Antibody immobilization technique using protein film for high stability and orientation control of the immobilized antibody

Abstract

The development of antibody immobilization techniques is essential for creating antibody-based biomaterials. Although numerous methods for antibody immobilization have been demonstrated, low stability and disordered orientation of the immobilized antibody remain an important problem. In this work, an original antibody immobilization technique using a protein film, which achieved a high stability and orientation control of the immobilized antibody, has been described. In this method, an antibody-immobilized albumin film was prepared by adding the cross-linked albumin solution to the substrate, where antibodies were attached in uniform orientation, followed by subsequent drying, and detaching the formed film from the substrate by heating at 120 °C in a dry state. Antibodies in the film showed high antigen-binding capacity, at a level comparable to the oriented immobilized antibody using protein G. The stability of antibodies in the film was found to be significantly high; their antigen-binding capacity was completely retained even after storage at 40 °C in a dry state for one month. Thus, this approach provides useful information to immobilize the antibody on solid surfaces while controlling its orientation and increasing its stability.