A Humanized Single Chain Fragment Variable Constructed to Use in Breast Cancer Immunotherapy

Abstract

The mouse B cell hybridoma line C3A8 was the starting material to generate monoclonal antibody to react specifically toward epidermal growth factor receptors (EGFR) of breast cancer cell line (MCF‐7). Phage display technologies were applied to produce molecular form of those IgG antibodies. The integrated cloning, screening, and phage display system allowed to use extended primers, which were incorporate the entire mouse VH and VL and their linker sequences collected in the Kabat database. The product was called single‐chain fragment variable (scFv) antibodies. The best candidate scFv sequences, based on preliminary enzyme‐linked immunosorbent assay (ELISA) screening data gave 100% identical sequences in genbank data base, while a band of 68kD was appeared in nitrocellose membrane in western blot test had used to characterize the specific reaction of scFv towards MCF‐7 breast cancer cells. Further, immuno‐fluorescence test clearly proved that the scFv recognized the MCF‐7 antigen epitopes which is localized in MCF‐7 nuclear. Also, 99% of the cells numbers were bind to scFv protein as measured by flow cytometry analysis. Herein the recombinant antibody technology is a rapid and effective approach to the development of the next generation of breast cancer diagnosis and immune‐therapy.