It has been observed that converting scFv formatted antibodies to full-length IgG often associates with loss of affinity. We aim to address this issue in this paper by establishing an integrated affinity maturation method applying yeast display technology platform. To demonstrate that, we employed a human thrombopoietin receptor targeting antibody named 3D9 which was identified previously from a combinational antibody library in scFv-Fc fusion protein form. We have observed that significant potency loss happened when 3D9 was transformed to full-length IgG form. In this study, we tested whether the potency of the full-length IgG can be improved by affinity maturation of 3D9 using a modified Fab yeast display platform. An efficient CDR3 targeted mutagenesis strategy was designed for Fab library with pre-designed CDR diversity. Next generation sequencing was also used for evaluation of the enrichment process and investigation of sequence-function relationship of the antibody. A variant with improved affinity and higher potency was identified. The study demonstrates that the strategy we used here are efficient for optimizing affinity and activity of full-length IgGs.

中文翻译：

---

全长IgG形式的TpoR靶向抗体的亲和力成熟，可增强激动剂活性。

已经观察到，将scFv格式化的抗体转化为全长IgG通常与亲和力丧失有关。我们旨在通过使用酵母展示技术平台建立集成的亲和力成熟方法来解决此问题。为了证明这一点，我们使用了人类血小板生成素受体靶向抗体3D9，该抗体先前已从scFv-Fc融合蛋白形式的组合抗体库中鉴定出来。我们已经观察到，当3D9转化为全长IgG形式时，发生了明显的效能损失。在这项研究中，我们测试了使用改良的Fab酵母展示平台通过3D9的亲和力成熟是否可以提高全长IgG的效力。针对具有预先设计的CDR多样性的Fab库设计了有效的CDR3靶向诱变策略。下一代测序也用于评估富集过程和研究抗体的序列-功能关系。鉴定了具有改善的亲和力和较高效力的变体。该研究表明，我们在此处使用的策略对于优化全长IgG的亲和力和活性非常有效。