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Selection of scFvs specific for the HepG2 cell line using ribosome display.
Zhou, Lei; Mao, Wei-Ping; Fen, Juan; Liu, Hong-Yun; Wei, Chuan-Jing; Li, Wen-Xiu; Zhou, Feng-Yun.
Afiliación
  • Zhou L; Jiangsu Province Key Laboratory of Molecular and Medical Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210046, P R China.
J Biosci ; 34(2): 221-6, 2009 Jun.
Article en En | MEDLINE | ID: mdl-19550038
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.
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Colección: 01-internacional Banco de datos: MEDLINE Asunto principal: Ribosomas / Cadenas Pesadas de Inmunoglobulina / Neoplasias Hepáticas Tipo de estudio: Prognostic_studies Límite: Animals / Female / Humans Idioma: En Revista: J Biosci Año: 2009 Tipo del documento: Article
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Colección: 01-internacional Banco de datos: MEDLINE Asunto principal: Ribosomas / Cadenas Pesadas de Inmunoglobulina / Neoplasias Hepáticas Tipo de estudio: Prognostic_studies Límite: Animals / Female / Humans Idioma: En Revista: J Biosci Año: 2009 Tipo del documento: Article