Recombinant antibodies for antibody arrays

描述（由申请人提供）：我们提出开发一种快速、低成本和高通量的方法，用于产生具有改进性质的重组工程抗体，用于蛋白质组学和免疫测定应用。工程化抗体构建体如重组Fab片段和嵌合抗体将通过直接修饰产生抗体的杂交瘤细胞系中的抗体基因来产生和表达，从而消除对基因克隆和转染的需要。在该I期SBIR中，我们将通过修饰十（10）个杂交瘤细胞系来证明可行性，这些杂交瘤细胞系产生MSD商业免疫测定试剂盒中使用的抗体。我们将证明，我们可以修饰细胞系，使得它们将其抗体表达为重组Fab片段或重组嵌合体，所述重组嵌合体包含与能够进行位点特异性标记的蛋白质结构域连接的Fab片段。 使用来自已建立的试剂盒的抗体将使得直接比较新抗体构建体与原始全长抗体的测定性能变得简单。该计划的长期目标是利用嵌合构建体的有益特性-较低的非特异性相互作用和通过位点特异性标记控制固定化的能力-以开发用于蛋白质组学应用的高密度阵列。这些改进的多重免疫测定将在研究和临床诊断应用中具有重要价值。

DESCRIPTION (provided by applicant): We propose to develop a rapid, low cost and high-throughput method for generating recombinant engineered antibodies with improved properties for proteomic and immunoassay applications. Engineered antibody constructs such as recombinant Fab fragments and chimeric antibodies will be generated and expressed by directly modifying the antibody genes in antibody-producing hybridoma cell lines, thereby eliminating the need for gene cloning and transfection. In this Phase I SBIR, we will demonstrate feasibility by modifying ten (10) hybridoma cell lines that produce antibodies used in MSD's commercial immunoassay kits. We will demonstrate that we can modify the cell lines so that they express their antibodies as recombinant Fab fragments or recombinant chimeras comprising Fab fragments linked to protein domains that enable site- specific labeling. The use of antibodies from established kits will make it straightforward to directly compare the assay performance of the new antibody constructs to the original full length antibodies. The long term goal of the program is to take advantage of beneficial properties of the chimeric constructs - lower non-specific interactions and the ability to control immobilization through site-specific labeling - to enable the development of high density arrays for proteomic applications. These improved multiplexed immunoassays will have significant value in both research and clinical diagnostics applications.