Factors that affect protein adsorption on nanostructured titania films. A novel spectroelectrochemical application to sensing

Nanoporous, thick (8 mum) films of titania (TiO2) were prepared and used for the immobilization of proteins. A detailed study has been made into the factors influencing protein adsorption on TiO2. Among these, we investigated pH, ionic strength of solution, protein surface charge, protein size, and immobilization time. Protein immobilization is found to be remarkably stable, attributed to secondary binding processes occurring after the initial immobilization. We also investigated the electrochemical properties of these films using cyclic voltammetry and spectroelectrochemistry and found that not only was direct reduction of the Fe-III-heme to Fe-II-heme of both cytochrome-c and hemoglobin possible but that all the protein in the film is electroactive. We further demonstrate the use of a hemoglobin/TiO2 film as an aerobic sensor for nitric oxide. Optical sensing is demonstrated, with a limit of detection of 1 muM.