现感兴趣的蛋白质、翻译后修饰、蛋白质水解加工和执行生物功能的蛋白质网络。因此，所有这些项目都已经过了发现阶段，并且已经确定了具体和实用的蛋白质和肽靶点。这些项目面临的直接挑战是超越定性描述，通过量化蛋白质和肽的变化来深入探索这些发现。这一目标无法通过典型的基于抗体的western blot、ELISA甚至我们目前的线性离子阱质谱仪来实现，因为这些方法由于缺乏灵敏度、特异性和/或吞吐量，通常既不可行也不实用。靶向肽定量作为一种高度选择性和敏感性的方法，正在成为定量评估蛋白质动态变化、蛋白质网络、翻译后修饰、生物标记物验证和利用的明确选择。为了实现这一目标，PI和联合研究者与拟议仪器的用户合作，正在积极开发使用稳定同位素标记肽和多反应监测（MRM）工作流程的方法，用于人类样品以及动物和细胞模型系统中的蛋白质定量。多反应监测正在成为定量液相色谱串联质谱（LC/MS/MS）实验的标准技术；然而，这些MRM工作流程的生成和应用与我们目前的离子阱平台是有限的。因此，安装和使用所提出的质谱仪对于开发和实施靶向肽定量方法至关重要，从而产生相当大的新知识和见解。总的来说，虽然生物多样性的提议项目是统一的，需要发展，验证和实施定量假设驱动的质谱方法。因此，虽然每个研究项目在科学价值和完整性上都是独立的，但这个综合建议的附加价值是巨大的。