Thesis (Ph. D.)--University of Hong Kong, 2007. / Title proper from title frame. Also available in printed format.

The aim of this work is to better understand the factors and mechanisms leading to the supramolecular organization of collagen layers adsorbed on polymers. Native collagen adsorption on polystyrene (PS) and plasma-oxidized polystyrene (PSox) substrates revealed that the adsorbed layer consists in two parts: a dense and thin sheet (~ 10 nm) in which fibrils are formed, as revealed by atomic force microscopy, and an overlying thick layer (~ 200 nm) which contains protruding molecules, as revealed by quartz crystal microbalance with dissipation monitoring. The protruding molecules are in low density but modify noticeably the local viscosity. Faster and enhanced fibril formation takes place on hydrophobic compared to hydrophilic substrate. As a result of drastic thermal denaturation, the ability of collagen to assemble into fibrils is lost and the number of protruding molecules responsible for higher viscosity is reduced. Radiochemical measurements showed that collagen molecules are more easily displaced when adsorbed on a hydrophobic substrate compared to a hydrophilic substrate. This may explain why fibril formation occurs more readily on the more hydrophobic substrate, but is in contrast with higher surface affinity. The possible explanation of this paradox by the quick formation of a dense layer of collagen molecules having a smaller number of contact points with a very hydrophobic surface could not be demonstrated by a comparison of adsorption procedures. Comparing different collagen sources revealed various modes of aggregation with different characteristics regarding size and order (large fibers in solution, smaller fibrils to featureless underneath layer in the adsorbed phase). Moreover, collagen aggregation in the solution is a process competing with adsorption: more aggregated solutions behave like less concentrated solutions regarding the adsorbed amount and fibril formation in the adsorbed phase. It must be emphasized that interpretation of the QCM-D data, which is based on fitting physical quantities according to a model, has to be performed very carefully, and requires the examination of the sensitivity of the fitted data to the fitting parameters.

This thesis uses surface sensitive tools to characterize the effect of a solid surface on immobilized biomacromolecules. This includes understanding how the surface can change the affinity of these macromolecules to small molecules compared to bulk studies. Two classes of immobilized biomacromolecules, the supported lipid bilayer (SLB) and the Lac repressor protein (LacI), are characterized using microcantilever sensors and quartz crystal microbalance with dissipation (QCM-D). The first part of this thesis reports the use of microcantilever beams, an ultrasensitive sensor for measuring the surface free energy changes on a substrate induced by molecular adsorptions, to probe the interaction between a solid surface and a phospholipid bilayer. This sensing method integrates two well-developed techniques: solid-supported lipid bilayers (SLBs) and the microcantilever (MC) sensors. Studying the adsorption free energy of lipid bilayers on a solid surface allows better characterizing of the formation and stability of SLBs. Microcantilever converts the Gibbs free energy change taking place on its surface into a mechanical deformation. As molecules physisorb or chemisorb onto the surface of the microcantilevers, the microcantilevers bend, either due to induced compressive or tensile stresses, which result from the surface free energy change. By monitoring the deflection values of the microcantilevers, the real-time surface free energy change during the SLB formation can be detected. This thesis has led to the development of a novel biosensor--lipid membrane coated microcantilevers--to detect the adsorption, insertion, aggregation and solubilizing effect of membrane-active substances, such as surfactants and peptides, on the phospholipid membranes. To better characterize the surface free energy, SLBs doped with charged lipids or cholesterol are shown to alter the surface free energy. We can predict this change in surface free energy using a thermodynamic model. Application of this membrane-coated cantilever is put into use for detecting how amphiphilic molecules interact with SLBs, as well as for probing the abrupt conformational change of SLBs during a temperature induced phase-transition. This study systematically demonstrates various usage aspects of microcantilever to characterize the SLBs, and how this technique may advance the biophysical knowledge of the lipid membrane, one of the essential building blocks of life. The second part of this thesis reports the use of both microcantilever sensors and QCM-D to measure the adsorption free energy and mass of a model protein, the Lac repressor (LacI), and compare how a modified T334C mutant that includes a cysteine group to orient the protein on the gold surface through a covalent sulfur bonds retains its binding capabilities over that of wild type LacI. The main challenge of this work is to unravel how the adsorption of biomacromolecules at the solid/liquid interface leads to surface free energy changes and ultimately changes the stress of the underlying solid surface (the cantilever). The uses of microcantilever sensors and QCM to probe the interactions that take place on SLBs and surface-bound proteins have the advantage of being a sensitive, real-time, and label-free technique.

The interest in tiny wireless applications raises the demand for an integrated reference oscillator with the same performance as the macroscopic quartz crystal reference oscillators. The main challenge of the thesis is to prove that it is possible to build a MEMS based oscillator that approaches the accuracy level of existing quartz crystal oscillators. The MEMS resonator samples which Philips provides are measured and an equivalent electrical model is designed for them. This model is used in the simulations of the Pierce oscillator and the transresistance amplifier oscillator that are evaluated in this thesis. Finally the Pierce oscillator is implemented in the A BCD2 process and manufactured at Philips Semiconductors in Nijmegen, The Netherlands. A test board, for measuring the Pierce oscillator together with a MEMS resonator or a quartz crystal resonator, is built. The Pierce oscillator is then measured with a quartz crystal resonator. In order to simulate the higher series resistance of the MEMS resonators a resistor is put in series with the quartz crystal. The Pierce oscillator is working with a series resistance of 1 kΩ. With higher series resistance the Pierce oscillator stops working. In circuit simulations the Pierce oscillator is working with a series resistance of about 5 kΩ in the MEMS resonator model. To be sure whether the Pierce oscillator has enough gain for the MEMS resonators, it needs to be measured with them. Temperature variations in the MEMS resonators need to be handled and the phase noise performance of the oscillator must be improved, in order for the MEMS based reference oscillator to be a successful replacement for the quartz crystal reference oscillator.

The primary objective of this study was to develop a sensitive and cost-effective sorption system to analyze adsorption and diffusion of different gases on micro porous materials and nanotubes. A high pressure Quartz Crystal Microbalance (QCM) based adsorption apparatus for single-component gas was developed. A QCM is an acoustic-wave resonator in which the acoustic wave propagates through the crystal. Therefore, it is highly responsive to addition or removal of small amounts of mass adsorbed or deposited on the surface of the crystal. This mass sensitivity makes the QCM an ideal tool for the study of gas adsorption. The QCM-based adsorption apparatus is advantageous over the commercialized none-gravimetric and gravimetric equipment in a way that it is low-cost, highly sensitive and accurate for mass sorption applications, satisfactorily stable in a controlled environment, and can be used for thin films. The high pressure apparatus was calibrated using Matrimid 5218, whose thermodynamic properties and adsorption parameters are known. The Matrimid was spin-coated onto a 14 mm-diameter QCM, and sorption equilibrium data for were obtained for CO₂ gas at 25, 30, 48, and 52 ºC and partial pressure range between 0 to 4 bar. In order to compare the experimental data with available literature data, the experimental data was fitted into a dual-mode adsorption model. The model results from Henry's law and a Langmuir mechanism. Comparison of the experimental adsorption isotherm of Matrimide for CO₂ gas with literature data showed reasonable agreement between the experimental and literature data. In this study, the adsorption parameters of aluminosilicate nanotubes are observed. Aluminosilicate nanotubes are ideal materials for chemical sensing, molecule separation, and gas storage; hence, there is a need for adsorption and diffusion data on this material. The adsorption of CO₂, N₂, and CH₄ gases on aluminosilicate nanotubes samples has been studied in the temperature range of 20° to 120° Celsius and pressure range of 0 to 8 bar. The experimental results yield the CO₂ and N₂ heat of adsorptions of -32.9 and -28.1 kJ/mol respectively.

The contamination of drinking water by microbial pathogens is recognized as one of the most pressing water supply problems of our day. To minimize the impact of pathogens and parasites on the environment and public health, accurate methods are needed to evaluate their presence and concentration. Although various techniques exist to detect certain pathogens in water (e.g., immunofluorescence or PCR techniques), these are time- and labor-intensive. A direct, real-time method for detection and quantification of target organisms would thus be very useful for rapid diagnosis of water safety. A quartz crystal microbalance with dissipation monitoring (QCM-D) based biosensor for detection of waterborne pathogens (i.e., Escherichia coli O157:H7) was developed. The detection platform is based on the immobilization of affinity purified antibodies onto gold coated QCM-D quartz crystals via a cysteamine self-assembled monolayer. The results show that the optimal sensor response is the initial slope of the dissipation shift. A highly log-log linear response is obtained for detection of E. coli O157:H7 over a broad range of cell concentration from 3 x 105 to 1 x 109 cells/mL. The prepared biosensor also exhibits a log-log linear working range from 107 to 109 cells/mL for E. coli K12 D21, a non-pathogenic model organism. The biosensor also shows satisfactory selectivity using Bacillus subtilis . To our knowledge, this is the first study demonstrating the use of the slope of the dissipation shift as a sensor response when using QCM-D technology. / Keywords: Biosensor, QCM-D, E. coli O157:H7, polyc1onal antibodies, dissipation slope, cysteamine, self-assembled monolayer

Ice-associating proteins (IAPs) are proteins that interact directly with ice crystals, either by offering a site for nucleation, i.e. ice nucleating proteins (INPs), or by binding to nascent crystals to prevent addition of more water molecules, i.e. antifreeze proteins (AFPs). AFPs have been found to inhibit the formation of clathrate-hydrates, ice-like crystalline solids composed of water-encaged guest molecules. Study of AFP-hydrate interaction is leading to a greater understanding of AFP adsorption and of the mechanism behind the “memory effect” in hydrates, wherein previously frozen crystals reform more quickly after a brief melt. AFP is currently the only known memory inhibitor. Such a low-dosage hydrate inhibitor (LDHI) is of great interest to the oil and gas industry, as hydrate formation and reformation in the field is a huge problem. Bacterial AFPs, though largely uncharacterized, may be the best candidates for large-scale production of hydrate inhibitors, given the difficulties in obtaining AFP from other sources. The popular kinetic inhibitors (KIs) polyvinylpyrrolidone (PVP) and polyvinylcaprolactam (PVCap) were used for points of comparison in experiments exploring the hydrate-inhibition activity of several ice-associating bacteria and proteins. The addition of the soil microbe, Chryseobacterium, increased the average lag-time to tetrahydrofuran (THF) hydrate formation by 14-fold, comparable to PVP or PVCap. Samples containing Pseudomonas putida, a bacterium having both ice-nucleation protein (INP) and AFP activity, had lag-times double that of the control. Solutions with P. putida and Chryseobacterium sometimes formed hydrate slurries of stunted crystal nuclei instead of solid crystals. No inhibition of memory or nucleation was noted in bacterial assays, however bacteria with INP activity was linked to unusually rapid memory reformation. Quartz crystal microbalance experiments with dissipation (QCM-D) showed that a tight adsorption to SiO2 and resistance to rinsing are correlated with a molecule’s inhibition of hydrate formation and reformation. These results support a heterogeneous nucleation model of the memory effect, and point to the affinity of AFP for heterogeneous nucleating particles as an important component of memory inhibition. / Thesis (Master, Biology) -- Queen's University, 2008-05-30 15:20:38.749

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Nanoporous molecular sieve materials like metal organic frameworks (MOFs) and metal oxide nanotubes (AlSiNTs) have found a wide range of technological applications in catalysis, separations, and ion exchange due to their salient features over other contemporary sensing materials. As a result, these materials can function as a chemical recognition layer that relies on analyte adsorption and they have shown to selectively adsorb specific gas molecules from mixtures. The characterization of gas adsorption in these materials is performed predominantly by commercial gravimetric equipment, whose capital and operating costs are generally high and require relatively large amounts of sample. Thus, it is desirable to obtain a reliable measure of the gas transport properties of these materials over a substantial range of pressure and temperature by non-gravimetric methods. The objective of this thesis is to investigate the adsorption and diffusion characteristics of recently-identified nanoporous materials through the development and use of a high-pressure/high-temperature quartz crystal microbalance (QCM) device. In this regard, this thesis is divided into four main objectives, viz. (1) Design and development of high temperature/ high pressure QCM device, (2) Measurement and analysis of adsorption characteristics in nanoporous materials, (3) Diffusion measurement and analysis in polymer thin films and (4) Diffusion measurement and analysis in MOF crystals. The results obtained in Objectives 2-4 will allow us to make important recommendations regarding the use of specific nanoporous materials in molecular separation applications and also lead to significant understanding of gas uptake thermodynamics in nanoporous materials via the application of analytical models to the experimental data.

Organic electrical energy storage (EES) is a growing field of research that is expected to play an important role in the future, as the need for sustainable EES increases. Conducting redox polymers (CRPs), i.e. conducting polymers with incorporated redox active moieties e.g. as pendant groups (PGs), are proposed as a promising class of compounds for this purpose. Redox cycling of the PGs can be utilized for high charge storage capacity, while the conducting polymer backbone provides fast charge transport through the material. Some of the major challenges with small-molecule systems for EES could be solved by using CRPs, e.g. capacity fading due to dissolution of the active compound, and high resistance due to slow charge transport between molecules. The latter issue is often solved by adding large amounts of conducting additives to the active material, drastically lowering the specific capacity. In this project, CRPs are shown to be able to function in battery cells without any additives, making both high capacity and high power possible. Although several CRPs have been reported in the literature, very few detailed studies have been conducted on the electrochemical processes of the two systems (i.e. the conducting polymer backbone and the redox active PGs). An important factor to consider in CRP design is the possibility for interaction between the two redox systems, which could be either beneficial or detrimental to the function as EES material. In this thesis, CRP model systems composed of hydroquinone functionalized polypyrrole have been studied, and they exhibit separate redox reactions for the PGs and the backbone, overlapping in potential. Significant interaction between them was observed, as oxidation of the PGs has severe impact on the backbone: When the oxidized and hydrophobic p-benzoquinone PGs are formed, they pack and force the polymer backbone to twist, localizing the bipolarons, and decreasing the conductivity. This is accompanied by a contraction of the polymer film and expulsion of electrolyte. Overall, the interaction in these polymers is destructive for their EES function, and it could be eliminated by introduction of a long linker unit between the PGs and the backbone.