Experimental Animal Models, Department of Veterinary and Animal Sciences, University of Copenhagen, Denmark.
Global Drug Discovery, Novo Nordisk, Måløv, Denmark.
J Immunol Methods. 2019 Feb;465:20-26. doi: 10.1016/j.jim.2018.11.014. Epub 2018 Nov 27.
Translation across species of immunoassay results is often challenging due to the lack of cross-species reactivity of antibodies. In order to investigate the biology of insulin and IGF1 receptors, we generated new versatile monoclonal assay antibodies using the extracellular domain of the insulin/IGF1 hybrid receptor as the bait protein in the Adimab yeast antibody discovery platform and as the antigen in a rabbit monoclonal antibody platform. The resulting antibody clones were screened for receptor specificity as well as cross-species reactivity to both tissue and cell line derived samples. Using these strategies, we were able to identify highly specific insulin receptor monoclonal antibodies that lack cross-reactivity to the IGF1 receptor using the Adimab platform and a highly specific IGF1 receptor monoclonal antibody that lacks cross-reactivity to the insulin receptor using the rabbit antibody platform. Unlike earlier monoclonal antibodies reported in the literature, these antibodies show cross-species reactivity to the extracellular domains of mouse, rat, pig, and human receptors, indicating that they bind conserved epitopes. Furthermore, the antibodies work well in several different assay formats, including ELISA, flow cytometry, and immunoprecipitation, and therefore provide new tools to study insulin and IGF1 receptor biology with translation across several species and experimental model systems.
由于抗体缺乏跨物种反应性，因此将免疫测定结果从一个物种翻译到另一个物种通常具有挑战性。为了研究胰岛素和 IGF1 受体的生物学特性，我们使用胰岛素/IGF1 杂合受体的细胞外结构域作为诱饵蛋白在 Adimab 酵母抗体发现平台中，并作为抗原在兔单克隆抗体平台中生成了新的多功能单克隆检测抗体。筛选所得的抗体克隆以确定其受体特异性以及对组织和细胞系来源样本的交叉反应性。使用这些策略，我们能够使用 Adimab 平台鉴定出缺乏与 IGF1 受体交叉反应性的高度特异性胰岛素受体单克隆抗体，并且使用兔抗体平台鉴定出缺乏与胰岛素受体交叉反应性的高度特异性 IGF1 受体单克隆抗体。与文献中报道的早期单克隆抗体不同，这些抗体显示出对小鼠、大鼠、猪和人受体的细胞外结构域的跨物种反应性，表明它们结合保守表位。此外，这些抗体在几种不同的检测形式中效果良好，包括 ELISA、流式细胞术和免疫沉淀，因此为研究胰岛素和 IGF1 受体生物学特性提供了新的工具，可以在多个物种和实验模型系统中进行翻译。
