High sensitivity liquid chromatography tandem mass spectrometry system

描述（由申请人提供）：对于本提案中描述的8个NIH支持的核心实验室和11名研究者，液相色谱/三重四极杆质谱（LC/MS-MS）在药物代谢物和肽的定量中起着重要作用。灵敏度和选择性通常存在问题，因为分析物的丰度固有地低，或者样品大小非常有限，并且分析物通常存在于高度复杂的生物基质中。拟议的仪器的主要用途将是在NIH支持的基础和临床项目中对常规小分子分析物进行定量。第二个用途将是使用我们正在开发的方法定量复杂生物基质（如组织）中的治疗和功能蛋白质。为此目的考虑了两种仪器：Thermo Finnigan Quantum Ultra AM MS/MS，具有精确的质量测量选项，以及Applied Biosystems API 5000 MS/MS。这两种仪器都代表了最新技术水平的重大进步，但存在明显的优势和妥协。我们对这些工具的评价和决定，以何种工具最好地满足我们的用户的需求进行了说明。拟议的仪器将被纳入现有的药物科学质谱设施，这是最全面的和大量使用的学术研究设施在西纽约的扩展地理区域。它目前拥有三个三重四极杆（LC-MS/MS）仪器，一个离子阱（LC/MSn）系统和一个新获得的具有多维LC功能的线性阱/四极杆系统（MDLC/LTQ）。该设施为从事基础，应用和临床生物医学研究的各种研究人员提供仪器访问。该设施的利用率很高，在过去3个日历年中，每台仪器平均为调查人员提供了至少1600小时的有用数据采集。然而，该设施缺乏具有所提出的仪器将带来的灵敏度和选择性的仪器，并且现有MS设备的灵敏度和选择性的限制在关键方面阻碍了所描述的项目，最常见的是当样品体积有限时，例如在儿科临床试验或小型啮齿动物疾病模型中。此外，目标蛋白的标记肽的定量是我们设施中不断增长的活动，并且通常最感兴趣的蛋白质是低丰度的蛋白质。没有其他区域性设施能够提供必要的扩展访问，以支持开发新的方法来量化低拷贝细胞蛋白质转录物或治疗上重要的蛋白质药物。鉴于我们的研究人员参与了基础和临床科学的研究，拟议的仪器将对危及生命的疾病的新疗法的发展以及我们对生化和生理过程的基本理解产生直接，重大的影响。相关性以高灵敏度和选择性定量细胞和组织中的药物、其代谢物和/或蛋白质生物标志物的能力是许多旨在改善疾病的治疗或诊断或了解生物过程的基本机制的研究项目的基本和共同需求。拟议的仪器将为这一地理区域带来必要的最先进仪器，从而推动许多寻求改善艾滋病和癌症等严重疾病治疗的项目。

DESCRIPTION (provided by applicant): For 8 core NIH-supported laboratories and 11 investigators described in this proposal, liquid chromatography/triple quadrupole mass spectrometry (LC/MS-MS) plays an important role in the quantification of drugs metabolites, and peptides. Sensitivity and selectivity often presents a problem because the analytes are either intrinsically low in abundance, or sample size is very limited, and the analytes generally are present in highly complex biological matrices. The major use of proposed instrument will be the quantification of conventional small-molecule analytes in both basic and clinical NIH-supported projects. A second use will be the quantification of therapeutic and functional proteins in complex biological matrices such as tissues, using methods we are developing. Two instruments were considered for this purpose: the Thermo Finnigan Quantum Ultra AM MS/MS, with the accurate mass measurement option, and the Applied Biosystems API5000 MS/MS. Both instruments represent a significant advance in the state-of-the-art, but present distinct advantages and compromises. Our evaluation of these instruments and the decision as to which instrument best serves the needs of our users is described. The proposed instrument will be integrated into the existing Pharmaceutical Sciences mass spectrometry facility, which is the most comprehensive and heavily utilized academic research facility in the extended geographic region of Western New York. It currently houses three triple quadrupole (LC-MS/MS) instruments, an ion trap (LC/MSn) system, and a newly-acquired linear trap/quadrupole system with multidimensional LC capabilities (MDLC/LTQ). This facility provides instrument access to a diverse population of researchers engaged in basic, applied, and clinical biomedical research. The facility is highly utilized, and has provided investigators with an average of at least 1600 hours of useful data acquisition per instrument over the last 3 calendar years. However, the facility lacks an instrument possessing the sensitivity and selectivity that the proposed instrument would bring, and limitations in the sensitivity and selectivity of the existing MS equipment hinders the described projects in key ways, most often when sample volumes are limited, e.g. in pediatric clinical trials or in small rodent disease models. Furthermore, quantification of marker peptides for target proteins of interest is a growing activity in our facility, and often the proteins of greatest interest are those of low abundance. No other regional facility exists to provide the extended access necessary to support the development of new methods for quantification of low-copy cellular protein transcripts or therapeutically important protein drugs. Given the involvement of our investigators in studies of both basic and clinical sciences, the proposed instrument would have direct, major impact on the development of new therapies for life-threatening diseases, as well as on our basic understanding of biochemical and physiological processes. Relevance The ability to quantify drugs, their metabolites, and/or protein biomarkers in cells and tissues, with high sensitivity and selectivity, is an essential and common need of many research projects that aim to improve the therapy or diagnosis of diseases, or to understand the basic mechanisms biological processes. The proposed instrument will bring to this geographic area the state-of-the art instrumentation necessary, thereby advancing many projects that seek to improve the therapy of serious diseases such as AIDS and cancer.