Collaborative Research: Single Cell Proteomics

该奖项为一个联合项目提供支持，该项目旨在开发基于质谱法的蛋白质序列分析的新分析策略。这项工作涉及路易斯安那州立大学的研究人员，他们擅长制造微米大小的设备，以及他们在辛辛那提大学的合作者，他们擅长高分辨率质谱分析。微加工技术将用于开发集成的微系统，预计可以分析单个细胞中的蛋白质。微流体的使用将有助于将蛋白质维持在足够高的浓度，以便进行分析，即使在细胞中发现的每种蛋白质的数量都在3000分子左右。在这个项目中使用的设备是使用聚合物和微制造技术来制造热压聚合物的高纵横比模塑模具。这些装置将通过研究含有至少150种不同蛋白质的叶绿体衍生膜来测试和改进。在拟议的努力范围内，这些研究将扩大到检查蛋白质组，即完整的阿拉伯芥叶绿体和蓝绿藻聚囊藻的整个细胞的全部蛋白质。如果完全成功，该仪器将允许对单个细胞进行生化分析，从而有助于更好地理解基因表达和细胞表型的关系。

This award provides support for a joint project aimed at the development of aggressive new analytical strategies for protein sequence analysis based on mass spectrometry. The effort involves investigators at Louisiana State University, who are accomplished in the manufacture of micrometer-sized devices, and their collaborator at the University of Cincinnati, who is skilled in high resolution mass spectrometry. Microfabrication techniques will be used to develop integrated micro-systems expected to permit analysis of the proteins in a single cell. The use of microfludics will help maintain the proteins at a high enough concentration to permit analysis even though the amount of each protein found in a cell is on the order of 3000 molecules. The devices to be used in this project are built using polymers and micromanufacturing techniques to fabricate high-aspect-ratio molding dies for hot embossing polymers. The devices will be tested and refined through studies of chloroplast-derived membranes thought to contain at least 150 different proteins. Within the context of the proposed effort, these studies will be extended to examine the proteome, i.e., the entire complement of proteins, of intact chloroplasts of Arabadopsis and of entire cells of the blue-green algae Synechocystis. If fully successful, the instrumentation will allow biochemical analyses of individual cells and thus contribute to a better understanding of the relationship of gene expression and cellular phenotype.