Abstract
Before any novel biochemical device is fabricated, one needs to know that the technology and methodology being used are the most efficient. In the case of immobilised proteins, one needs to know that the method used will not harm the protein and that the conjugation technique used facilitates the highest binding efficiencies for the protein in question. Three methods from literature (non-specific binding, non-covalent binding and covalent binding) were used to conjugate bovine serum albumin (BSA) to single-walled carbon nanotubes (SWNTs). Efforts to use simple methods for protein concentration determination were hampered by the strong absorbance and interference from the nanotubes themselves. FITC tagged BSA was then used to investigate the conjugation methods. This technique was also subject to interference from the nanotubes when they were present. However, the subtraction of the amount of BSA removed from the nanotubes during washing was found to be accurate enough to compare the different conjugation techniques used. This showed that non-specific binding was the best method for conjugation, with approximately 79% of the original BSA remaining on the nanotubes after eight washes. The non-covalent and covalent method showed efficiencies of 54 and 60% respectively. A novel method for protein binding was investigated. This involved using a short DNA oligonucleotide as a tether for binding. The protein concentration was investigated as before using fluorescent BSA. This showed that approximately 88% of the BSA originally added remained on the nanotubes after washing. This value is higher than the other methods investigated and therefore this was the best technique. In addition to BSA conjugation, the enzyme urease was attached using non-specific binding and non-covalent conjugation. An attempt was made to determine the amount of urease bound through an assessment of its catalytic activity. This investigation was unable to accurately determine the amount of urease present, but did show there to be a difference in the amount of urease bound to SWNTs using either non-specific binding and non-covalent binding. The investigation of the best method for protein immobilisation in this thesis has shown that the novel DNA mediated conjugation is the most efficient, closely followed by the simplest method, non-specific binding. Both of these methods can allow the protein to remain structurally intact, an important factor to consider in the creation of novel devices.