SYSTEM FOR QUANTITATIVE 3D CONFOCAL LIGHT MICROSCOPY

我们对细胞及其功能结构的理解 与遗传和发育事件的关系直到最近 是慢慢获得的。这是因为研究细胞结构的工具 要么使用(电子显微镜)繁琐，要么在 细胞由于缺乏分辨率(常规光学显微镜)。其目的是 这项建议的目的是获得一个研究功能三维的系统 基于革命性的共焦新技术的细胞体系结构 光学显微镜。该系统将由七个核心小组使用 细胞和发育生物学家共同参与了七个 专注于所涉及的分子事件的独立研究领域 在细胞内细胞器的通讯、组装和发育中。这些 主要研究内容有：1)形态梯度的产生机制及其 在胚胎发育中的应用；2)合子的物理基础 线粒体膜混合及其遗传后果；3)分析 极化细胞中蛋白质分选的机制；4)分析 蛋白质折叠及其在细胞器运动中的作用 膜；5)内肿瘤形成的筛选和特征 转基因小鼠的不同组织；6)生理和遗传 核染色质附着部位的分析；7) 转位复合体的构建和遗传分析 通过核包膜进行运输。 这一核心用户群体研究的小区内位置是多样的； 然而，我们都需要一种快速和灵敏的分析技术 组织中0.1微米到1.0微米分辨率范围内的细胞结构 以及厚度在0.1至0.5 mm范围内的单元格。不同的成员 的核心小组已经探索了激光扫描共聚焦的用途 并发现这项新开发的技术可以使 可用于分析的新可能性，而以前没有 可用。

Our understanding of the functional architecture of the cell and its relationship to both genetic and developmental events has until recently been gained slowly. This is because the tools for studying cell structure have been either tedious to use (electron microscopy) or ineffective in cells due to lack of resolution (conventional light microscopy). The aim of this proposal is to acquire a system for studying the functional 3-D architecture of cells based on a revolutionary new technique of confocal light microscopy. This system will be utilized by a core group of seven cell and developmental biologists who collectively are engaged in seven independent areas of research which focus on the molecular events involved in intracellular organelle communication, assembly and development. These areas are: 1) the mechanism of morphometric gradient generation and its utilization in the developing embryo; 2) the physical basis of zygotic mitochondrial membrane mixing and its genetic consequences; 3) analysis of the mechanism of protein sorting in the polarized cell; 4) analysis of protein folding and its role in protein movement through organelle membranes; 5) screening and characterization of tumorogenesis within different tissues of the transgenic mouse; 6) the physical and genetic analysis of chromatin attachment sites within the cell nucleus; 7) architecture and genetic analysis of the translocation complex mediating transport through the nuclear envelope. The intracellular locations studied by this core user group are diverse; however, we share the need for a rapid and sensitive technique for analysis of cell architecture in the 0.1 to 1.0 micron resolution range in tissues and cells with thicknesses in the range 0.1 to 0.5 mm. Different members of the core group have explored the use of laser scanning confocal microscopy and have found that this newly developed technique makes available new possibilities for analysis which previously were not available.