Acoustic coupling of transverse waves as a mechanism for the label-free detection of biomolecular interactions.
Published: 2004
Language: eng
Number of pages: 185
Description: This thesis presents the application of the Thickness Shear Mode (TSM) Acoustic Wave Sensor to the study of interactions between biological molecules and between biological molecules and small molecules.The final biological interaction is that between the enzyme Taq DNA Polymerase and a DNA sequence. This is a step in the Polymerase Chain Reaction which is traditionally performed in solution but is being developed here as a surface based technique by immobilizing the Taq enzyme to the surface of the TSM sensor in order to be able to monitor the PCR steps acoustically.This thesis shows that the TSM sensor can be used to detect binding interactions and monitor conformational changes of the HSA.This has been further investigated by applying the principle that molecules containing triple bonds will crosslink in the presence of UV light and that this crosslinking essentially involves no mass change. Monolayers that contain triple bonds and those that do not were exposed to UV light while in the TSM sensor format. The resulting surfaces were further characterized by Secondary Ion Mass Spectrometry.Other interactions that have been studied are those between HSA and the drug cisplatin and HSA and its monoclonal antibody.Four linking systems for the covalent attachment of the biological molecules to the surface of the gold electrode of the sensor were employed, namely thioctic acid, cysteamine, 11-mercaptoundecanoic acid and an N-hydroxysuccinimide ester. All attachment protocols involve the ability of the thiols to form gold-sulphur bonds at the metal surface and be chemically activated at the distal end for covalent attachment.The acoustic parameters associated with the binding interaction of immobilized Human Serum Albumin (HSA) and the drug warfarin where measured on line in real time. Depending on the linking monolayer, different directional combinations of series resonant frequency and motional resistance are observed for the binding of warfarin. These responses are ascribed to acoustic coupling phenomena.It is known that HSA will undergo conformational changes on the binding of warfarin and in the pH is altered. The TSM sensor shows that the HSA and the monolayer itself are affected by pH.
Notes: Adviser:  Michael Thompson.

Thesis (Ph.D.)--University of Toronto, 2004.

Electronic version licensed for access by U. of T. users.

Source: Dissertation Abstracts International, Volume: 65-05, Section: B, page: 2390.