Fluorescent labeling can improve the detection sensitivity in capillary electrophoretic (CE) separations down to attomolar concentrations. However, most fluorescent labels are not compatible with CE because their fluorescence properties and charge states are pH-dependent, they are often hydrophobic and they have a tendency to significantly change the properties of the analytes after labeling. A group of fluorescent labeling reagents have been prepared whose fluorophores have properties that are optimized for CE separations. These fluorophores have fluorescence properties and charge states that are independent of pH in the 2 < pH < 11 range. Their excitation maxima are also compatible with the 488 nm line of the Argon ion laser. A mono-cationic acridine-based fluorescent label was prepared and was found to not shift the pI of a labeled model protein in capillary isoelectric focusing separation (cIEF). Lower loading, due to increased sensitivity, led to better resolution of closely spaced isoform peaks having a pI = 0.05. A tri-anionic pyrene-based fluorescent labeling reagent was also synthesized and was used in the sodium dodecyl sulfate capillary gel electrophoresis (SDS-CGE) separation of proteins. The fluorophore led to an LOQ in the nM range, and did not alter the migration behavior of proteins in the sieving matrix. A third fluorescent labeling reagent was developed as a solid phase reagent (SPR) where the fluorophore was immobilized on a solid surface through a cleavable anchor. The fluorophore is di-anionic and is based on pyrene. The SPR was designed to allow the simultaneous capture and labeling of an analyte and the efficient release of the label-analyte conjugate under mild acidic conditions. The use of the SPR allowed the labeling of a diamine whose concentration was in the low nanomolar range. The SPR opens up the possibility for mono-labeling and proportional multiple labeling of proteins.

Die Plasmamembranspannung von eukaryotischen Zellen soll maßgeblich zur Regulation von zellulären Prozessen wie der Zellmigration, Mitose, Endo- und Exozytose, Membranreparatur, Osmoregulierung und Zellspreiten beitragen, welche zu einer Veränderung der Membranfläche und ihrer Deformation führt. In dieser Arbeit wurde die epitheliale Zelllinie MDCK II (Madin-Darby Canine Kidney) benutzt, um spannungsgesteuerte Oberflächenregulierung zu untersuchen. Indentationsexperimente kombiniert mit dem Herausziehen von Membrannanoröhren wurden mit Hilfe des Rasterkraftmikroskops (Atomic Force Microskope, AFM) durchgeführt, um lokale Variationen in der Membranspannung und überschüssiger Membranfläche als Funktion von äußeren Reizen abzuschätzen. Die verwendeten externen Stimuli beinhalten eine Veränderung der Funktionalität des Actomyosin-Cortexes durch die Wirkung von Blebbistatin und Cytochalasin D, sowie die Manipulation der Zytoskelett-Membran Adhäsionspunkte durch Einzel-Mikroinjektion. Die Injektion von Neomycin verhindert die Anbindung von ERM-Proteinen an das Lipid Phosphatidylinositol-(4,5)-bisphosphat (PIP2) und bewirkt somit die Abkopplung des Zytoskeletts von der Plasmamembran. Als Gegenexperiment diente die Injektion des Lipids PIP2 selbst, welches zur Erhöhung der Anzahl der Zytoskelett-Membran Adhäsionspunkte führte. Weiterhin wurden die als Membranreservoire dienenden Mikrovilli durch den Entzug von Cholesterol entfernt. Auswirkung auf das Vorhandensein von Membranreservoiren hat ebenfalls die Veränderung des osmotischen Drucks innerhalb der Zellen. Zusätzlich wurden die elastischen Eigenschaften von apikalen Zellmembran-Fragmenten von konfluenten MDCK II Zellen untersucht, welche Aufschluss über die intrinsischen Membraneigenschaften ohne den Einfluss des Zytosols und Zytoskeletts geben konnten. Abschließend wurde die Mechanik von adhärierenden und spreitenden Zellen untersucht. Zusammenfassend kann gesagt werden, dass die Plasmamembran, bestehend aus einer Phospholipiddoppelschicht, lateral schwer ausdehnbar ist aufgrund ihrer flüssig-kristallinen Natur. Durch das Vorhandensein von dynamischen Membranreservoiren wie Mikrovilli, die schnell auf Veränderungen der Membranspannung durch Membranhomöostase reagieren, werden zellulare Prozesse wie die Zellmotilität oder die Anpassung an osmotischen Stress ermöglicht. In der vorliegenden Arbeit gelang es gleichzeitig, die Membranspannung und die Verfügbarkeit von Membranfläche von adhärenten konfluenten als auch von adhärierenden und spreiten Zellen zu messen. Die durchgeführten Experimente ergaben ein detailliertes Bild wie sich die zelluläre Oberflächenregulierung in der Membranmechanik widerspiegelt.

The synthesis of the three 6”-deoxy-6”-thio glycolipid analogues β-D-Gal-(1→6)-β-D-Gal-(1→4)-β-D-Glu-(1→OCH2)-[1,2,3]-triazole-1-dodecane, β-D-Gal-(1→4)-β-D-Glu-(1→4)- β-D-Glu-(1→OCH2)-[1,2,3]-triazole-1-dodecane and β-D-Gal-(1→4)-β-D-Glu-(1→4)-β-D-Glu-(1→OCH2)-[1,2,3]-triazole-1-octadecane is presented here. Glycosylation at position O-4’ of a propargyl cellobioside glycosyl acceptor and position O-6’ of a propargyl lactoside glycosyl acceptor with a 6-thio-6-deoxy galactosyl donor gave rise to two unique trisaccharides that in turn underwent copper-catalyzed azide-alkyne cycloadditions with either 1-azidododecane or 1-azidooctadecane. The potential for each of these analogues to function as tethers of lipid bilayers to Au(111) was assessed primarily by differential capacitance experiments. Deposition of a bilayer of DMPC/cholesterol (70:30) by Langmuir-Blodgett (LB) transfer followed by Langmuir-Schaefer (LS) touch to a self-assembled monolayer of the O-6’ linked analogue, diluted with 1-β-D-thioglucose, failed. This led to simplifying the target architecture to diagnose the quality of the monolayers. A monolayer of the known monosaccharide 1-octadecane-4-(6-thio-β-D-galacto-pyranosyloxymethyl)-[1,2,3]-triazole1 prepared by LB transfer was found to support a lipid monolayer deposited by LS touch and this bilayer had the lowest minimum capacitance observed of 0.9 µF/cm2. An attempt to produce a bilayer by the same method using the trisaccharide bearing the C-18 alkane chain failed and this was attributed to high water solubility, which gave rise to poor organization at the air-water interface. A self-assembled monolayer of this variant went forward to produce a poor quality bilayer with a minimum capacitance of 7.1 µF/cm2, which was the lowest value obtained for the trisaccharide series of analogues. / Natural Sciences and Engineering Research Council of Canada (NSERC)

The increased interest in the offshore wind resource in both industry and academic and the extension of the wind field where offshore wind turbine can be deployed has stimulated quite a number of offshore wind turbines concepts. This thesis presents a design of mono-column platform supported for 5 MW baseline wind turbine developed by the National Renewable Energy Laboratory (NREL), with a single tether anchored to the seabed. The design, based on the pioneer concept SWAY, results from parametric optimized design processes which account for important design considerations in the static and dynamic view, such as the stability, natural frequency, performance requirements as well as the economic feasibility. Fully coupled aero-hydro-servo-elastic model is established in the time-domain simulation tool FAST (Fatigue, Aerodynamics, Structures, and Turbulence) with the hydrodynamic coefficients from HydroGen, an indoor program providing same outputs as the commercial software WAMIT. The optimized model is verified by imitating the frequency-domain approach in FAST and thus comparing the results with the frequency-domain calculations. A number of simulations with various wind and wave conditions are run to explore the effect of wind speed and wave significant height in various water depths. By modifying the optimized model to a downwind turbine with the nacelle rigidly mounted on the tower and the single tether connected to the platform by a subsea swivel, the modified models are more closed to the original SWAY-concept wind turbine. These models are compared based on the platform motion, tether tension, displacement, nacelle velocity and acceleration, resonant behavior as well as the damping of the coupled systems. The results of these comparisons prove the advantage of the modified model in performance. The modified model has also clarified itself a good candidate for deep water deployment.

京都大学 / 0048 / 新制・論文博士 / 博士(人間健康科学) / 乙第13303号 / 論人健博第6号 / 新制||人健||5(附属図書館) / 京都大学大学院医学研究科人間健康科学系専攻 / (主査)教授 青山 朋樹, 教授 黒木 裕士, 教授 松田 秀一 / 学位規則第4条第2項該当 / Doctor of Human Health Sciences / Kyoto University / DFAM

[pt] O presente trabalho apresenta um estudo sobre uma estrutura de contenção projetada para a duplicação da ferrovia FEPASA no Km 74 entre Santos e São Paulo, que consiste em uma cortina ancorada por tirantes. Essa estrutura foi instrumentada a fim de se medirem as cargas nos tirantes com uso de células de carga elétricas instaladas na cabeça dos mesmos. As medições de carga nos tirantes foram feitas durante os ensaios de recebimento, no momento da incorporação e também após a conclusão da obra. As cargas nos tirantes sofrem redução de seu valor no momento imediato a sua incorporação, durante a construção e após da data de terminação das obras da cortina. Constatou-se que houve perda de carga nos tirantes ao longo do tempo, chegando a 54 por cento de perda em um deles. Enfatizando a importância da instrumentação de campo para compreender melhor o comportamento das estruturas, na parte inicial desta dissertação é apresentado um desenvolvimento teórico e calibração de dezesseis eletroníveis como proposta para a medição de deslocamentos horizontais em obras geotécnicas. / [en] This work presents a study about a retaining structure designed for the duplication of the rail FEPASA on the 74th km between Santos and São Paulo. This structure, an anchored retaining wall, was instrumented in the anchors heads with strain gauges in order to monitor its loads. The load measurements occurred during the performance test, locking and also after the works were concluded. A decrease on anchors loads is noticed in the moment immediately after the locking, during construction and after the works finished. It was observed that a loss of load in the anchors occurred to a maximum of 54 per cent. Highlighting the relevance of field monitoring to better understand the structures behavior, in the first part of this dissertation we present a theoretical background as well as the calibration of sixteen electrolevels as a proposal for the measurement of horizontal displacements in Geotechnical works.

[pt] O presente trabalho tem por objetivo desenvolver uma formulação e um sistema que seja uma ferramenta auxiliar para o engenheiro de projeto na definição de modelos de bielas e tirantes de estruturas bi-dimensionais. No sistema desenvolvido o usuário pode partir de um modelo complexo, com um número grande de barras nas mais variadas direções. Essa solução inicial pode se basear nos resultados em tensão de uma análise linear elástica por elementos finitos e na experiência do projetista. Todavia, como o usuário terá muitas dúvidas quanto ao melhor modelo as ser usado, ele pode superpor no modelo inicial todas as soluções que achar possíveis. A função do sistema é escolher entre todas as soluções admissíveis (aquelas que respeitam equilíbrio e resistência) a que for mais econômica. Define-se como solução mais econômica aquela que corresponder ao menor consumo de armadura. A formulação do problema conduz a um problema de Programação Linear (PL) e o programa LINDO [2] é utilizado para resolvê-lo. A visualização do modelo é também parte do sistema desenvolvido e é feito a partir dos resultados gerados pelo programa LINDO. O trabalho se divide basicamente em três partes. Na primeira parte é feita uma revisão sobre modelos de bielas e tirantes e sobre fundamentos de programação linear. A segunda parte consta da formulação do problema e por ultimo os resultados do programa são comparados com aqueles fornecidos pela literatura. A ferramenta desenvolvida neste trabalho pode ser de grande utilidade para a definição de modelos de bielas e tirantes mas ela nem de longe tem a pretensão de substituir a experiência, a intuição e o bom senso do engenheiro de projeto que deve sempre prevalecer, inclusive na análise crítica dos resultados do programa. / [en] The present work has the aim of developing a formulation and a computational system, which should be used by the structural engineers an auxiliary tool in the definition of strut and tie models of bidimensional structures. In the developed system the user can define initially a very complex system with a great number of bars and many different directions. This initial solution can be based on both the stress results of a linear elastic finite element analysis and the experience of the engineer. As the user may be in doubt about the best model to be used he may superimpose in the initial model all the allowable solutions he may find. The aim of the system is to choose among all the allowable solution (these ones wich satisfy equilibrium and strenght) the more economical one. As more economical solution is considered the one which correspond to the least consumption of reinforcement. The problem formulation leads to a linear programming problem (LP) and the LINDO program [2] is used for solving it. The visualization of the model and results is also part of the system. It is based on the data generated by the LINDO program. The work is basicaly divided in three parts. In the first one a revision about strut and tie models as well as the basic knowledge in linear programming is presented. In the second part the problem formulation is shown and the last the results of the program are compared with those old the technical literature. The computational tool presented here may be of great utility in the definition of strut and tie models, but it has not by far the intention and the feeling of the engineers which shall ever prevail, inclusive in the critic analysis of the program results. / [es] El presente trabajo tiene como objetivo desarrollar un sistema que sirva como herramienta auxiliar para el ingeniero de proyecto en la definición de modelos de bielas y tirantes de extructuras bidimensionales. En el sistema desarrollado, el usuario puede partir de un modelo complejo, con un número grande de barras en varias direcciones. Esa solución inicial puede basearse en los resultados en tensión de un análisis lineal elástica por elementos finitos y en la experiencia del proyectista. Como el usuario tendrá muchas dudas respecto al mejor modelo as ser usado, él puede superponer en el modelo inicial todas las soluciones que juzgue posibles. La función del sistema es elegir entre todas las soluciones admisibles (aquellas que respetan equilibrio y resistencia) la que sea más económica. Se define como solución más económica aquella que correspdonda al menor consumo de armadura. La formulación del problema conduce a un problema de Programación Lineal (PL) que es resuelto con ayuda del programa LINDO [2]. La visualización del modelo también es parte del sistema desarrollado y parte de los resultados generados por el programa LINDO. El trabajo se divide básicamente en tres partes. En la primeira parte se realiza una revisión de los modelos de bielas y tirantes y de los fundamentos de programación lineal. La segunda parte contiene la formulación del problema y la presentación de los resultados del programa se comparan con los reportados en la literatura. La herramienta desarrollada en este trabajo puede ser de gran utilidad para la definición de modelos de bielas y tirantes pero no tiene la pretensión de substituir la experiencia, la intuición y el buen senso del ingeniero de proyecto que debe siempre prevalecer, incluso en el análisis crítico de los resultados del programa.

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Currency has changed a lot, and the introduction of the Internet sped up the evolution of the currency. Digital currency introduced many benefits compared to physical currencies. Ideas such as cryptocurrencies work as an option for other means of payment. During the recent pandemic, interest in new digital currencies has increased, leading to more research on digital currency. With the introduction of new currencies and their increased popularity, many central banks have started looking into the idea of innovating currency. All this new research has coined the term central bank digital currency. As of today, there is no single idea on how a digital currency should work or be implemented. With many variations, the future is still unclear. There seem to be vulnerabilities to solve and many potential ways to improve current systems. When building this new currency it is crucial to know what different use cases could demand from the implementation. In conclusion, the results show that digital currency is still in early development, with central bank digital currency research showing promise. It is theoretically possible to create a better transaction solution contra traditional currencies. More research is needed on the topic of digital currency, but there could be incremental improvements to today’s currency leading to better future solutions. / Valuta har förändrats genom tiderna och introductionen av internet skyndade på denna utveckling. Digital valuta har introducerat många fördelar jämfört med fysiska valutor. Fler idéer som till exempel kryptovalutor har introducerats som alternativa betalmedel. Under den senaste pandemin så har intresset för nya digitala valutor ökat, vilket har lett till mer forskning inom området av digitala valutor. På grund av växande popularitet och nya digitala valutor så har många central banker börjat testa idéen om att nyskapa valuta, och med detta så har termen centralbanks valuta skapats. Det finns inte idag en enda lösning på hur digitala valutor ska fungera eller bli implementerade. Med många varianter så är framtiden fortfarande oklar. Det verkar finnas problem att lösa och många möjliga sätt att förbättra existerande system. Vid byggandet av denna nya valuta så är det extremt viktigt att veta vad som är viktigt och hur det kan finnas olika krav beroende på hur valutan ska användas. Enligt resultaten så är digital valuta fortfarande tidigt i sin utveckling och forsking gällande centralbanks styrda digitala valutor verkar lovande. Mer forsking kommer att behövas inom området digital valuta, men det kan komma många små förbättringar på dagens valuta som leder till bättre framtida lösningar.

During vertebrate development, gastrulation leads to the formation of three distinct germlayers. In zebrafish a central process is the delamination and the ingression of single cells from a common ancestor tissue - that will lead to the formation of the germlayers. Several molecules have been identified to regulate this process but the precise cellular mechanisms are poorly understood. Differential adhesiveness, a concept first introduced by Steinberg over 40 years ago, has been proposed to represent a key phenomena by which single hypoblast cells separate from the epiblast to form the mesendoderm at later stages. In this work it is shown that differential adhesion among the germlayer progenitor cells alone cannot predict germlayer formation. It is a combination of several mechanical properties such as cell cortex tension, cell adhesion and membrane mechanical properties that influence the migratory behavior of the constituent cells.