Monoclonal Antibody Resources for C. elegans

线虫单克隆抗体资源

• 批准号: 7127597
• 负责人: MICHAEL L NONET
• 金额: $ 29.78万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-30 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7127597
• 关键词: Monoclonal; Antibody; Resources; C.; elegans

DESCRIPTION (provided by applicant): Antibodies (Abs) are widely used to detect specific cellular components both in situ and in biochemical experiments aimed at defining protein-protein interactions, protein processing, and reversible protein modifications such as phosphorylation. Polyclonal Abs are relatively simple to make and are widely used in molecular and cellular studies. However, such reagents are limited in quantity and thus rarely made widely available. Monoclonal Abs are more complicated to make, but hybridomas secreting the monoclonal antibody can be passaged ad infinitum permitting the identical reagent to be used by many different research group over long periods of time. Polyclonal and monoclonal Abs are available commercially that are directed against a huge array of vertebrate cellular components. By contrast, few such commercial reagents are available for work with model organisms such as C. elegans probably because the market for these reagents is too small to make them commercially viable. I propose to develop a 'tool kit' consisting of monoclonal Abs directed against a series of proteins that label discrete cellular components and subcellular compartments of C. elegans. The compartments include organelles common to all cells (e.g. endoplasmic reticulum, the Golgi, endosomes), organelles of specialized cells (e.g. synaptic vesicles), subcellular structures common to all cells (e.g. nuclear membrane, centrosomes and caveoli) and components that label domains in the developing embryo, germline, and nervous system. The cellular components include macromolecular complexes (e.g. splicesomes, proteosomes, replication origin complexes) and common cellular proteins (e.g clathrin, kinesin, dynein and intermediate filaments). The Abs will be thoroughly characterized and made publically available through the Developmental Studies Hybridoma Bank. The tool kit will be extremely useful for C. elegans researchers. It will provide a set of standardized renewable reference antibody tools available to the entire community. Secondly, it will greatly facilitate localizing other cellular components by providing a large set of cellular markers that can be used coordinately with rabbit polyclonal antibodies for co-localization studies. Finally, it will stimulate the use of biochemical approaches, such as subcellular fractionation and immunoprecipitations, by providing means to detect specific cellular compartments in extracts and the presence of specific components in immunoprecipitates.

描述（由申请人提供）： 抗体 (Abs) 广泛用于原位检测和生化实验中检测特定细胞成分，旨在定义蛋白质-蛋白质相互作用、蛋白质加工和可逆蛋白质修饰（例如磷酸化）。多克隆抗体制备相对简单，广泛用于分子和细胞研究。然而，此类试剂的数量有限，因此很少能广泛使用。单克隆抗体的制备更为复杂，但分泌单克隆抗体的杂交瘤可以无限传代，从而允许许多不同的研究小组在很长一段时间内使用相同的试剂。多克隆和单克隆抗体可在商业上买到，它们针对大​​量脊椎动物细胞成分。相比之下，很少有这样的商业试剂可用于模型生物，例如线虫，可能是因为这些试剂的市场太小，无法使其商业化。我建议开发一个“工具包”，其中包含针对一系列蛋白质的单克隆抗体，这些蛋白质标记了秀丽隐杆线虫的离散细胞成分和亚细胞区室。这些区室包括所有细胞共有的细胞器（例如内质网、高尔基体、内体）、特化细胞的细胞器（例如突触小泡）、所有细胞共有的亚细胞结构（例如核膜、中心体和小窝）以及标记域的组件发育中的胚胎、种系和神经系统。细胞成分包括大分子复合物（例如剪接体、蛋白体、复制起点复合物）和常见细胞蛋白（例如网格蛋白、驱动蛋白、动力蛋白和中间丝）。 Abs 将被彻底表征并通过发育研究杂交瘤库公开。该工具包对于线虫研究人员来说非常有用。它将提供一套可供整个社区使用的标准化可再生参考抗体工具。其次，它将通过提供大量细胞标记物来极大地促进其他细胞成分的定位，这些细胞标记物可以与兔多克隆抗体配合使用进行共定位研究。最后，它将通过提供检测提取物中特定细胞区室以及免疫沉淀物中特定成分的存在的方法，刺激生物化学方法的使用，例如亚细胞分级分离和免疫沉淀。