A Universal Multiplex Assay System for High-Throughput Clinical Applications

描述（由申请人提供） 摘要：基因组学、蛋白质组学和代谢组学的最新进展使得“组学”技术成为用于鉴定人类疾病的候选生物标志物的强大的基于发现的工具;然而，迄今为止，通过利用这些技术来建立用于临床实践的新生物标志物还没有成功。主要瓶颈在于缺乏有效的高通量验证工具。为了克服这一瓶颈，我建议开发一种新的“无试剂”质谱为基础的通用多重检测系统，将提供高通量的定量测量数百个低丰度蛋白质和代谢物分析物，独立于抗体为基础的试剂。该技术平台的目标是通过提供分析物信号强度的>1000倍增强（足以检测低丰度物质）和分辨率的>5000倍改善（用于极高特异性检测）来实现对当前MS平台的深刻进步。这些进展将通过开发和集成1）具有纳米电喷雾阵列的新型亚环境压力电离源，2）先进的离子漏斗接口，3）新型多级气相离子迁移技术来实现（与场不对称离子迁移谱法耦合的微分迁移率分析器）用于分离和选择感兴趣的分析物，（4）一种新的三级五四极杆（QqQqQ）质谱计，用于进一步分离和检测离子。优化的平台将具有每天>100个样品的潜在分析通量，广泛分析低丰度候选生物标志物而无需富集的灵敏度，以及同时监测多达1000种分析物的多路复用能力。灵敏度或检测动态范围至少3-4个数量级的增强（即，达到与当前ELISA相当或上级的水平），这是相对于当前分析平台的显著进步。这种新颖的分析系统是一种颠覆性技术，将通过仪器商业化彻底改变生物医学研究、当前医疗实践和未来医疗保健的许多领域以及一般生物医学研究领域。 公共卫生相关性：本项目的目标是开发一种新型的基于质谱的通用多重检测系统，用于高通量临床发现和验证人类疾病的候选生物标志物。该开发将通过加速生物标志物的发现和临床实践的实施，对当前的生物医学研究产生重大的临床和转化影响。使用新的测定系统可以有效地筛选用于各种人类疾病的大量候选生物标志物以进行验证，从而发现用于改善疾病诊断、预后和治疗的标志物。

DESCRIPTION (Provided by the applicant) Abstract: Recent advances in genomics, proteomics, and metabolomics make the "omics" technologies powerful discovery-based tools for identifying candidate biomarkers for human diseases; however, it has not been successful so far to establish new biomarkers for clinical practice by utilizing these technologies. The main bottleneck lies in the lack of effective tools for high-throughput validation. To overcome this bottleneck I propose to develop a novel "reagent-free" mass spectrometry-based universal multiplex assay system that will provide high throughput quantitative measurements for hundreds of low-abundance protein and metabolite analytes, independent of antibody-based reagents. The goal for this technology platform is to achieve a profound advance over current MS-platforms by providing >1000-fold enhancement in analyte signal intensities, sufficient for detecting low-abundance species, and >5000-fold improvement in resolving power for extremely high specificity detection. These advances will be achieved by developing and integrating 1) a novel subambient pressure ionization source with nanoelectrospray array, 2) advanced ion-funnel interfaces, 3) novel multi-stage gas-phase ion mobility technology (differential mobility analyzer coupled to field asymmetric ion mobility spectrometry) for separating and selecting analytes of interest, and (4) a new triple-stage pentaquardrupole (QqQqQ) mass spectrometer for further isolating as well as detecting ions. The optimized platform will have a potential analytical throughput of >100 samples per day, sensitivity for broadly analyzing low-abundance candidate biomarkers without enrichment, and the multiplexing power to monitor up to 1000 analytes simultaneously. At least 3-4 orders of magnitude enhancement in sensitivity or detection dynamic range (i.e., to a level comparable or superior to current ELISA) is anticipated, a significant advance over current assay platforms. Such a novel assay system is a disruptive technology that will revolutionize many areas of biomedical research, current medical practice, and the future of health care, as well as the biomedical research field in general through instrument commercialization. Public Health Relevance: The objective of this project is to develop a novel mass spectrometry-based universal multiplex assay system for high throughput clinical discovery and validations of candidate biomarkers of human diseases. The development will have major clinical and translational impact on current biomedical research by accelerating biomarker discovery and implementation in clinical practices. Large number of candidate biomarkers for various human diseases can be effectively screened for validation using the new assay system to discover markers for improved disease diagnosis, prognosis, and treatment.