Molecular Engineering of the Biosensor Interface.

The research described in this thesis concerns the investigation of technologies for the molecular engineering of the biosensor interface. Two avenues of investigation have been explored: the use of polymer matrices to modify the properties and functions of optical biosensor interfaces and the conjugation of photochromic dyes to protein systems to achieve photomodulation of protein function for biosensor applications. A comparison of the industry standard polymer, carboxymethyl dextran (CMD) was made against carboxymethyl cellulose and mixed systems, including a novel synthetic polymer, carboxylated polynoxylin. While CMD was found to provide the highest surface loading of protein, mixed polymers demonstrated the ability to allow prediction of surface loading, and showed features such as improved resistance to biological degradation. A novel method of depositing interfaces was investigated, allied to a study of liquid handling methods. A system was developed that allowed a printed heterogeneous array to be produced which showed preferential binding of specific analytes to defined areas of the sensor, whilst the other printed arrays retaining a high degree of non-specific interaction. The use of photochromic dyes to modulate protein function was applied to glucose oxidase and horseradish peroxidase. From the initial results, a hypothesis regarding the mechanism of photomodulation and its effect concerning the molecular weight of the conjugated protein was proposed. This was examined by the photomodulation of members of the peroxidase super family, antibodies and Fab fragments. From these results, the hypothesis was proved to be correct but incomplete, and was modified to include the disruption of the hydration shell around the protein caused by photochromic switching. Further research directly related to these experiments, and in novel fields of investigation have been proposed.