High-throughput reformatting of phage-displayed antibody fragments to IgGs by one-step emulsion PCR
High-throughput reformatting of phage-displayed antibody fragments to IgGs by one-step emulsion PCR
复制标题
通过一步乳液 PCR 将噬菌体展示的抗体片段高通量重新格式化为 IgG。
DOI：
10.1093/protein/gzz004
复制
发表时间：
2018-11-01
影响因子：
2.4
通讯作者：
Zhang, Hongkai
中科院分区：
文献类型：
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作者：
Liu, Yaohui;Gu, Manping;Zhang, Hongkai
单链抗体是噬菌体展示抗体库中最常用的形式。分离的scFv需要重新格式化为全长IgG以用于进一步表征。在不破坏VH-VL配对的情况下将scFv高通量重新格式化为IgG对于IgG形式的所有抗体的彻底筛选是非常苛刻的。在此，我们开发了一种基于乳液中重叠延伸PCR的策略，以将scFv重新格式化为IgG，同时保持可变区配对的准确性和复杂性。使用CD 40作为示例靶标，我们将噬菌体展示衍生的CD 40结合scFv文库重新格式化为IgG哺乳动物展示文库，并分离高亲和力CD 40结合IgG。这种稳健和可靠的抗体重组方法可以整合到任何基于噬菌体展示的抗体药物发现中。
Single-chain variable fragment (scFv) is the most common format for phage display antibody library. The isolated scFvs need to be reformatted to full-length IgGs for further characterization. High throughput reformatting of scFv to IgG without disrupting VH-VL pairing is of great demanding for exhaustive screening of all antibodies in IgG format. Herein, we developed a strategy based on the overlap extension PCR in emulsion to reformat scFv to IgG while maintain the accuracy and complexity of variable region pairing. Using CD40 as an example target, we reformatted phage display derived CD40 binding scFv library to IgG mammalian display library and isolated high affinity CD40 binding IgGs. This robust and reliable antibody reformatting approach could be integrated into any phage display based antibody drug discovery.