Cryoultramicrotomy for Subcellular Localization of Molecules

用于分子亚细胞定位的冷冻超薄切片术

• 批准号: 9970143
• 负责人: Marcia Miller
• 金额: --
• 依托单位: Beckman Research Institute City of Hope
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-15 至 2001-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9970143&HistoricalAwards=false
• 关键词: Cryoultramicrotomy; Subcellular; Localization; Molecules

9970143Abstract This project involves the acquisition, installation and use of instrumentation that will allow investigators at the City of Hope to localize molecules quantitatively in thin cryosections of tissue imaged by electron microscopy. Knowledge of the molecular basis of life has grown considerably more detailed since the inception of electron microcopy for the examination of ultrathin sections of resin embedded tissues. The need to be able to examine particular molecules at subcellular sites has driven the development of a means to make ultrathin sections of frozen cells and tissues. Such cryosections can be labeled robustly with antibodies and other molecules such as fusion proteins and derivitized RNAs, and immunogold to localize these molecules to regions of specialized cellular activity. City of Hope investigators have vigorous programs that can take advantage of this type of information in their studies of cell function. Projects include studies of synaptic transmission; CD44a and its role in a morphogenetic model; the fate of products of transgenes such as Bcl-XL and mdr1; the fate of the major histocompatibility complex encoded B-G molecules in virally-infected cells; and localization of p53. The equipment being acquired will make possible the quantitative analysis of cellular localization and co-localization in these and other projects at the City of Hope using a combination of ultrathin cryosections and immunogold labeling.

9970143Abstract 该项目涉及仪器的采购、安装和使用，这些仪器将使希望之城的研究人员能够在电子显微镜成像的组织薄冷冻切片中定量定位分子。自从用于检查树脂包埋组织超薄切片的电子显微镜问世以来，对生命分子基础的了解已经变得更加详细。能够检查亚细胞位点的特定分子的需求推动了一种制作冷冻细胞和组织超薄切片的方法的开发。这种冷冻切片可以用抗体和其他分子（例如融合蛋白和衍生化 RNA）和免疫金进行牢固标记，以将这些分子定位到特定细胞活性的区域。希望之城的研究人员制定了强有力的计划，可以在细胞功能研究中利用此类信息。项目包括突触传递的研究； CD44a 及其在形态发生模型中的作用； Bcl-XL 和 mdr1 等转基因产物的命运；病毒感染细胞中主要组织相容性复合体编码的 B-G 分子的命运；和p53的定位。正在购买的设备将使希望之城的这些项目和其他项目结合超薄冷冻切片和免疫金标记对细胞定位和共定位进行定量分析成为可能。