[Construction of chimeric anti-c-Met antibody random mutagenesis libraries using mammalian cell surface display]

Objective To construct a random mutagenesis library of 3E1D7, a chimerical antibody against c-mesenchymal epithelial transition factor (c-Met), using mammalian cell surface display. Methods Antibody genes with randomly mutated complementarity-determining region 3 (CDR3) part were inserted into the mammalian expression plasmid pSZI-CD to construct the random mutagenesis library using double enzyme digestion. Reconstructed plasmids were then cloned into CHO cells by transfection. The expression level of antibodies on the surface of CHO cells was checked by C6 PLUS flow cytometry. Results 3E1D7 random mutagenesis library was successfully constructed with a volume of 5.52×10⁶ in diversity on gene level. Sequence analysis showed that all 20 clones randomly picked from the library coded for 20 different mutated amino acid sequences in open reading frames. After transfection, the expression of full-length antibodies on CHO cell surfaces could be detected by flow cytometry. Conclusion A random mutagenesis library of a certain anti-c-Met antibody has been successfully constructed with an exhibitable diversity of 5.52×10⁶, which would be a useful platform for further screening of therapeutic antibodies.