Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Komaba 3-8-1, Meguro-ku, Tokyo 153-8902, Japan.
Nucleic Acids Res. 2011 Feb;39(3):e14. doi: 10.1093/nar/gkq1122. Epub 2010 Nov 9.
The recent development of screening strategies based on the generation and display of large libraries of antibody fragments has allowed considerable advances for the in vitro isolation of monoclonal antibodies (mAbs). We previously developed a technology referred to as the 'ADLib (Autonomously Diversifying Library) system', which allows the rapid screening and isolation in vitro of antigen-specific monoclonal antibodies (mAbs) from libraries of immunoglobulin M (IgM) displayed by the chicken B-cell line DT40. Here, we report a novel application of the ADLib system to the production of chimeric human mAbs. We have designed gene knock-in constructs to generate DT40 strains that coexpress chimeric human IgG and chicken IgM via B-cell-specific RNA alternative splicing. We demonstrate that the application of the ADLib system to these strains allows the one-step selection of antigen-specific human chimeric IgG. In addition, the production of chimeric IgG can be selectively increased when we modulate RNA processing by overexpressing the polyadenylation factor CstF-64. This method provides a new way to efficiently design mAbs suitable for a wide range of purposes including antibody therapy.
基于抗体片段的产生和展示的大规模文库的筛选策略的最新发展，使得体外分离单克隆抗体（mAbs）取得了相当大的进展。我们之前开发了一种称为“ADLib（自主多样化文库）系统”的技术，该技术允许从鸡 B 细胞系 DT40 展示的免疫球蛋白 M（IgM）文库中快速筛选和分离体外抗原特异性单克隆抗体（mAbs）。在这里，我们报告了 ADLib 系统在生产嵌合人 mAbs 中的一种新应用。我们设计了基因敲入构建体，以生成共表达嵌合人 IgG 和鸡 IgM 的 DT40 株，通过 B 细胞特异性 RNA 可变剪接。我们证明，将 ADLib 系统应用于这些菌株可以一步选择抗原特异性人嵌合 IgG。此外，当我们通过过表达多聚腺苷酸化因子 CstF-64 来调节 RNA 加工时，可以选择性地增加嵌合 IgG 的产生。这种方法为设计适合广泛用途的 mAbs 提供了一种新方法，包括抗体治疗。
