Direct Immobilization of Gold-Binding Antibody Fragments for Immunosensor Applications

摘要

A novel method that enables antibody fragments to benimmobilized on a sensor substrate with a high bindingncapability using molecular recognition has been devel-noped. Using genetic engineering, we fabricated bispecificnrecombinant antibody fragments, which consist of twonkinds of antibody fragments: a gold antibody fragment andna target molecule antibody fragment. Surface plasmonnresonance (SPR) analysis indicated that these gold-nbinding bispecific antibody fragments bind directly to thengold substrate with high affinity (KD ∼ 10 9 M). Aboutn70% of the bispecific antibody fragments immobilizednon the gold substrate retained their target protein-nbinding efficiency. The Sips isotherm was used tonassess the heterogeneity in antibody affinity for thenbispecific antibody fragments. The results showed thatnthe immobilized bispecific antibody fragments exhib-nited an increased homogeneity of affinity (KD) to targetnmolecules when compared with monospecific antibodynfragments immobilized by conventional methods. Thenuse of bispecific antibody fragments to directly im-nmobilize antibody fragments on a solid-phase substratenoffers a useful platform for immunosensor applications.