CORE--ANTIBODY AND IMMUNOCYTOCHEMISTRY

核心--抗体与免疫细胞化学

• 批准号: 6301887
• 负责人: ROBERT J BLOCH
• 金额: $ 17.82万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-05-01 至 2001-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6301887
• 关键词: astrocytes; biomedical facility; chickens; chimeric proteins; confocal scanning microscopy; enzyme linked immunosorbent assay; immunocytochemistry; immunofluorescence technique; immunoprecipitation; laboratory mouse; laboratory rabbit; monoclonal antibody; neurons; protein purification; western blottings

Antibodies are now routinely used to identify proteins in the central nervous system, to estimate the amounts of different kinds of proteins present, and to determine in which cellular population and subcellular region particular proteins are concentrated. Over the past 8 years, this program project has had a significant component dedicated to the preparation or use of antibodies, but it was limited largely to a single research project. We now propose to expand the use of antibodies in this program to take greater advantage of their potential for use in addressing questions related both to the structure and function of astrocytes and neurons, and in elucidating how these to major cell populations in the brain interact. The purpose of this Core is to provide and characterize immunological reagents, as well as expertise in their use, to all the participants in this program project. Some antibodies, available from commercial sources or from outside collaborators, will be characterized and, if necessary, further purified to allow them to be used to immunoprecipitation, immunoblotting, and immunofluorescence experiments. The Core will also generate its own antibodies. Purified proteins, polypeptide antigens, fusion proteins, and even crude protein mixtures, will be used as antigens for the generation of polyclonal antibodies in rabbits or chickens, and monoclonal antibodies in mice. The Core will select appropriate antigens, preparing then "in house" whenever possible. Animals will also be injected and boosted "in house", and pre-immune, and immune sera will be collected. The Core will purify these antibodies as needed, characterize their specificity by ELISA, immunoblotting, and immunoprecipitation, and test their usefulness in immunofluorescence protocols. Experimental assistance in all these techniques, as well as in confocal laser scanning microscopy, will be provided to all participants in the program. We anticipate that all of the projects as well as the Tissue Culture Core in this program project will benefit from the Immunology Core.

现在通常使用抗体来识别中央的蛋白质 神经系统，以估计不同种类的蛋白质的量 目前，并确定在哪个细胞种群和亚细胞中 区域特定的蛋白质浓缩。在过去的8年中， 计划项目的重要组成部分致力于 制备或使用抗体，但主要限于一个 研究项目。我们现在建议在此扩大抗体的使用 计划更大地利用其用于解决的潜力 与星形胶质细胞的结构和功能有关的问题和 神经元，并阐明这些神经元如何在主要细胞种群中 大脑相互作用。 该核心的目的是提供和表征免疫学 对所有参与者的试剂以及其使用方面的专业知识 这个程序项目。一些抗体，可从商业来源获得 或从外部合作者中进行表征，如有必要， 进一步纯化以允许它们用于免疫沉淀， 免疫印迹和免疫荧光实验。核心也将 产生自己的抗体。纯化的蛋白质，多肽抗原， 融合蛋白，甚至粗蛋白混合物都将用作抗原 为了产生兔子或鸡中的多克隆抗体，以及 小鼠的单克隆抗体。核心将选择适当的抗原， 随时准备“在房子里”。动物也会被注射 并将收集“在房屋中”，免疫前和免疫血清。 核心将根据需要纯化这些抗体，表征它们 ELISA的特异性，免疫印迹和免疫沉淀和测试 它们在免疫荧光方案中的有用性。实验协助 在所有这些技术以及共聚焦激光扫描显微镜中， 将提供给该计划的所有参与者。 我们预计所有项目以及组织培养核心 在这个程序中，项目将受益于免疫学核心。