Development of Next Generation Mass Spectrometric Instrumentation for Proteomics

项目概要/摘要 目前，大多数用于蛋白质组学的质谱（MS）分析是连续进行的。 模式，其中样品中的各种物质被一个接一个地选择和询问。作为 顺序模式MS是依次检查每个物种所需的有限时间的结果， 在灵敏度、速度和分析所有离子的能力方面受到不可避免的限制， 当离子束的成分复杂且快速变化时。这些限制 保留了大量的生物学和生物医学，包括，例如，深层单细胞蛋白质组 分析，目前的MS技术无法达到。在本提案中，我们重申， 通过并行执行MS（通过 类似于下一代DNA测序），从而克服了 目前的商业质谱仪主要以顺序模式操作。 在这里，我们建议开发新的MS仪器，以大规模并行方式执行MS 方式，考虑到两个主要目标： 目的1.提高蛋白质组分析的灵敏度、速度和深度， 最终超过1000倍目前的顺序MS方法可以比作采样 尼亚加拉福尔斯，其中大部分样品被浪费。我们提出的平行 MS技术旨在消除这种巨大的浪费。 目标2.在MS分析之前，在真实的时间中过滤噪声以消除不需要的离子背景， 从而使感兴趣的样品离子的MS利用率最大化， 噪声比，以及提供增加的动态范围来测量非常低的丰度 在高丰度组分存在下的组分。 这些目标的成功实现将使深，全面，高吞吐量 蛋白质组分析的情况下，样品的可用性是有限的， 由于它们低丰度，或者当需要单细胞分析以 解决生物学或生物医学问题。这一奋进的成功将推动 提出了许多领域的基础和应用生物医学研究，需要蛋白质组学分析， 这种方式类似于通过开发NeXT所取得的巨大进步， 代DNA测序。

Project Summary/Abstract Currently, most mass spectrometric (MS) analyses for proteomics are performed in sequential mode, wherein various species in a sample are selected and interrogated one after another. As a consequence of the finite time needed to examine each species in turn, sequential mode MS suffers from inescapable limitations in sensitivity, speed and ability to analyze all ions, especially when the composition of the ion beam is complex and rapidly changing. These limitations have kept vast tracts of biology and biomedicine, including, for example, deep single cell proteome analysis, out of reach of the current MS technology. In the present proposal, we posit that sensitivity, speed and dynamic range can be vastly improved by performing MS in parallel (by analogy to Next Generation DNA Sequencing), thus overcoming the technical barriers inherent to current commercial mass spectrometers that operate largely in sequential mode. Here, we propose to develop new MS instrumentation to execute MS in a massively parallel manner, with two major objectives in mind: Objective 1. Increase the sensitivity, speed and depth of proteome analyses by up to and ultimately beyond 1000-fold. The current sequential MS approaches can be likened to sampling the Niagara Falls with a bucket, where the majority of the sample is wasted. Our proposed parallel MS technology is designed to eliminate this immense waste. Objective 2. Filter noise in real time to eliminate unwanted ion background prior to MS analysis, thereby maximizing the MS utilization of the sample ions of interest and the resulting signal-to- noise ratios, as well as providing increased dynamic range to measure very low abundance components in the presence of highly abundant components. Successful attainment of these objectives will allow deep, comprehensive, high throughput analyses of proteomes in cases where sample availability is limiting, where the components of interest elude detection due to their low abundance, or when single cell analysis is needed to address the biological or biomedical question at hand. Success in this endeavor will propel forward many areas of basic and applied biomedical research that require proteomic analyses in a manner analogous to the immense progress that has been made through development of Next Generation DNA Sequencing.