Expression and purification of a human anti-cyclin D1 single-chain variable fragment antibody AD5 and its characterization

Abstract

CyclinD1 plays an important role in cell cycle progression. Increasing evidence indicates that cyclinD1 is overexpressed in the majority of tumor cells and has become a potential target for tumor therapy. However, little research has been done on the specific inhibition of cyclinD1 for cancer therapy. With the rapid development of the phage display antibody library technique, single-chain variable fragment (scFv) antibodies have emerged, which have tremendous application prospects in cancer therapy and diagonosis. In this study, a human scFv binding specifically to cyclinD1 (AD5) that was derived from a human semi-synthetic scFv phage library was expressed in the soluble form in Escherichiacoli (E.coli) HB2151 cells. To characterize AD5, soluble AD5 was purified successfully through ammonium sulfate precipitation and affinity chromatography from the culture supernatant of AD5/HB2151. ELISA assay revealed that purified soluble AD5 could specifically bind to human recombinant cyclinD1 with approximately (1.19±0.056)x107M-1 affinity constant and showed approximately 52% competitive inhibition with the anti-cyclinD1 polyclonal antibody for binding to cyclinD1 invitro. These results suggest that the scFv antibody against cyclinD1 may be a novel potential tool for targeting cyclinD1 in cancer therapy and diagnosis.