NINDS/UC Davis NeuroMab Hybridoma Facility

描述(由申请人提供)： 这项建议的具体目的是在NINDS/加州大学戴维斯分校的NeuroMab设施中开发一个针对脑内使用而优化的单抗(MAbs)的综合文库。这一提议是因为需要极大地扩大这种大脑优化的mAbs的可用性，用于基础、翻译和临床神经科学研究。高通量后基因组方法正在以快速的速度产生数据，这些方法涉及大脑发育、神经元可塑性以及神经和精神障碍的分子机制。作为进一步基础研究或治疗学发展的潜在靶点的候选基因的确认，取决于这些基因的蛋白质产物的特性。针对已定义的已消失产物的单抗可以作为大脑中表达的基因清单与深入了解其产品如何决定大脑功能之间的重要桥梁。然而，许多必要的试剂要么无法获得，要么在有的情况下用于哺乳动物的大脑时缺乏有效性和特异性。在人类、非人类灵长类动物和啮齿类动物的大脑中使用经过优化的高质量、可靠的单抗(即神经单抗)对于神经科学的各个领域都是至关重要的。将首先利用人类、小鼠和大鼠基因组计划产生的丰富数据，产生与神经元蛋白片段相对应的重组和/或合成免疫原，从而建立一个全面的神经单抗文库。这些将用于强化免疫方案，在相对较短的免疫期内产生大量分泌免疫球蛋白的杂交瘤。这些大型杂交瘤细胞池将筛选出识别异种细胞中同源抗原的单抗，然后对整个阳性细胞池进行全面的生化和免疫组织化学分析，以确定其在脑内的有效性和特异性。由此产生的大脑优化的神经单抗将以非常低的成本提供给研究界，作为组织培养上清液或作为浓缩的免疫球蛋白制剂。分泌神经单抗的杂交瘤也将免费提供。研究人员将使用这些神经单抗来确定发育中的、成人的、老年的和患病的大脑中同源抗原的存在和相对丰度，它们的细胞和亚细胞定位，功能相关的翻译后修饰，以及蛋白质-蛋白质相互作用。此外，NeuroMabs可能会在蛋白质的直接功能分析、诊断程序和治疗学中发现更多的应用。

DESCRIPTION (provided by applicant): The specific aim of this proposal is to develop a comprehensive library of monoclonal antibodies (mAbs) optimized for use in the brain (i.e. NeuroMabs) at the NINDS/UC Davis NeuroMab Facility. This proposal is driven by the need to greatly expand the availability of such brain-optimized mAbs for use in basic, translational and clinical neuroscience research. Data are being generated at a rapid rate from high throughput post-genomic approaches addressing molecular mechanisms of brain development, neuronal plasticity, and neurological and psychiatric disorders. The validation of candidate genes as potential targets for further basic research, or for the development of therapeutics, relies on characterization of the protein products of these genes. MAbs against defined gone products can serve as the crucial bridge between the inventory of genes expressed in the brain, and insights into how their products determine brain function. However, many of the necessary reagents are either unavailable, or when available suffer from a lack of efficacy and specificity when used in mammalian brain. The availability of high-quality, reliable mAbs that have been optimized for use in human, non-human primate, and rodent brain (i.e. NeuroMabs) is of utmost importance to virtual all areas of neuroscience. The generation of a comprehensive library of NeuroMabs will be pursued by first taking advantage of the wealth of data emerging from the human, mouse and rat genome projects to generate recombinant and/or synthetic immunogens corresponding to fragments of neuronal proteins. These will be used in an intense immunization protocol that yields large numbers of IgG-secreting hybridomas from a relatively short immunization period. These large hybridoma pools will be screened for those mAbs that recognize the cognate antigen in heterologous cells, and then the entire positive pool subjected to comprehensive biochemical and immunohistochemical analyses of their efficacy and specificity in brain. The resultant brain optimized NeuroMabs will be made available at very low cost to the research community as tissue culture supernatants or as concentrated IgG preparations. The NeuroMab secreting hybridomas will also be made freely available. Investigators will use these NeuroMabs for determining the presence and relative abundance of the cognate antigens in developing, adult, aged, and diseased brain, their cellular and subcellular localization, functionally relevant post-translational modifications, and protein-protein interactions. Moreover, NeuroMabs may find additional i applications in direct functional analyses of proteins, in diagnostic procedures, and as therapeutics.