The use of sol-gel immobilization in the fabrication of microarrays is a relatively new approach that has shown potential to become a leading methodology in this field. However, there are a limited number of assay systems that have been reported using this method. Furthermore, methods to produce high-density sol-gel-derived microarrays have not been reported. Herein, two novel assays utilizing sol-gel-derived microarrays are presented. In the first case, the solid phase of sol-gel-derived microarrays was employed as a detection platform for monitoring the activity of glycogen synthase kinase 3-β (GSK3β) in solution using a phosphospecific stain. Using this assay format, the ability to detect hyperphosphorylated product over the pre-phosphorylated substrate was demonstrated and a z' value of 0.49 was obtained, indicating amenability to small molecule screening. Secondly, a fluorogenic assay for acetylcholinesterase (AChE) was development that is compatibility with sol-gel derived microarrays and standard imaging instrumentation. A thiol-reactive fluorogenic dye, typically used for detection of thiolated oligonucleotides, was successfully used to monitor AChE activity both in solution and in silica. Further, a functional sol-gel-derived AChE microarray was fabricated and activity on array was detected. We have also reported on the optimization of materials for the fabrication of high-density kinase microarrays using sol-gel immobilization. By employing a directed criteria-based screen, optimal materials were quickly and efficiently identified. Two materials, 1.5SS/1PV A/Glycerol and 0.25DGS, were identified as the optimal materials for fabrication of sol-gel-derived functional microarrays. / Thesis / Master of Science (MSc)

<p> A new technique for the systematic identification of metal-protein complexes combining gel chromatography with either neutron activation analysis or radioactive tracer methods has been proposed. The technique has been tested on the copper in serum situation to evaluate the results obtained on a well-known system.</p> <p> It was then applied to manganese in serum, manganese in erythrocytes and copper in erythrocytes. The results indicate that serum contains two manganese-binding proteins, one of low molecular weight and relatively labile in nature, the other of higher molecular weight and incorporating radioactive manganese in vivo at some definite time interval subsequent to the isotope's administration.</p> <p> Manganese in erythrocytes occurs as a porphyrin bound to apoglobin as a manganese analogue of hemoglobin.</p> <p> Copper in erythrocytes appears to exist in two forms - one firmly bound to erythrocuprein, the other more loosely bound to the same protein.</p> / Thesis / Doctor of Philosophy (PhD)

Listeria monocytogenes is a deadly, Gram-positive foodborne pathogen that is ubiquitous in the environment. The bacterium expresses a number of virulence and stress adaptation proteins that support its pathogenic capabilities. Two-dimensional gel electrophoresis (2-DE) was used to map L. monocytogenes surface proteins, which play a central role in virulence, and to examine protein expression by L. monocytogenes grown on ready-to-eat meat, an important source of Listeria infections. A novel method for solubilization of surface proteins from L. monocytogenes for 2-DE was developed. Additionally, the unique proteome expressed by L. monocytogenes grown on a meat matrix was uncovered. The developed solubilization method will facilitate efforts to identify and routinely compare surface proteins of Listeria by 2-DE. Furthermore, the 2-DE database of proteins expressed by L. monocytogenes grown on a meat matrix will allow further understanding of the interactions of Listeria with its food environment that influence its ability to cause disease.

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The functionality of a novel 3-dimensional betavoltaic battery design will be investigated to improve conversion efficiency over existing planar devices. A beta-emitting isotope of nickel, Ni-63, is embedded in the volume of empty space between self-assisted p-i-n junction gallium phosphide nanowires to improve the beta capture efficiency. Parameters such as nanowire pitch, diameter, and height will influence the efficiency and were investigated thoroughly. Material selection was performed based on the following considerations. Gallium phosphide is chosen to achieve a high open circuit voltage under beta exposure. Ni-63 has an optimal beta energy spectrum for a nanowire device and a half-life of 101 years for long term application. The majority of the work focused on the development of the fabrication process, particularly the radioactive source deposition. The method used for embedding the source was a citrate-based sol-gel which was spun onto the sample. This method was modified for this nanowire application and specific challenges to the process are outlined. Furthermore, the obstacles of working with radioactive materials will be discussed. The first nanowire-based betavoltaic device is reported to produce beta-generated current and achieved a beta conversion efficiency of 0.03%. Investigation of the junction was performed to provide future improvements to the efficiency. Additionally, simulated IV curves for a non-active sample exhibited a possible conversion efficiency of 1.92%. / Thesis / Master of Applied Science (MASc)

The sol-gel process has been employed to generate hybrid inorganic-organic network materials. Unique ceramers were prepared based on an alkoxysilane functionalized soft organic oligomer, poly(propylene oxide) (PPO), and tetramethoxysilane (TMOS). Despite the formation of covalent bonds between the inorganic and organic constituents, the resulting network materials were phase separated, composed of a silicate rich phase embedded in a matrix of the organic oligomer chains. The behavior of such materials was similar to elastomers containing a reinforcing filler. The study focused on the influence of initial oligomer molecular weight, functionality, and tetramethoxysilane, water, and acid catalyst content on the final structure, mechanical and thermal properties. The sol-gel approach has also been exploited to generate thin, transparent, abrasion resistant coatings for metal substrates. These systems were based on alkoxysilane functionazized diethylenetriamine (DETA) with TMOS, which generated hybrid networks with very high crosslink densities. These materials were applied with great success as abrasion resistant coatings to aluminum, copper, brass, and stainless steel. In another study, intercalated polymer-clay nanocomposites were prepared based on various epoxy networks montmorillonite clay. This work explored the influence of incorporated clay on the adhesive properties of the epoxies. The lap shear strength decreased with increasing clay content. This was due to a reduction in the toughness of the epoxy. Also, the delaminated (or exfoliated) nanocomposite structure could not be generated. Instead, all nanocomposite systems possessed an intercalated structure. The final project involved the characterization of a series of metallocene catalyzed linear polyethylenes, produced at Phillips Petroleum. Polyolefins synthesized with such new catalyst systems are becoming widely available. The influence of molecular weight and thermal treatment on the mechanical, rheological, and thermal behavior was probed. Although the behavior of this series of metallocene polyethylenes was not unlike that of traditionally catalyzed materials, this work is one of the first comprehensive studies of these new linear polyethylenes. The main distinction between the metallocene and traditional Ziegler-Natta catalyzed polyethylenes is the narrow molecular weight distributions produced by the former (for this series of materials, 2.3< Mw  Mn <3.6). / Ph. D.

Nature is able to convert chemical energy into mechanical work under modest conditions, i.e., physiological pH and ambient temperature and pressure. One of the most interesting systems is muscle modeled as the "sliding filament" system. The sliding filament system is a combination of a thin actin filament and a thick myosin filament that slide over one another by breaking the "energy-rich" pyrophosphate bond of ATP. The energy from ATP hydrolysis is used for mechanical motion and the energy lost during this process is used to heat our body. In biology, the sliding filament system is taken as a fairly effective model. For engineering systems, the energy lost to heat needs to be reduced to build an efficient energy converter. In our research, we use a phosphate charged protein, casein, and react it with divinyl sulfone (DVS) through a Michael addition reaction to produce a cross-linked gel. The protein gel could be ephosphorylated at standard conditions using bovine phosphatase (bp) and re-phosporylated using casein kinase. When attached to the protein, the negatively charged phosphate groups cause the gel to expand from repulsion. When removed, the protein contracts. Therefore, work is realized without sliding friction, which is the origin of the large energy loss in muscle. FT-IR spectroscopy allows us to follow the two biochemical reactions. We also show a thermodynamic analysis of the work and offer an estimation of the most basic term. / Master of Science

Aufgrund des unbegrenzten Angebots sowie ihrer physikalischen und biologischen Eigenschaften nimmt die Bedeutung von synthetischen Knochenersatzmaterialien stetig zu. Im Fokus der Forschung steht dabei vor allem die Verbesserung der Festigkeit dieser Materialien, die bisher nur in nicht lasttragenden Bereichen eingesetzt werden, die Biokompatibilität gegenüber körpereigenen Zellen sowie die Resorbierbarkeit. In der vorliegenden Arbeit sollten deshalb dualhärtende Bruschit-Zemente mit Organosilicat-Matrizen und mit Baghdadit substituierte Bruschit-Zemente auf die oben genannten Eigenschaften untersucht werden. Für die dualhärtenden Zemente ergab sich die höchste Druckfestigkeit für Silicat-Matrizen auf Basis von TEOS mit 16,27 MPa gegenüber Bruschit. Für die Hybrid-Silicat-Matrizen konnte hingegen keine Steigerung der langfristigen Druckfestigkeit gezeigt werden. Aufbauend auf unsere Untersuchungen sollte darüber hinaus die initiale Druckfestigkeit der verwendeten Systeme in den ersten 24 Stunden untersucht werden, da die Silicat-Matrizen zu deren Erhöhung beitragen könnten. Alle Zemente mit Bruschitanteil befanden sich zudem nach spätestens 10 Stunden im pH-neutralen Bereich zwischen 7,26 und 7,31. Bei den dualhärtenden Zementen mit Silicat-Matrizes ergab sich die beste Biokompatibilität. Diese zeigte sich jedoch nur bei den Bruschit-Zementen mit TEOS und 1,8-Bis-(triethoxysilyl)octan mit einer relevanten Signifikanz. MG-63 zeigte nach 9 Tagen eine 8-28x Erhöhung der Zellaktivität und eine 6-28x Erhöhung der Zellzahl gegenüber der Bruschit-Kontrolle. Auch bei RAW 264.7 konnte auf den Zementen mit beiden Präkursor nach 15 Tagen 90 ng/ml DNA detektiert werden, während diese auf Bruschit kaum zu detektieren war. Dies lässt sich einerseits durch eine Steigerung der Biokompatibilität durch den Silicat-Anteil und die Veränderung der Porengröße erklären, andererseits auch durch eine mögliche Vernetzung mit dem Abbindeverzögerer Citrat, der die Adhärenz von Zellen reduziert. Eine weitere Steigerung könnte zudem durch die Verwendung von besser biokompatiblen Abbindeverzögerern wie Phytat erreicht werden. / Due to the unlimited supply and their physical and biological properties, the importance of synthetic bone replacement materials is constantly increasing. Research is focussing primarily on improving the strength of these materials, which have so far only been used in non-load-bearing areas, their biocompatibility with the body's own cells and their resorbability. In the present study, dual- setting brushite cements with organosilicate matrices and brushite cements substituted with baghdadite were therefore to be analysed for the above-mentioned properties. For the dual-setting cements, the highest compressive strength was found for silicate matrices based on TEOS with 16.27 MPa compared to brushite. In contrast, no increase in long-term compressive strength could be demonstrated for the hybrid silicate matrices. Based on our investigations, the initial compressive strength of the systems used in the first 24 hours should also be investigated, as the silicate matrices could contribute to their increase. All cements with brushite content were also in the pH-neutral range between 7.26 and 7.31 after 10 hours at the latest. The dual-setting cements with silicate matrices showed the best biocompatibility. However, only the brushite cements with TEOS and 1,8-bis-(triethoxysilyl)octane showed a relevant significance. After 9 days, MG-63 showed an 8-28x increase in cell activity and a 6-28x increase in cell count compared to the brushite control. In the case of RAW 264.7, 90 ng/ml DNA was also detected on the cements with both precursors after 15 days, while this was hardly detectable on brushite. This can be explained on the one hand by an increase in biocompatibility due to the silicate content and the change in pore size, and on the other hand by possible cross-linking with the setting retarder citrate, which reduces the adherence of cells. A further increase could also be achieved through the use of more biocompatible setting retarders such as phytate.