An Integrated Microfluidic System for the Combining Top-Down and Bottom-Up Proteomics

项目总结/摘要 系统生物学旨在了解健康的生物系统如何工作，以及当 它们停止正常工作并变得病态， 并模拟它们如何变化和相互作用。生物系统可以是细胞、组织、有机体，甚至是细胞。 生物群落。蛋白质构成了细胞内几乎所有的结构，因此它们在很大程度上 影响与疾病相关的适当生物功能和功能障碍。因此，通过改善 蛋白质组分析可以提高我们对生物系统的正常功能和功能障碍的理解。 该研究项目的长期目标是提供改进的蛋白质组分析方法， 增加了可以从生物样品中分析的蛋白质的细节。即， 鉴定的蛋白质型将大大提高，并超过用肽鉴定的蛋白质的数量 基于碎片离子谱的方法。为了在蛋白质型检测中实现这一大的改进， 将开发一种新型的微流控系统，该系统将整合完整的蛋白质和肽识别策略 以无缝和高度自动化的格式。拟议工作的具体目标将包括： 一种高效的二维快速样品分离微流控系统的研制 处理.将通过鲍曼不动杆菌分析确定器械性能 和人Jurkat细胞，分别代表相对简单和复杂的蛋白质组。除了 作为一个相对简单的模型，A.鲍曼不动杆菌是临床相关的，因为它引起人类感染， 正在以惊人的速度获得抗生素耐药性。人类jurkat细胞参与了人类免疫系统 反应，并已高度研究和表征。这种高度的jurkat细胞表征将 提供了一个很好的基准，以比较所提出的微流控蛋白质组学系统的性能 与其他常用的蛋白质组学方法相比。

Project Summary/Abstract Systems biology seeks to understand how healthy biological systems work, and what goes wrong when they stop working properly and become diseased gaining a holistic view of all of the biochemical components and modeling how they change and interact. The biological system can be a cell, tissue, organism, or even community of organisms. Proteins form nearly all of the machinery within a cell and consequently they greatly influence proper biological function and dysfunction that is associated with disease. Thus, by improving proteome analysis we can improve our understanding of proper function and dysfunction of biological systems. The long-term objective of this research project is to provide improved methods for proteome analysis that increase the detail of the proteins that can be analyzed from a biological sample. Namely, the number of proteoforms identified will be greatly improved and surpass the number of proteins identified with peptide fragment ion spectra based methods. To achieve this large improvement in the detection of proteoforms, a novel microfluidic system will be developed that will integrate intact protein and peptide identification strategies in a seamless and highly automated format. The specific aims of the proposed work will include development of a high-efficiency two-dimensional separation and development of a microfluidic system for rapid sample processing. The performance of the device will be determined through the analysis of Acinetobacter baumannii and human jurkat cells, which represent relatively simple and complex proteomes, respectively. In addition to being a relatively simple model, A. baumannii is clinically relevant because it causes infections in humans and is gaining antibiotic resistance at an alarming rate. Human jurkat cells are involved in the human immune response and have been highly studied and characterized. This high degree of jurkat cell characterization will provide an excellent benchmark to compare the performance of the proposed microfluidic proteomic system against other commonly used proteomics approaches.