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Mitochondrial proteins are presently believed to reside and function only within the mitochondria, under normal physiological conditions. However, in recent years many examples have come to light where proteins originally identified on the basis of extramitochondrial functions, upon characterization were found to be bonafide mitochondrial proteins. This raises important questions concerning the cellular functions of mitochondrial proteins. To investigate this phenomenon of mitochondrial proteins being present at extramitochondrial locations, we have examined by means of immunoelectron microscopy, the subcellular localization in normal rat tissues of a number of well characterized mitochondrial proteins which are encoded either by the nucleus (chaperonin 10 (Cpn10) and mitochondrial aspartate aminotransferase (mAspAT)), or by mitochondrial DNA (Cytochrome c oxidase subunits I and II (COX I and II)), using highly specific antibodies. Both Cp10 and mAspAT have already been shown to be involved in extramitochondrial functions. Cpn10 has been shown to be identical to early pregnancy factor, which is an immunosuppressant and growth factor found in maternal serum. mAspAT is identical to fatty acid binding protein isolated from plasma membranes of several cell types, involved in the transport of long chain free fatty acids. Immunofluorescent labeling of BS-C-1 African monkey kidney cells with these antibodies showed characteristic mitochondrial labeling. In all tissues examined, the antibodies showed strong labeling of mitochondria, as was expected. In several tissues, the binding was seen exclusively within mitochondria. However, in a variety of tissues such as pancreas, anterior pituitary and kidney, these antibodies showed strong and specific labeling of one or more of the following compartments- pancreatic zymogen granules, growth hormone granules, secretory granules in islet cells, red blood cells, condensing vacuoles in kidney and on the cell surface of different regions of the kidney. All of the observed labeling with these antibodies, both within mitochondria and in other compartments, was abolished upon omitting the primary antibodies or upon adsorption of the Cpn10 antibody (in the case of Cpn10) with purified recombinant protein. The extramitochondrial localization of these proteins, particularly those encoded by mitochondrial DNA provides strong evidence that these proteins have reached these sites by exiting from mitochondria. These observations have important implications concerning the roles of mitochondria and mitochondrial resident proteins in different mitochondrial diseases. Also, the function of these proteins and the precise mechanism by which they reach these extramitochondrial sites is of great interest. / Thesis / Master of Science (MSc)

The characterization of nanomaterials involved in dynamic processes are conventionally conducted using microscopy, spectroscopy and other physical/chemical methods through the pseudo-dynamic approach. In details, the dynamics processes are recorded by repeating or terminating the process multiple times. However, the above approach can lead to missing important transition information and inducing contamination for mechanistic studies. This motivates the efforts to develop real time characterization techniques which can probe the dynamic change of nanoparticles in their native operating environments. With the capability of probing structural change at the nanoscale, in situ transmission electron microscopy, has shown great potential in studies and applications of various processes. Targeting at conducting precise analysis, which has been limited by many uncertainties from electron beam effects and the miniaturized reaction cell used for TEM, the work presented herein pursues a quantitative characterization of a few electrochemical and biological processes through in situ liquid-phase transmission electron microscopy. In this work, the in situ transmission electron microscopy system is evaluated by comparing the in situ results with those from standard experiments to show its capabilities in studying dynamic processes. The in situ system is quantitatively calibrated to obtain the optimized observation conditions to avoid detectable electron beam interference, solution depletion and achieve sufficient resolution for analysis through micrometer thick liquid. These form the fundamentals for the in situ studies. Moreover, a comprehensive analysis protocol is established by incorporating multiple ex situ and post situ characterizations. Using this optimized in situ system, the mechanism of electrodeposition of gold on carbon electrode is studied. The in situ results allow quantitative analysis of the growth process. The prevailing diffusion limited three dimensional growth model is examined. A study of the effect of supporting electrolyte on the electrodeposition of palladium is also conducted. The self-limiting, surface diffusion and aggregation/recrystallization growth model is found to describe the early stage of growth, rather than the classical Volmer-Weber growth model. A further study is conducted on the structural evolution of palladium nanoparticles under electrochemical cycling. The mechanisms involved in this process, including electrodeposition, dissolution, hydrogen co-deposition and hydrogen desorption, are studied. The supporting electrolyte, HCl, is found to enhance the dissolution of deposited palladium clusters and induce movements and aggregation of the deposits during the hydrogen interaction process to form chain-like and irregular clusters, which provide direct experimental proof on the morphology formation of palladium with hydrogen involvement. Ultimately, the in situ technique is applied to the study of calcium phosphate biomineralization. Combined with multiple post situ characterization techniques, the study provides direct experimental evidence of the non-classical pre-nucleation and attachment growth of calcium phosphate structures. This demonstrates the potential of the in situ technique for studying the mechanisms involved in biological processes. / Dissertation / Doctor of Philosophy (PhD) / Nanostructured materials have been widely used in various fields. In situ transmission electron microscopy, a technique used to characterize nanomaterials involved in different dynamic processes in their operating environments, is an advanced tool over the traditional characterization methods such as ex situ microscopy and spectroscopy. However, there are several challenges in applying this in situ technique to processes occurring in liquid media. In this thesis, an in situ transmission electron microscopy system is applied to study the mechanisms of structural changes during different processes in liquids with both high spatial and temporal resolution. Protocols to evaluate and optimize the in situ system are developed to provide results comparable with those from their actual applications. Then in situ studies on the structural evolution of nanomaterials during electrochemical processes are performed and different theoretical models are applied to describe these processes. Finally, this technique is extended to investigate biomineralization to show its capabilities in future studies on biological processes.

Magnetotactic bacteria are a group of prokaryotes that share the ability to align with external magnetic fields, due to the presence within their cytoplasm of one or several chains of nanometer-sized magnetic crystals called the magnetosomes. The orientation of the chain within the cell is critical for magnetotaxis, which allows these bacteria to swim along the geomagnetic field lines. To do so, the magnetic moment and thus the chain need to lie parallel to the swimming direction which, for elongated bacteria such as AMB-1, is roughly parallel to the long axis of the cell. In most studies, the alignment between the magnetic moment and the cell axis is taken for granted, however no precise measurement has been performed to confirm this. In this thesis, experiments performed to test this assumption are presented, and the results show that for most studied bacteria the alignment is not perfect. The effect on the orientation distributions is discussed and accounted for in the analysis performed to measure the magnetic moment of individual bacteria. A second project presented in this thesis is focused on the biomineralization process in AMB-1. Magnetotactic bacteria synthesize crystals characterized by a well-controlled morphology and a high chemical purity, which makes them interesting for biomedical applications. To study how these crystals are produced, we used scanning trans- mission X-ray microscopy, and preliminary results show that this tool is suitable for studying this complex process. The methods developed and improved during this MSc to perform these experiments are presented, and the first results show an evolution in the spectroscopy of the magnetosomes as they grow. / Thesis / Master of Science (MSc)

Liquid cell electron microscopy has been proven to provide high spatial and temporal resolution for studying liquid layers and the solid-liquid interface at the micro and nano scale. The in situ environment allows for spatial and spectral characterization and quantification of the dynamics and kinetics involved with structural and chemical changes of nanostructures, which has seen application is fields of materials science, electrochemistry, corrosion, biomaterials, and nanophysics. The rapid growth of in situ liquid cell electron microscopy has motivated the fabrication of a custom liquid cell for improved control over the experimental conditions, including cell dimensions and materials. In this work, the process flow and micro-fabrication of a custom liquid cell system are proposed and executed, with proof of operation through the in situ imaging of suspended gold nanoparticles and electrochemical characterization. The in situ TEM system is improved upon by forming 1μm diameter holes through the viewing membranes, removing the background noise contribution from imaging electrons through the windows. This allows for high resolution liquid cell imaging. This improved system is used to study the oxidative etching conditions for palladium nanocrystals, which are commonly used as catalysis for hydrogen fuel cells. The dendritic etching is studied through native etching under exposure to hydrochloric acid without the presence of oxidizing species, followed by the radiolytic generation of oxidizing radicals via the microscope electron beam, and finally by the application of electrical biasing. / Thesis / Master of Applied Science (MASc) / Characterization of nanomaterials has been available for several decades and has aided in the improvement of material design, such as steel strength and corrosion resistance, electrical systems such as those involved in computers and smartphones, and biological sensing and detection. Observation of dynamic process which occur at the interface between solid and liquid phases, or purely within liquid layers, has always been a challenging topic due to the difficulty of finding a stable environment for both solids and liquids to exist at the nano scale within a measurement device. In situ liquid cell electron microscopy offers the ability to image this interface with real-time data acquisition for recording of dynamics and kinetics at the nano scale. Previous work has shown the liquid cell to provide high spatial and temporal resolution of systems in an environment which mimics their native operating conditions. This thesis addresses current developments for in situ systems and works to develop a custom liquid cell to further the applications of the liquid cell and provide improved control over experimental conditions. The work then aims to improve on current technology by increasing the spatial resolution obtainable. Finally, the technique is applied to study the structural changes of nanocrystals under various etching conditions as a demonstration of its’ capabilities.

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No description available.

The displacement damage induced by primary recoils of fusion neutrons in tungsten and alloys has been studied with self-ion irradiations, followed by damage characterization with electron microscopy. Tungsten and alloys (≤ 5 wt.% Re, Ta, V) were implanted with 2 MeV W+ ions over a dose range of 3.3×1017 - 2.5×1019 W+m-2 at temperatures ranging from 300 to 750°C. Dislocation loops of b = ½<111> (> 60%) and b = <100> were identified, and that ½<111> loops were found more thermally stable. Among loops that were large enough for nature determination, at least 50% were found to be of interstitial type, with larger fractions in high-temperature and high-dose conditions. The diameter of loops did not exceed 20 nm, with the majority being ≤ 5 nm. The loop number density varied between 1022 and 1023 m-3. The effects of ion dose, irradiation temperature, composition and grain orientation on damage microstructure were investigated. In-situ irradiations (150 keV W+ ions) were carried out as a complement to the bulk implantations. Qualitative trends in loop size, geometry and nature with irradiation dose and temperature were similar to bulk irradiated specimens. Also, the dynamics of defects and their effects on the damage evolution were explored. In-situ annealing of irradiated thin foils was performed to investigate the thermal stability of radiation damage in tungsten. The majority of microstructure transformations were completed within 15 min of annealing. However, extended durations did favour the increase of loop size and the fraction of ½<111> loops.

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