Oriented Self-Assembling Protein Monolayers for Antibody Capture on Gold Surfaces

摘要

The immunoassay is a powerful tool in diagnostics; antibody technology provides exquisitely specific capture of biological markers for disease. In the last 15 years there has been a drive to transfer the benefits of the immunoassay onto the surfaces of advanced electronic biosensors. Traditional methods of antibody immobilisation such as adsorption and chemical coupling have some disadvantages that are magnified in the arena of ultrasensitive miniaturised electronic detection of antibody-antigen interaction (Table 1). In order to address some of these problems we have developed a method for creating oriented, stable, self-assembled monolayers of protein using engineered bacterial outer membrane proteins (omp) as scaffolds for fusion proteins [1,2]. The core technology requires the fusion of a protein of interest to a scaffold protein with self assembling properties. The fusion protein is assembled in a monolayer by a simple 'apply-and-wash' process. Gaps between the proteins are filled in with filler molecules such as PEG-thioalkanes, leaving only the protein of interest exposed (Figure 1). This method overcomes many of the problems associated with traditional methods of protein application to surfaces. The scaffold protein is highly stable with a melting temperature of 88°C. It is resistant to protease digestion and remains intact in SDS and extremes of pH. This basic technology was used to create a set of proteins for antibody immobilisation.