earticle

영어

Single-chain Fv antibodies that VH and VL domains of whole antibodies (whole Abs) are genetifally linked via a flexible linker (G4S)3, have been expected as a next generation of ligand molecular recognition agent in clinical diagnosis. ScFvs can be efficiently produced in an insoluble fraction of recombinant E. coli cells with much lower production cost than the conventional whole Ab, while immobilization method of scFv with high density, activity and uniform orientation is requisite to increase antigen-binding activity of scFv on the solid support to great potential extent. Recently, we succesfully identified the affinity peptide tag which possessed strong binding affintiy against the surface of hydrophilic polystyrene (phi-PS) support for immunodiagnostics. By genetic fusion or chemical conjugation of PS-tag with target proteins such as enzyme as well as antibody, they were able to be immobilized with maintaining much higher density as well as remaining activity due to site-specific attachment of PS-tag onto the surface of phi-PS plate. Especially, it was revealed that use of PS-tag-fused scFvs (scFv-PS) as a ligand antibody resulted in enhancement of sentivity 10 times higher than that of conventional whole Ab. Furthermore, a solid-phase refolding method that refolding and immobilization processes of scFv-PS were simultaniously performed, was successfully developed. Consequently, production cost of scFv-immobilized immunodiagnostic support could become one-tenth or further lower than that of whole Ab-immobilized support. This method was applicable to a variety of antibody species. Thus, the immobilization method for PS-tag-fused scFv developed in this stugy will be considerably useful for production of high-performance antibodyimmobilized diagnostic support.