[Construction of human full-length renal cell carcinoma patient-specific antibody library by mammalian cell surface display].
摘要
OBJECTIVE: To construct human renal cell carcinoma patient-specific full-length antibody library using mammalian cell surface display technique. METHODS: Peripheral blood mononuclear cells (PBMC) were isolated from patients with renal cell carcinoma. The repertoires of kappa light chain (LCkappa) and heavy chain variable region (VH) of antibody were amplified by RT-PCR. The LCkappa and VH libraries were inserted into the vector pDGB-HC-TM separately, and the ligated libraries were transformed into competent E.coli TOPO10 to construct the renal cell carcinoma patient-specific antibody heavy and light chain libraries. 293T cells were co-transfected with the libraries and the full-length human antibodies expressed on the surface of 293T cells were analyzed by flow cytometry. RESULTS: The libraries of renal cell carcinoma-specific antibody kappa light chain (LCkappa) and heavy chain (IgG1) were constructed. The expression of the full-length human antibodies on the surface of 293T cell was confirmed by flow cytometry. The libraries showed an expressible combinatory diversity of 7.5x10(10). CONCLUSION: The expressible antibody library provides a useful platform for screening of renal cell carcinoma-specific antibodies.