A novel pre-defined CDR library for selection of affinity reagents

用于选择亲和试剂的新型预定义 CDR 库

• 批准号: 9266529
• 负责人: MICHAEL P WEINER
• 金额: $ 65.01万
• 依托单位: ABCAM, INC.
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-15 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9266529
• 关键词: novel; pre; defined; CDR; library

 DESCRIPTION (provided by applicant): The most commonly used method for generating antibodies is through immunization of animals. However, this method is generally low-throughput, expensive, time-consuming, and the antibodies generated are not always renewable. Recombinant antibodies (rAb), like single chain variable fragments (scFv), have many attractive attributes compared to polyclonal antisera and monoclonal antibodies derived from hybridomas. They are renewable through overexpression in the appropriate heterologous host, they are easily stored and transferred as DNA, and they can be genetically engineered as fusions to various enzymes, fluorescent proteins, and epitope tags. While a number of approaches for generating recombinant affinity reagents exist, the cost and throughput of current technologies represent significant roadblocks to the development of a comprehensive and broadly available resource of renewable affinity reagents. We believe that improvements to both gene synthesis technologies and the increased affordability of high-throughput DNA sequencing can be leveraged to create antibody discovery pipelines based on synthetic biology that rival animal immune systems. In this proposal, we will build a high-throughput pipeline for recombinant antibody development in as few as 21 days. The proposed platform takes advantage of pre-designed diversity, next-generation sequence analysis, and advanced molecular biology techniques to enable the rapid identification of specific antibodies. Although our screening platform is being developed with single-chain variable fragment antibodies (scFvs), the technology is applicable to both Fab and yeast display libraries. High-throughput conversion of the scFvs to full immunoglobulin G (IgG) will be integrated within the pipeline so that the antibodies can be directly validated in the desired final format. In genomics, it was thought that the $1000 genome would be the inflection point at which whole genome sequencing would become commonplace. But even at $10,000 per genome, researcher uptake was phenomenal and new ways of using the NextGen sequencers, like ChIP Seq, RNA-seq, etc. were invented. In a similar fashion, we believe that at $5000 and 4 weeks, researchers will begin to develop new applications where the cost of producing antibodies is no longer a relevant factor and speed becomes everything. The ready availability of low cost, high quality affinity reagents will potentially accelerate all aspects of basic science research, provide diagnostics for disease biomarkers, and serve as a proof of concept for therapy. Like the $1000 genome, we think antibody identification and production can eventually go to under $100 and less than 2 weeks. At that point, whole proteome analyses for many different organisms and disease states will be possible. And new methods will arise.

 描述（由申请方提供）：最常用的产生抗体的方法是通过动物免疫。然而，这种方法通常是低通量、昂贵、耗时的，并且产生的抗体并不总是可再生的。重组抗体（rAb），如单链可变区片段（scFv），具有许多有吸引力的属性相比，来自杂交瘤的多克隆抗血清和单克隆抗体。它们通过在适当的异源宿主中过表达而可再生，它们易于作为DNA储存和转移，并且它们可以作为与各种酶、荧光蛋白和表位标签的融合物进行遗传工程改造。虽然存在许多用于产生重组亲和试剂的方法，但当前技术的成本和通量代表了开发全面且广泛可用的可再生亲和试剂资源的重大障碍。我们认为，基因合成技术的改进和高通量DNA测序的可负担性的提高可以用来创建基于合成生物学的抗体发现管道，与动物免疫系统相媲美。在本提案中，我们将在短短21天内建立一个用于重组抗体开发的高通量管道。该平台利用预先设计的多样性、下一代序列分析和先进的分子生物学技术，能够快速识别特异性抗体。虽然我们的筛选平台是用单链可变片段抗体（scFv）开发的，但该技术适用于Fab和酵母展示文库。将scFv高通量转化为完整的免疫球蛋白G（IgG）将被整合在管道内，使得抗体可以以所需的最终形式直接验证。在基因组学中，人们认为1000美元的基因组将是全基因组测序变得普遍的转折点。但即使是每个基因组10，000美元，研究人员的吸收也是惊人的，并且发明了使用NextGen测序仪的新方法，如ChIP Seq，RNA-seq等。以类似的方式，我们相信，在5000美元和4周，研究人员将开始开发新的应用程序，其中生产抗体的成本不再是一个相关的因素和速度成为一切。低成本、高质量亲和试剂的现成可用性将潜在地加速基础科学研究的各个方面，为癌症提供诊断， 疾病生物标志物，并作为治疗的概念证明。就像1000美元的基因组一样，我们认为抗体的鉴定和生产最终可以达到100美元以下，不到2周。到那时，许多不同生物体和疾病状态的全蛋白质组分析将成为可能。新的方法将会出现。