CORE--RADIOIMMUNOASSAY OF AND PRODUCTION OF ANTISERA

核心--抗血清的放射免疫测定和生产

• 批准号: 6577257
• 负责人: CATHERINE L RIVIER
• 金额: $ 19.74万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6577257
• 关键词: antiserum; biomedical facility; follistatin; guinea pigs; immunological substance; inhibin; laboratory rabbit; laboratory rat; peptide hormone; radioimmunoassay; synthetic peptide

This Program Project focuses on the role of activins/inhibins and their binding proteins, receptors and downstream modulators in the regulation of reproductive function, cellular growth and development. The first aim of this Program is to characterize the structure of activin-activin/receptor complexes and physical interactions between activin, its receptors and binding proteins, and downstream regulatory molecules. The second aim is to explore the physiologic and pathophysiologic significance of these molecules at the cellular and system levels and to study the control of protein expression and secretion as well as modes of action. These studies require high affinity, high titer antibodies of appropriate specificity for immunoprecipitation, immunoblot radioimmunoassay (RIA), Immunohistochemistry, and/or immunoneutralization techniques. This Core will produce and characterize polyclonal antisera needed by the immunoneutralization techniques. This Core will produce and characterize polyclonal antisera needed by the Projects of this Program to carry out structural, functional and mechanistic studies. We will use synthetic peptides conjugated to carrier proteins as immunogens to develop antibodies directed towards gene products predicted from the cDNA sequences or from amino acid sequencing of tryptic fragments of purified natural products. Alternatively, proteins produced using in vitro cell expression systems and provided by individual Projects will be injected directly into host animals for production of anti-sera. To reduce cross-reactivity with antigens other than the protein of interest, selected antisera will be immunoaffinity purified using the cognitive peptide, recombinant protein, or fusion protein covalently bound to an agarose- based gel. Physiologic and pharmacologic studies proposed by the Projects in this Program are critically dependent upon our ability to measure relevant hormones in a variety of biological fluids. The core will provide RIAs of pituitary gonadotropic hormones and sex steroids. The Core is responsible for ensuring quality control of these assays. The core will be under the overall responsibility of Drs. C. Rivier (RIA analysis of biological fluids, 10% effort) and W. Vale (antibody production, 3% effort). Day to day operation of the antisera production and purification component of Core B will be under the supervision of J. Vaughan (10% effort).

该程序项目的重点是激活素/抑制素及其结合蛋白，受体和下游调节剂在生殖功能，细胞生长和发育中的作用。该程序的第一个目的是表征激活素活性蛋白/受体复合物的结构以及激活素，其受体和结合蛋白与下游调节分子之间的物理相互作用。第二个目的是探索这些分子在细胞和系统水平上的生理和病理生理意义，并研究蛋白质表达和分泌的控制以及作用方式。这些研究需要具有适当特异性的高滴度抗体，具有适当特异性的免疫沉淀，免疫印迹放射免疫测定（RIA），免疫组织化学和/或免疫中性化技术。该核心将产生并表征免疫中和技术所需的多克隆抗血清。该核心将产生并表征该计划的项目所需的多克隆抗血清，以进行结构，功能和机械研究。我们将使用与载体蛋白结合的合成肽作为免疫原来开发针对从cDNA序列预测的基因产物或纯天然产物胰蛋白酶片段的氨基酸测序的抗体。或者，使用体外细胞表达系统产生的蛋白质并由各个项目提供的蛋白质将直接注入宿主动物中，以生产抗疾病。为了降低与感兴趣蛋白以外的抗原的交叉反应性，使用认知肽，重组蛋白或融合蛋白共价结合与琼脂糖基凝胶的认知肽，重组蛋白或融合蛋白进行免疫亲和力纯化。该计划中该项目提出的生理和药理学研究在很大程度上取决于我们测量各种生物流体中相关激素的能​​力。核心将提供垂体性促性腺激素激素和性类固醇的动力。核心负责确保对这些测定的质量控制。核心将承担DRS的总体责任。 C. rivier（生物流体的RIA分析，10％的努力）和W. Vale（抗体产生，3％的努力）。 Antisera生产和核心B的纯化成分的日常运行将在J. Vaughan的监督下（10％的努力）。