High Throughput Measurement of Cellular Signaling

细胞信号传导的高通量测量

基本信息

批准号：
7105123
负责人：
JOHN Michael RAMSEY
金额：
$ 42.5万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-09-01 至 2008-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): We will develop automated microfluidic devices for the high throughput measurements of kinase activation in human cells. These devices will ultimately be capable of detecting and characterizing the states of signal transduction pathways in cells by measuring and comparing the activity of multiple different kinases simultaneously in a large number of individual cells. The devices will automatically perform all of the processing and measurement steps required for high throughput measurements including cell transport, loading of kinase substrate peptides into the cells, cell lysis, and the separation and detection of the released peptides. The ratios of phosphorylated to nonphosphorylated substrates will be used as a measure of the degree of activation of each kinase assayed in each cell. Kinases play crucial roles in virtually all aspects of cellular physiology and are the focus of intense basic and pharmaceutical science research, thus these measurements have widespread applicability in the biomedical sciences. Moreover, by virtue of the heterogeneity of cellular behavior among individual cells in a population, high throughput single cell assays are critical for understanding how signaling networks interact to control cell physiology. Developing a technology, therefore, that is capable of determining the number, identities, and degree of activation of kinases in single cells will substantially advance our understanding of the molecular mechanisms of cellular signaling in health and disease.

描述（由申请人提供）：我们将开发自动化的微流体设备，用于人类细胞中激酶激活的高吞吐量测量。这些设备最终将能够通过测量和比较大量各个单个细胞中的多种不同激酶的活性来检测和表征细胞中信号转导途径的状态。设备将自动执行高吞吐量测量所需的所有处理和测量步骤，包括细胞转运，激酶底物肽在细胞中的负载，细胞裂解以及释放肽的分离和检测。磷酸化与非磷酸化底物的比率将用作每个细胞中每个激酶的激活程度的度量。激酶在细胞生理学的几乎所有方面都起着至关重要的作用，并且是激烈的基础和药学科学研究的重点，因此这些测量值在生物医学科学中广泛适用。此外，由于人群中个体细胞之间细胞行为的异质性，高吞吐量单细胞测定对于理解信号网络如何相互作用与控制细胞生理非常重要。因此，开发一种能够确定单个细胞中激酶的数量，身份和激活程度的技术将大大提高我们对健康和疾病中细胞信号的分子机制的理解。