earticle

영어

For the realization of immunoaffinity (IA) biosensors, antibodies have been usually immobilized on the metal surface of transducers as a molecular recognition layer. As the antigen-binding sites of antibodies are located at Fab region, the sensitivity of the IA biosensors is influenced on the orientation of antibodies. In this work, Z domain of protein A with IgG-binding activity was expressed on the outer membrane of E.coli as a fusion protein of AIDA-1 by using Autodisplay method1-2). The outer membrane of E.coli with Z-domain was isolated to be colloid particle with a diameter of 100 nm according to ref.3). Prepared E.coli outer membrane was coated on the gold
surface and the IgG-binding activity was tested by the binding assay with fluorescence labeled IgG. In comparison to the intact outer membrane, the outer membrane with Z-domain showed significant amount of IgG binding activity. The outer membrane of E.coli with Zdomain was treated to the gold surface of SPR biosensor. After antihIgG antibodies were immobilized to the Z-domain, hIgG antibodies were measured as a target analyte. The LOD of SPR biosensor with
the outer membrane of E.coli with Z-domain and intact E.coli outer membrane was estimated to be 1.6 ng/ml, 316 ng/ml, respectively. This result shows that the outer membrane of E.coli with Z-domain could increased the sensitivity of SPR biosensor through the orientation control of antibodies at the molecular recognition layer.