Emulsion phage display for high-throughput isolation of affinity binders

描述（由申请人提供）：我们建议开发一种廉价的高通量方法来选择和鉴定针对未修饰和修饰的蛋白质、肽和非肽抗原的抗体。本提案的具体目标是完成概念验证，使用油乳剂中的水滴来鉴定和分离单链猪瘟病毒抗体。重组噬菌体产生大肠杆菌细胞的区隔化用于单独查询每个scFv结合抗原包被珠的能力。我们建议在Her2蛋白上发现的磷酸肽表位上开发和测试这些方法。SBIR和随后的II期目标的成功完成将对新的治疗靶点、免疫治疗、预后和诊断生物标志物的开发产生相当大的直接影响。公共卫生相关性：基因阵列使我们对细胞和组织中发生的潜在生物过程的理解取得了重大突破。这些增进的认识已经导致了诊断和治疗主要疾病的新方法，包括癌症、糖尿病和阿尔茨海默氏症。我们现在正进入一个后基因组时代，对蛋白质组学工具的需求变得越来越重要。蛋白质组学工具的技术挑战远远大于基因组计划中遇到的挑战。本提案描述了用于测量蛋白质丰度的抗体的高通量制造。此次SBIR的成功完成和我们未来的联合目标将对新的治疗靶点以及免疫治疗、预后和诊断生物标志物的开发产生重大的直接影响。

DESCRIPTION (provided by applicant): We propose to develop an inexpensive high-throughput means of selecting and identifying antibodies to both unmodified and modified protein, peptide, and non-peptide antigens. The specific goals of this proposal are to complete a proof-of-concept using aqueous droplets in an oil emulsion for identification and isolation of scFv antibodies. Compartmentalization of recombinant-phage-producing E.coli cells is used to individually query the ability of each scFv to bind to antigen-coated beads. We propose to develop and test these methods on a phosphopeptide epitope found on the protein Her2. The successful completion of this SBIR and subsequent Phase II goals will have considerable direct impact on the development of new therapeutic targets, and immunotherapeutic, prognostic, and diagnostic biomarkers. PUBLIC HEALTH RELEVANCE: Gene arrays enabled great breakthroughs in our understanding of the underlying biological processes that occur in cells and tissues. These increased understandings have already led to new approaches to diagnosis and treatment of major diseases, including cancer, diabetes and Alzheimer's. We are now entering a post-genomic era where the need for proteomics tools is becoming increasingly important. The technical challenges for proteomic tools are far greater than those encountered during the genome project. This proposal describes the high-throughput manufacture of antibodies for measuring protein abundance. The successful completion of this SBIR and our combined future goals will have significant direct impact on the development of new therapeutic targets as well as immunotherapeutic, prognostic, and diagnostic biomarkers.