单克隆抗体
抗体
噬菌体
表位
噬菌体展示
免疫
抗原
生物
免疫系统
分子生物学
抗体库
病毒学
cDNA文库
杂交瘤技术
互补DNA
免疫学
噬菌体
大肠杆菌
基因
遗传学
作者
Thi Thu Ha Nguyen,Jong Seo Lee,Hyunbo Shim
出处
期刊:Methods in molecular biology
日期:2023-01-01
卷期号:: 93-106
标识
DOI:10.1007/978-1-0716-3381-6_6
摘要
Rabbits have distinct advantages over mice as a source of target-specific antibodies. They produce higher affinity antibodies than mice and may elicit strong immune response against antigens or epitopes that are poorly immunogenic or tolerated in mice. However, a great majority of currently available monoclonal antibodies are of murine origin because of the wider availability of murine fusion partner cell lines and well-established tools and protocols for fusion and cloning of mouse hybridoma. Phage display selection of antibody libraries is an alternative method to hybridoma technology for the generation of target-specific monoclonal antibodies. High-affinity monoclonal antibodies from non-murine species can readily be obtained by constructing immune antibody libraries from B cells of the immunized animal and screening the library by phage display. In this article, we describe the construction of a rabbit immune Fab library for the facile isolation of rabbit monoclonal antibodies. After immunization, B-cell cDNA is obtained from the spleen of the animal, from which antibody variable domain repertoires are amplified and assembled into a Fab repertoire by PCR. The Fab genes are then cloned into a phagemid vector and transformed to E. coli, from which a phage-displayed immune Fab library is rescued. Such a library can be biopanned against the immunization antigen for rapid identification of high-affinity, target-specific rabbit monoclonal antibodies.
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