Kim Sang Jick, Jang Myeong Hee, Ahn Hyun Joo, Kim Jin Hong, Lim Ji Hye, Ryu Chun Jeih, Lim Nam-Kyu, Kim Keun-Soo, Park Mi-Ju, Park Insoo, Hong Hyo Jeong
Therapeutic Antibody Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejon 305-333, South Korea.
J Immunol Methods. 2008 Jan 1;329(1-2):176-83. doi: 10.1016/j.jim.2007.10.009. Epub 2007 Nov 9.
In a previous study, we generated a murine hepatitis B virus (HBV)-neutralizing monoclonal antibody (mAb), KR127, that binds to an epitope (amino acids 37-45, NSNNPDWDF) of the preS1 antigen. Furthermore, an epitope tag, S1 (NANNPDWDF), was developed for protein tagging. The aim of the present study was to develop a high-affinity antibody to the same preS1 epitope. Mice were immunized with the N-terminal domain of human thrombopoietin fused to the S1 tag (nTPO-S1), and a phage-displayed chimeric Fab library was constructed and screened by panning against nTPO-S1. A high-affinity antibody (3-34) was selected that binds to the preS1 antigen. The IgG molecules of 3-34 showed approximately nine-fold higher affinity (K(D) 1.2 nM) for preS1 compared with KR127 (K(D) 10.4 nM), competed with KR127 for binding to the epitope, and bound to HBV particles. This study provides a simple and efficient way to develop a high-affinity antibody to a defined epitope by phage display of an immune antibody library.
在先前的一项研究中，我们制备了一种小鼠乙型肝炎病毒（HBV）中和单克隆抗体（mAb）KR127，它可与前S1抗原的一个表位（氨基酸37 - 45，NSNNPDWDF）结合。此外，还开发了一种用于蛋白质标记的表位标签S1（NANNPDWDF）。本研究的目的是开发一种针对同一前S1表位的高亲和力抗体。用与S1标签融合的人血小板生成素N端结构域（nTPO - S1）免疫小鼠，构建噬菌体展示嵌合Fab文库，并通过对nTPO - S1进行淘选来筛选该文库。筛选出一种与前S1抗原结合的高亲和力抗体（3 - 34）。与KR127（解离常数K(D)为10.4 nM）相比，3 - 34的IgG分子对前S1的亲和力高约9倍（K(D)为1.2 nM），能与KR127竞争结合该表位，并且能与HBV颗粒结合。本研究提供了一种通过免疫抗体文库的噬菌体展示来开发针对特定表位的高亲和力抗体的简单有效方法。
