Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology · 2010
[Expression and neutralizing activity analysis of humanized single-chain antibody against anthrax protective antigen].
Аннотация на английскомСтатьяПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод в очереди
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Li Bing, Zhang Jun, Xu Jun-jie, Liu Shu-ling, Fu Ling, Li Jian-min, Chen Wei
Аннотация
AIM: In order to reduce the human anti-mouse antibody (HAMA) response, anti-anthrax protective antigen scFv-5E1 has been humanized.
METHODS: The authors have constructed a "humanized" antibody fragments by grafting the complementarity-determining regions (CDRs) of the murine anti-PA scFv-5E1 to human framework regions which showed maximal homology to the scFv-5E1 sequence. The humanized VH-5E1 and VL-5E1 DNA fragments were then joined by fusion PCR. The expression vectors named pET-hscFv-5E1 was constructed by cloning the humanized 5E1 scFv gene into the Nde I/EcoR I site. The transformed E.coli BL21(DE3) cells were propagated and induced by IPTG.
RESULTS: Expression product was found as inclusion body with expected size by SDS-PAGE analysis. Soulable scFv showed binding ability to the protective antigen by ELISA analysis. The purified scFv showed good neutralizing activity in a cell model.
CONCLUSION: The humanized scFv exhibited good binding and neutralizing activity. It lays a foundation for the development of therapeutic whole molecular antibody.
METHODS: The authors have constructed a "humanized" antibody fragments by grafting the complementarity-determining regions (CDRs) of the murine anti-PA scFv-5E1 to human framework regions which showed maximal homology to the scFv-5E1 sequence. The humanized VH-5E1 and VL-5E1 DNA fragments were then joined by fusion PCR. The expression vectors named pET-hscFv-5E1 was constructed by cloning the humanized 5E1 scFv gene into the Nde I/EcoR I site. The transformed E.coli BL21(DE3) cells were propagated and induced by IPTG.
RESULTS: Expression product was found as inclusion body with expected size by SDS-PAGE analysis. Soulable scFv showed binding ability to the protective antigen by ELISA analysis. The purified scFv showed good neutralizing activity in a cell model.
CONCLUSION: The humanized scFv exhibited good binding and neutralizing activity. It lays a foundation for the development of therapeutic whole molecular antibody.