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名研究人员，LC/MS-MS(LC/MS-MS)在药物代谢物和多肽的定量中发挥着重要作用。灵敏度和选择性往往是一个问题，因为分析物要么本身丰度低，要么样本量非常有限，而且分析物通常存在于高度复杂的生物基质中。建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传统小分子分析物进行量化。第二个用途将是使用我们正在开发的方法，对复杂生物基质(如组织)中的治疗性和功能性蛋白质进行量化。为此考虑了两种仪器：具有精确质量测量选项的Thermo Finnigan Quantum Ultra AM MS/MS和应用生物系统API5000 MS/MS。这两种仪器都代表着最先进的重大进步，但也存在明显的优势和折衷。描述了我们对这些工具的评价以及关于哪种工具最能满足我们用户需求的决定。拟议的仪器将被整合到现有的制药科学质谱学设施中，这是纽约西部广阔地理区域内最全面和最频繁使用的学术研究设施。它目前拥有三台三重四极(LC-MS/MS)仪器、一个离子陷阱(LC/MSN)系统和一个新获得的具有多维LC能力的线性陷阱/四极系统(MDLC/LTQ)。这一设施为从事基础、应用和临床生物医学研究的不同人群的研究人员提供了仪器。该设施的利用率很高，在过去3个历年中，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，该设施缺乏具有拟议仪器将带来的灵敏度和选择性的仪器，而且现有MS设备的灵敏度和选择性的限制在关键方面阻碍了所述项目，最常见的是在样本量有限的情况下，例如在儿科临床试验或小型啮齿动物疾病模型中。此外，在我们的设施中，对感兴趣的目标蛋白的标记多肽的量化是一个日益增长的活动，并且通常最感兴趣的蛋白质是那些低丰度的蛋白质。不存在其他区域设施来提供必要的扩展准入，以支持开发新的方法来量化低拷贝细胞蛋白转录本或具有治疗意义的蛋白质药物。鉴于我们的研究人员参与了基础和临床科学的研究，拟议的仪器将对威胁生命的疾病的新疗法的开发以及我们对生化和生理过程的基本理解产生直接和重大的影响。相关性高灵敏度和选择性地对细胞和组织中的药物、其代谢物和/或蛋白质生物标记物进行量化的能力是许多旨在改进疾病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要。拟议的工具将为这一地理区域带来必要的最先进的工具，从而推动许多旨在改善艾滋病和癌症等严重疾病的治疗的项目。

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.