APPLICATIONS OF HIGH SENSITIVITY DETECTION

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 该项目的总体目标是通过开发的方法和仪器的改进来扩大高灵敏度流式细胞术(HSFCM)的生物学应用，从而实现多参数测量能力。流式细胞术通过提供对单个细胞颗粒的快速、定量和灵敏的多参数测量，使生物医学科学取得了重大进展。个体分析产生了总体分析没有揭示的关于群体异质性的信息，并且允许比在整体阶段进行测量时更精确地测量个体属性。然而，传统的流式细胞术的一个局限性是无法测量小于0.5埃的小颗粒或含有少于数百个荧光分子的暗颗粒。许多重要的生物粒子、分子和分子组件都属于这些类别。我们的方法是通过增强能力的HSFCM访问该测量域。我们已经确定了三个重要的生物学领域：通过基因组分析进行细菌鉴定；DNA单分子扫描在高通量基因组学中的应用；以及线粒体基因组和生化分析。多色和光散射检测方面的特定仪器增强将使这些新应用成为可能。该研究计划分为四个相关的具体目标：基于DNA片段大小的细菌指纹分析；单个DNA分子的高通量扫描；单个线粒体的分析；以及高灵敏度仪器的开发。这一研究开发的成果将开创一类新的高灵敏度的流式细胞仪，并将在亚细胞单分子水平上刺激流式细胞术的新领域。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The overall goal of this project is to expand the biological applications of High Sensitivity Flow Cytometry (HSFCM) through methods developed and through instrumentation improvements leading to a multiparameter measurement capability. Flow cytometry has enabled major advances in the biomedical sciences by providing rapid, quantitative and sensitive multiparameter measurements of individual cellular particles. Individual analysis produces information on population heterogeneity that is not revealed by ensemble analysis and allows more precise measurement of individual attributes than is possible when measurement is done in bulk phase. However, one limitation of conventional flow cytometry is the inability to measure small particles less than 0.5 ¿m or dim particles having less than several hundred fluorescent molecules. A wide variety of important biological particles, molecules, and molecular assemblies fall into these categories. Our approach is to access this measurement domain through enhanced capability HSFCM. We have identified 3 important biological areas to focus on: bacterial identification by genome analysis; DNA single molecule scanning applications in high throughput genomics; and mitochondrial genomic and biochemical analysis. Specific instrument enhancements in multi-color and light scatter detection will enable these new applications. The research plan is divided into four related Specific Aims: DNA Fragment Size-Based Analysis of Bacterial Fingerprints; High Throughput Scanning of Single DNA Molecules; Analysis of Individual Mitochondria; and High Sensitivity Instrumentation Development. The results of this research and development will create a new class of high sensitivity flow cytometry instrumentation and will stimulate a new field of flow cytometry at the sub-cellular single molecule level.