Selection of scFvs specific for the HepG2 cell line using ribosome display

Journal of biosciences

Feng-Yun Zhou¹  Wen-Xiu Li¹  Juan Fen¹  Hong-Yun Liu¹  Chuan-Jing Wei¹  Wei-Ping Mao1¹  Lei Zhou¹ 

[1] Jiangsu Province Key Laboratory of Molecular and Medical Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210046, P R China$$

关键词: Hepatocarcinoma;    ribosome display;    single-chain antibody;   

DOI  :  

来源: Indian Academy of Sciences

【 摘 要 】

The aim of this study was to construct a ribosome display library of single chain variable fragments (scFvs) associated with hepatocarcinoma and screen such a library for hepatocarcinoma-binding scFvs. mRNA was isolated from the spleens of mice immunized with hepatocellular carcinoma cell line HepG2. Heavy and k chain genes (VH and k) were amplified separately by RT-PCR, and an anti-HepG2 VH/k chain ribosome display library was constructed by assembling VH and k into the VH/k chain with a specially constructed linker by SOE-PCR. The VH/k chain library was transcribed and translated in vitro using a rabbit reticulocyte lysate system. In order to isolate specific scFvs, recognizing HepG2 negative selection on a normal hepatocyte line WRL-68 was carried out before three rounds of positive selection on HepG2. After three rounds of panning, cell enzyme-linked immunosorbent assay (ELISA) showed that one of the scFvs had high affinity for the HepG2 cell and lower affinity for the WRL-68 cell. In this study, we successfully constructed a native ribosome display library. Such a library would prove useful for direct intact cell panning using ribosome display technology. The selected scFv had a potential value for hepatocarcinoma treatment.

【 授权许可】

Unknown