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- W1013032156 abstract "Objective To construct and screen specific mouse Fab phage antibody library against EHEC O157:H7 Stx2. Methods BALB/c mice were immunized with inactivated Stx2 of EHEC O157:H7, and splenic lymphocytes were isolated for extraction of total RNA from which antibody light chain κ gene and heavy chain Fd gene were amplified by RT-PCR, identified by restriction analysis and inserted into phagemid pComb3X successively. The constructed recombinant vector was transformed to E.coli XL1Blue by electroporation, based on which the specific Fab phage antibody library against Stx2 of EHEC O157:H7 was constructed by superinfection with helper phage M13K07. The specific Fab antibody against Stx2 of EHEC O157:H7 was screened using purified Stx2 as antigen. The binding activity of phage antibody to toxin antigen was determined by Western blot, and the obtained positive clones were sequenced. Results The Fab phage antibody library with a capacity of 1.56×10^7 was constructed, and three strong positive clones recognizing Stx2 specifically were screened, of which two reacted with Stx2A1, and the other one with Stx2B. The homologies of amino acid sequences of variable heavy (VH) and light (VL) chain domains were 98.5% and 99.6% respectively to those of murine Ig reported in GenBank. Conclusion The specific mouse Fab phage antibody library against EHEC O157:H7 Stx2 was successfully constructed, which laid a foundation of preparation of therapeutic humanized antibody against EHEC O157:H7 Stx2." @default.
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- W1013032156 date "2009-02-20" @default.
- W1013032156 modified "2023-09-23" @default.
- W1013032156 title "鼠源性抗EHEC O157:H7 Stx2噬菌体Fab抗体库的构建及筛选" @default.
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