[Construction of full-length human bladder cancer-specific antibody libraries based on mammalian display technology].

Objective To construct full-length human bladder cancer-specific antibody libraries for efficient display of full-length antibodies on the surface of mammalian cells.Methods The total RNA was isolated from peripheral blood mononuclear cells from patients with bladder cancer.The repertoires of IgGl heavy chain variable region(VH) and Kappa light chain were amplified by RT-PCR using specific primers.The antibody genes were inserted into the vector pDGB-HC-TM to construct the bladder-cancer-specific antibody libraries of heavy chains and light chains.Ten clones from each library were randomly picked for gene sequencing and transient transfection into FCHO cells to analyze antibody display on mammalian cell surface by flow cytometry after staining with corresponding fluorescent labeled antibodies.Results The libraries of bladder-cancer-specific antibody heavy chain(IgGl) and light chain(LCκ) were successfully constructed.Seven out of the 10 clones randomly selected from the heavy chain library and 9 out of the 10 clones from the light chain library showed correct open reading frame,coding for 7 unique VH and 9 unique LCκ.The combinatory library size reached 3.32×10~(11)[(1.7×10~6×70%)×(3.1×10~5×90%)].Conclusion We have successfully constructed a full-length human bladder-cancer-specific antibody library with a combinatory diversity of 3.32×10~(11) based on mammalian display technology,which can be used for screening monoclonal antibodies against bladder-cancer-associated antigens.