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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

MONITORING YEAST tRNA ADSORPTION BY QCM-D: AN OPPORTUNITY FOR OPTIMIZATION OF APTAMER SELECTION CONDITIONS

Shang, Jieting 11 1900 (has links)
RNA aptamers that bind to a wide range of targets with high affinity and specificity have been identified via the in vitro systematic evolution of ligands by exponential enrichment (SELEX). However, the process is quite unpredictable due in part to binding that occurs not only on the targets themselves but also on any of the other functional groups, moieties, or surfaces. Recent modelling work has shown that this level of “background binding” is a key parameter in the performance of aptamer selection processes. One strategy to minimize the amount of background binding is to pre-block those possible binding sites with a non-amplifiable nucleic acid molecule, such as yeast tRNA. It is also known that binding buffer conditions have strong effect on the binding affinity of nucleic acids. However, there are no detailed studies and little quantitative information available to guide the design of aptamer selection processes. In this study, the binding ability of yeast tRNA, which has comparable size with most RNA aptamer libraries, on both silicon dioxide and poly (ethylene terephthalate glycol) (PET-G) surfaces was studied using Quartz Crystal Microbalance with Dissipation (QCM-D). Silicon dioxide surface is a commonly used substrate for QCM-D tests on the adsorption behaviour of different nucleic acid. PET-G is a commonly used polymer substrate for the fabrication of microfluidic devices, which are advanced techniques for aptamer selection. The presence of specific divalent cations, for example Mg2+ over Ca2+, in binding buffers greatly enhanced the binding of yeast tRNA on silicon dioxide surfaces and PET-G surfaces. Proper NaCl concentration (100 mM) and MgCl2 concentration (5 mM) is necessary to enhance yeast tRNA binding on both surfaces. Yeast tRNA binding ability on silicon dioxide surfaces show more dependence on binding buffer pH than on PET-G surfaces. / Thesis / Master of Applied Science (MASc)
2

QUARTZ CRYSTAL MICROBALANCE STUDIES ON FRICTION MODIFIERS FOR LUBRICANT APPLICATIONS

Lehner, Carey 01 January 2015 (has links)
Lubricants are used in numerous applications to control friction and protect moving parts from fatigue. These fluids consist of a variety of surface active chemistries competing for the surface to provide performance. In order to develop fluids that meet the ever-increasing requirements (from legislation and manufacturers), techniques that can provide insight into surface adsorption, in real time, and relate it back to performance are critical. The objective of this work is to determine if Quartz Crystal Microbalance with Dissipation (QCM-D) is an effective technique to investigate surfactant adsorption in regimes that are common to the transportation lubricant industry. QCM-D is employed to quantify the mass, characterize the morphology, and quantify the kinetics of adsorption of common friction modifiers. The adsorption information is then compared to macroscopic properties (friction and corrosion prevention) to determine if this technique can aid in formulating future lubricants.
3

Analysis and metric development for the study of viscoelastic thin films utilising a quartz crystal microbalance

Mcnamara, Thomas January 2016 (has links)
The aim of this thesis is the creation of a set of tools for the quartz crystal microbalance (QCM-D) that aid in the measurement and quantification of soft viscoelastic thin films and experimental work demonstrating their use. The QCM-D is an acoustic technique that monitors structural changes occurring at the sensor's surface via changes in the sensor's resonance frequency and the rate of mechanical energy loss (dissipation). As a first approximation, the frequency shifts are used to measure mass changes on the sensor's surface, and dissipation shifts used to quantify changes in the rigidity of the film. Use of the QCM-D responses in this manner requires that the film is acoustically thin and rigid, limiting its application to soft films. To quantify mass and viscoelastic changes using the QCM-D, soft films either need to be approximated to a thin, rigid layer, or the frequency and dissipation responses modelled using a viscoelastic model. Such an approximation leads to the encompassment of all the viscoelastic properties into the single dissipation measurement in addition to potentially introducing errors in mass calculations. Existing commercial software allows for the deconvolution of film parameters such as the shear modulus and viscosity by fitting experimental data to a viscoelastic model. This analysis can only be done after the experimental data is collected however, and provides no guidance on future experiments, also commonly requiring an initial estimate of the parameter values under investigation. I have developed an experimental optimisation tool, termed the total parameter matrix sensitivity (TPM-sensitivity). It is defined as the Jacobian determinant of the QCM-D responses with respect to the parameters under investigation, e.g. the film's height, density, viscosity and shear modulus and the bulk fluid's density and viscosity. TPM-sensitivity is a measure of how readily resolvable and separable the film and bulk are when analysing the QCM-D responses. This enables the user to select the most mathematically important harmonics, and using this I was able to experimentally resolve the viscoelastic information of a soft film using frequency responses alone. I have also defined a classification system which categorises the QCM-D responses relative to a perfectly rigid and thin film. This provides guidance on the level of analysis required to gain information about the film parameters, with the limitations of commonly applied rules of thumb also demonstrated. Examples using these computational tools and metrics are also presented with data I obtained experimentally and from the literature. Of the experimental investigations, the curing process of a bulk elastomer is of particular importance due to the film being both soft and acoustically thick, demonstrating QCM-D use for a film not complying to either of thecommonly used film approximations.
4

Characterizing molecular-scale interactions between antimicrobial peptides and model cell membranes

Wang, Kathleen F 23 April 2014 (has links)
Due to the escalating challenge of antibiotic resistance in bacteria over the past several decades, interest in the identification and development of antibiotic alternatives has intensified. Antimicrobial peptides (AMPs), which serve as part of the innate immune systems of most eukaryotic organisms, are being researched extensively as potential alternatives. However, the mechanism behind their bactericidal capabilities is not well understood. Previous studies have suggested that AMPs may first attach to the cell membranes, leading to pore formation caused by peptide insertion, lipid removal in the form of peptide-lipid aggregates, or a combination of both mechanisms. In addition to the lack of mechanistic knowledge, a significant hurdle in AMP-based drug development is their potential cytotoxicity to mammalian cells. Understanding AMP interactions with eukaryotic model membranes would allow therapeutics to be tailored for preferential action toward specific classes of bacterial membranes. In this study, we developed novel methods of quartz crystal microbalance with dissipation monitoring (QCM-D) data analysis to determine the fundamental mechanism of action between eukaryotic and bacterial membrane mimics and select membrane-active AMPs. A new technique for creating supported membranes composed entirely of anionic lipids was developed to model Gram-positive bacterial membranes. Atomic force microscopy (AFM) imaging was also used to capture the progression of AMP-induced changes in supported lipid membranes over time and to validate our method of QCM-D analysis. QCM-D and AFM were used to investigate the molecular-scale interactions of four peptides, alamethicin, chrysophsin-3, sheep myeloid antimicrobial peptide (SMAP-29) and indolicidin, with a supported zwitterionic membrane, which served as a model for eukaryotic cell membranes. Since established methods of QCM-D analysis were not sufficient to provide information about these interaction mechanisms, we developed a novel method of using QCM-D overtones to probe molecular events occurring within supported lipid membranes. Also, most previous studies that have used AFM imaging to investigate AMP-membrane interactions have been inconclusive due to AFM limitations and poor image quality. We were able to capture high-resolution AFM images that clearly show the progression of AMP-induced defects in the membrane. Each AMP produced a unique QCM-D signature that clearly distinguished their mechanism of action and provided information on peptide addition to and lipid removal from the membrane. Alamethicin, an alpha-helical peptide, predominantly demonstrated a pore formation mechanism. Chrysophsin-3 and SMAP-29, which are also alpha-helical peptides of varied lengths, inserted into the membrane and adsorbed to the membrane surface. Indolicidin, a shorter peptide that forms a folded, boat-shaped structure, was shown to adsorb and partially insert into the membrane. An investigation of rates at which the peptide actions were initiated revealed that the highest initial interaction rate was demonstrated by SMAP-29, the most cationic peptide in this study. The mechanistic variations in peptide action were related to their fundamental structural properties including length, net charge, hydrophobicity, hydrophobic moment, accessible surface area and the probability of alpha-helical secondary structures. Due to the charges associated with anionic lipids, previous studies have not been successful in forming consistent anionic supported lipid membranes, which were required to mimic Gram-positive bacterial membranes. We developed a new protocol for forming anionic supported lipid membranes and supported vesicle films using a vesicle fusion process. Chrysophsin-3 was shown to favor insertion into the anionic lipid bilayer and did not adsorb to the surface as it did with zwitterionic membranes. When introduced to supported anionic vesicle films, chrysophsin-3 caused some vesicles to rupture, likely through lipid membrane disruption. This study demonstrated that molecular-level interactions between antimicrobial peptides and model cell membranes are largely determined by peptide structure, peptide concentration, and membrane lipid composition. Novel techniques for analyzing QCM-D overtone data were also developed, which could enable the extraction of more molecular orientation and interaction dynamics information from other QCM-D studies. A new method of forming supported anionic membranes was also designed, which may be used to further investigate the behavior of bacterial membranes in future studies. Insight into AMP-membrane interactions and development of AMP structure-activity relationships will facilitate the selection and design of more efficient AMPs for use in therapeutics that could impact the lives of millions of people per year who are threatened by antibiotic-resistant organisms.
5

Microfabricated acoustic sensors for the detection of biomolecules

Weckman, Nicole Elizabeth January 2018 (has links)
MEMS (Microelectromechanical Systems) acoustic sensors are a promising platform for Point-of-Care biosensing. In particular, piezoelectrically driven acoustic sensors can provide fast results with high sensitivity, can be miniaturized and mass produced, and have the potential to be fully integrated with sample handling and electronics in handheld devices. Furthermore, they can be designed as multiplexed arrays to detect multiple biomarkers of interest in parallel. In order to develop a microfabricated biosensing platform, a specific and high affinity biodetection platform must be optimized, and the microfabricated sensors must be designed to have high sensitivity and maintain good performance in a liquid environment. A biomolecular sensing system that uses high affinity peptide aptamers and a passivation layer has been optimized for the detection of proteins of interest using the quartz crystal microbalance with dissipation monitoring (QCM-D). The resulting system is highly specific to target proteins, differentiating between target IgG molecules and other closely related IgG subclasses, even in complex environments such as serum. Piezoelectrically actuated MEMS resonators are designed to operate in flexural microplate modes, with several modes shown to be ideally suited for fluid based biosensing due to improved performance in the liquid environment. The increase in quality factor of these MEMS microplate devices in liquid, as compared to air, is further investigated through the analytical and finite element modeling of MEMS fluid damping mechanisms, with a focus on acoustic radiation losses for circular microplate devices. It is found that the impedance mismatch at the air-water interface of a droplet is a key contributor to reduced acoustic radiation losses and thus improved device performance in water. Microplate acoustic sensors operating in flexural plate wave and microplate flexural modes are then integrated with a fluidic cell to facilitate protein sensing from fluid samples. Flexural plate wave devices are used to measure protein mass adsorbed to the sensor surface and initial results toward microplate flexural mode protein sensing are presented. Finally, challenges and areas of future research are discussed to outline the path towards finalization of a sensing platform taking advantage of the combination of the sensitive MEMS acoustic sensor capable of operating in a liquid environment and the specific and high affinity biomolecular detection system. Together, these form the potential basis of a novel Point-of-Care platform for simple and rapid monitoring of protein levels in complex samples.
6

Enrichment and Separation of Phosphorylated Peptides on Titanium Dioxide Surfaces : Applied and Fundamental Studies

Eriksson, Anna I. K. January 2013 (has links)
Protein phosphorylation is a very common posttranslational modification (PTM), which lately has been found to hold the keyrole in the development of many severe diseases, including cancer. Thereby, phosphoprotein analysis tools, generally based on specific enrichment of the phosphoryl group, have been a hot topic during the last decade. In this thesis, two new TiO2-based on-target enrichment methods are developed and presented together with enlightening fundamental results. Evaluation of the developed methods was performed by the analysis of: custom peptides, β-casein, drinking milk, and the viral protein pIIIa. The results show that: i) by optimizing the enrichment protocol (first method), new phosphorylated peptides can be found and ii) by the addition of a separation step after the enrichment (second method), more multi-phosphorylated peptides, which usually are hard to find, could be detected. The fundamental part, on the other hand, shows that the phosphopeptide adsorption is caused by electrostatic interactions, in general follows the Langmuir model, and the affinity increases with the phosphorylation degree. Here, however, the complexity of the system was also discovered, as the adsorption mechanism was found to be affected by the amino acid sequence of the phosphopeptide.
7

Understanding Submicron Foulants in Produced Water and their Interactions with Ceramic Materials

Medina, Sandra Constanza 11 1900 (has links)
Produced water (PW), or water associated with crude oil extraction, is the largest oily wastewater stream generated worldwide. The reuse and reclamation of these important water volumes are critical for more sustainable operation in the oilfield. Ceramic membrane filtration is a promising technology for PW treatment; however, fouling is the major drawback for a broader application. Fouling leads to higher resistance to flow, reducing membrane lifetime, and ultimately leading to higher capital expenditures and operating expenses. Further understanding of the interactions between PW foulants and the ceramic materials is needed for designing fouling control strategies and cleaning protocols for ceramic membranes. This work explored different techniques to characterize, visualize, and quantify the submicron PW contaminants content and its adsorption interactions with metal oxides. We visualized and characterized submicron oil droplets in oilfield PW samples by applying suitable advanced microscopy techniques. For the first time, crude oil droplets as small as 20 nm were found in oilfield PW together with other submicron contaminants. The adsorption studies performed by quartz crystal microbalance with dissipation (QCM-D) showed that the interactions of organic surface-active compounds with the metal oxides are driven by the nature of the surfactant and not by the surface properties. This has implications in the selection of the ceramic membrane material, wherein electrostatic interactions should not be taken as the only predicting factor of adsorption and fouling during PW treatment. Furthermore, our results suggested that the more fluid or viscoelastic-like the contaminant layer, the more difficult the cleaning process from the metal oxide. It demonstrates that the mechanical property of the attached films is a crucial factor in designing appropriate cleaning protocols for ceramic membranes. Finally, QCM-D and advanced microscopy techniques were applied to analyze adsorption and cleaning of contaminants in a complex Bahraini PW into alumina as a case study. Bacteria were found to attach irreversibly on the alumina surface, promoting nucleation points for calcium precipitates. The protocols developed in this work are suitable for understanding membrane fouling phenomena in the micron scale and could be implemented before filtration pilot testing to save time and expenses at larger scales.
8

Bio-inspired Cellulose Nanocomposites

Pillai, Karthik 07 October 2011 (has links)
Natural composites like wood are scale-integrated structures that range from molecular to the macroscopic scale. Inspired by this design, layer-by-layer (LbL) deposition technique was used to create lignocellulosic composites from isolated wood polymers namely cellulose and lignin, with a lamellar architecture. In the first phase of the study, adsorption of alkali lignin onto cationic surfaces was investigated using a quartz crystal microbalance with dissipation monitoring (QCM-D). Complete coverage of the cationic surface with alkali lignin occured at low solution concentration; large affinity coefficients were calculated for this system at differing pH levels. Adsorption studies with organosolv lignin in an organic solvent, and spectroscopic analysis of mixtures of cationic polymer with alkali lignin revealed a non-covalent interaction. The work demonstrated how noncovalent interactions could be exploited to molecular organize thin polyphenolic biopolymers on cationic surfaces. The second phase of the study examined the adsorption steps during the LbL assembly process to create novel lignocellulosic composites. LbL assembly was carried out using oxidized nanocellulose (NC) and lignin, along with a cationic polymer poly(diallyldimethylammonium chloride) (PDDA). QCM-D was used to follow the sequential adsorption process of the three different polymers. Two viscoelastic models, namely Johannsmann and Voigt, were respectively used to calculate the areal mass and thickness of the adsorbed layers. Atomic force microscopy studies showed a complete coverage of the surface with lignin in all the disposition cycles, however, surface coverage with NC was seen to increase with the number of layers. Free-standing composite films were obtained when the LbL process was carried out for 250 deposition cycles (500 bilayers) on a cellulose acetate substrate, following the dissolution of the substrate in acetone. Scanning electron microscopy of the cryo-fractured cross-sections showed a lamellar structure, and the thickness per adsorption cycle was estimated to be 17 nm. The third phase of the study investigated the effect of LbL ordering of the polymers versus a cast film composed of a blended mixture of the polymers, using dynamic mechanical analysis. A tan ï ¤ peak was observed in the 30 – 40 ºC region for both films, which was observed in the neat NC film. Heating of the samples under a compressive force produced opposite effects in the films, as the LbL films exhibited swelling, whereas the cast films showed densification. The apparent activation energy of this transition (65 – 80 kJ mol-1) in cast films, calculated based on the Arrhenius equation was found to be coincident to those reported for the ï ¢ transition of amorphous cellulose. The peak was seen to disappear in case of LbL films in the second heat, whereas it was recurring in case of cast films of the blended mixture, and neat NC films. Altogether, the together the work details a novel path to integrate an organized lignin and cellulose molecular structure, albeit modified from their native form, into a three-dimensional composite material. / Ph. D.
9

Studies of Biomacromolecule Adsorption and Activity at Solid Surfaces by Surface Plasmon Resonance and Quartz Crystal Microbalance with Dissipation Monitoring

Liu, Zelin 05 October 2010 (has links)
Self-assembly of polysaccharide derivatives at liquid/solid interfaces was studied by surface plasmon resonance spectroscopy (SPR) and quartz crystal microbalance with dissipation monitoring (QCM-D). Carboxymethyl cellulose (CMC) adsorption onto cellulose surfaces from aqueous solutions was enhanced by electrolytes, especially by divalent cations. A combination of SPR and QCM-D results showed that CMC formed highly hydrated layers on cellulose surfaces (90 to 95% water by mass). Voigt-based viscoelastic modeling of the QCM-D data was consistent with the existence of highly hydrated CMC layers with relatively low shear viscosities of ~ 10-3 N·s·m-2 and elastic shear moduli of ~ 105 N·m-2. Adsorption of pullulan 3-methoxycinnamates (P3MC) and pullulan 4-chlorocinnamates (P4CC) with different degrees of cinnamate substitution (DSCinn) onto cellulose, cellulose acetate propionate (CAP), poly(L-lactic acid) (PLLA), and methyl-terminated self-assembled monolayer (SAM-CH3) surfaces was also studied by SPR and QCM-D. Hydrophobic cinnamate groups promoted the adsorption of pullulan onto all surfaces and the adsorption onto hydrophobic surfaces was significantly greater than onto hydrophilic surfaces. SPR and QCM-D results showed that P3MC and P4CC also formed highly hydrated layers (70 to 90% water by mass) with low shear viscosities and elastic shear moduli. Finally, cellulose adsorption and activity on pullulan cinnamate (PC) and cellulose blend films were studied via QCM-D and in situ atomic force microscopy (AFM). The hydrophobicity of PC surfaces was controlled by adjusting the degree of cinnamate substitution per anhydroglucose unit (DSCinn). It was found that cellulase showed weak adsorption onto low DSCinn PC surfaces, whereas cellulase adsorbed strongly onto high DSCinn PC surfaces, a clear indication of the role surface hydrophobicity played on enzyme adsorption. Moreover, cellulase catalyzed hydrolysis of cellulose/PC and cellulose/polystyrene (PS) blend surfaces was studied. The QCM-D results showed that the cellulase hydrolysis rate on cellulose in cellulose/PC blend surfaces decreased with increasing DSCinn. AFM images revealed smooth surfaces for cellulose/PC (DSCinn = 0.3) blend surfaces and laterally phase separated morphologies for cellulose/PC (DSCinn ≥ 0.7) blend surfaces. The combination of QCM-D and AFM measurements indicated that cellulase catalyzed hydrolysis was strongly affected by surface morphology. The cellulase hydrolysis activity on cellulose in cellulose/PS blend surfaces was similar with cellulose/PC blend surfaces (DSCinn ≥ 0.7). These studies showed self-assembly of macromolecules could be a promising strategy to modify material surfaces and provided further fundamental understanding of adsorption phenomena and bioactivity of macromolecules at liquid/solid interfaces. / Ph. D.
10

Arabinoglucuronoxylan and Arabinoxylan Adsorption onto Regenerated Cellulose Films

Ni, Ying 10 January 2014 (has links)
Cellulose and hemicelluloses have attracted increasing interest as renewable biopolymers because of their abundance. Furthermore, the recognition of biomass as a sustainable and renewable source of biofuels has driven research into the assembly and disassembly of polymers within plant cell walls. Cellulose thin films are useful in the study of interactions between cellulose and hemicelluloses, and quartz crystal microbalances with dissipation monitoring (QCM-D), surface plasmon resonance (SPR) and atomic force microscopy (AFM) are widely used to investigate polymer adsorption/desorption at liquid/solid interfaces. In this study, smooth trimethylsilyl cellulose (TMSC) films were spincoated onto gold QCM-D sensors and hydrolyzed into ultrathin cellulose films upon exposure to aqueous HCl vapor. The adsorption of arabinoglucuronoxylan (AGX) and arabinoxylan (AX) onto these cellulose surfaces was studied. The effects of structure, molar mass and ionic strength of the solution were considered. Increasing ionic strength increased AGX and AX adsorption onto cellulose. While AGX showed greater adsorption onto cellulose than AX by QCM-D, the trend was reversed in SPR experiments. The combination of QCM-D and SPR data showed a greater amount of water was trapped within the AX films. Both adsorbed AGX and AX films were subsequently visualized by AFM. Images from AFM showed AGX and AX adsorbed as aggregates from water, while AGX and AX adsorbed from CaCl2 yielded smaller xylan particles with more numerous globular structures on the cellulose surfaces. Images from AFM of xylan films on bare gold surfaces also showed layers of uniform aggregates that were consistent with AX and AGX aggregation in solution. / Master of Science

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