Immobilizing antibodies on piezoelectric quartz crystal for diagnostic applications

The development of a stable and sensitive microgravimetric immunobiosensor requires stable and reproducible immobilization method for binding antibodies to the biosensing surface. The piezoelectric quartz crystal surface is coated with a specific antibody to which the specific antigen is adsorbed resulting in change of its resonance frequency. In the present study, silanization using 3-aminopropylthethoxysilane (3-APTES) has been used for activating the carrier surface. Protein A is covalently attached to the surface causing the oriented immobilization of human immunoglobulin G (human IgG) leaving its antigen binding sites free to interact with the specific antigen. The optimization of the immobilization was performed by inverted fluorescence microscopy, fluorescence spectroscopy, enzyme labeling and atomic force microscopy (AFM). A highly uniform monolayer distribution of protein A and the subsequent oriented immobilization of human IgG were shown in our studies.