The primary aim of the work presented in this thesis is to elucidate novel information regarding the uptake, storage, distributions, and functions of both copper and zinc in mammalian cells by predominantly using a combination of the high resolution imaging modalities, synchrotron radiation X-ray fluorescence microscopy (SXRF) and standard fluorescence imaging. Results from studies using cell permeable, metal ion selective fluorescent probes suggested the presence of labile pools of copper and zinc localized within the mitochondria and Golgi apparatus. Furthermore, SXRF imaging of a cell line defective in the copper transporter, Atox1, revealed intriguing differences in the Cu distribution of Atox1-/- cells compared to the corresponding wild-type cells. Finally, spatially well-resolved SXRF elemental maps of single, adherent mouse cells revealed remarkable changes in the distributions of both zinc and copper as the cells progressed through the cell cycle. Taken together, findings suggested major roles for copper and zinc within a native biological setting.

Cartilage tissue engineering has been emerging as a promising therapeutic approach, where engineered constructs or scaffolds are used as temporary supports to promote regeneration of functional cartilage tissue. Hybrid constructs fabricated from cells, hydrogels, and solid polymeric materials show the most potential for their enhanced biological and mechanical properties. However, fabrication of customized hybrid constructs with impregnated cells is still in its infancy and many issues related to their structural integrity and the cell functions need to be addressed by research. Meanwhile, it is noticed that nowadays monitoring the success of tissue engineered constructs must rely on animal models, which have to be sacrificed for subsequent examination based on histological techniques. This becomes a critical issue as tissue engineering advances from animal to human studies, thus raising a great need for non-invasive assessments of engineered constructs in situ. To address the aforementioned issues, this research is aimed to (1) develop novel fabrication processes to fabricate hybrid constructs incorporating living cells (hereafter referred as “construct biofabrication”) for cartilage tissue regeneration and (2) develop non-invasive monitoring methods based on synchrotron X-ray imaging techniques for examining cartilage tissue constructs in situ. Based on three-dimensional (3D) printing techniques, novel biofabrication processes were developed to create constructs from synthetic polycaprolactone (PCL) polymer framework and cell-impregnated alginate hydrogel, so as to provide both structural and biological properties as desired in cartilage tissue engineering. To ensure the structural integrity of the constructs, the influence of both PCL polymer and alginate was examined, thus forming a basis to prepare materials for subsequent construct biofabrication. To ensure the biological properties, three types of cells, i.e., two primary cell populations from embryonic chick sternum and an established chondrocyte cell line of ATDC5 were chosen to be incorporated in the construct biofabrication. The biological performance of the cells in the construct were examined along with the influence of the polymer melting temperature on them. The promising results of cell viability and proliferation as well as cartilage matrix production demonstrate that the developed processes are appropriate for fabricating hybrid constructs for cartilage tissue engineering. To develop non-invasive in situ assessment methods for cartilage and other soft tissue engineering applications, synchrotron phase-based X-ray imaging techniques of diffraction enhanced imaging (DEI), analyzer based imaging (ABI), and inline phase contrast imaging (PCI) were investigated, respectively, with samples prepared from pig knees implanted with low density scaffolds. The results from the computed-tomography (CT)-DEI, CT-ABI, and extended-distance CT-PCI showed the scaffold implanted in pig knee cartilage in situ with structural properties more clearly than conventional PCI and clinical MRI, thus providing information and means for tracking the success of scaffolds in tissue repair and remodeling. To optimize the methods for live animal and eventually for human patients, strategies with the aim to reduce the radiation dose during the imaging process were developed by reducing the number of CT projections, region of imaging, and imaging resolution. The results of the developed strategies illustrate that effective dose for CT-DEI, CT-ABI, and extended-distance CT-PCI could be reduced to 0.3-10 mSv, comparable to the dose for clinical X-ray scans, without compromising the image quality. Taken together, synchrotron X-ray imaging techniques were illustrated promising for developing non-invasive monitoring methods for examining cartilage tissue constructs in live animals and eventually in human patients.

<p>  </p> <p>Due to miniaturization of electronic devices new electronic packaging strategies, such as Heterogeneous Integration Packaging (HIP), are being developed. In HIP, the space in the package is strategically mapped out to maximize the placement of components including all types of materials. Thus, there is a need to develop and understand the behavior of lower-melting point metallic interconnects as they will be located next to lower melting point materials, such as polymers. </p> <p>The composition of alloying elements in Sn-rich solder plays a pivotal role in determining the microstructural properties of the solder joint. However, the complex mechanisms governing the solidification processes of Sn-In, and Sn-Bi alloys are still not fully understood. Furthermore, the experimental characterization of phase formation poses significant challenges.</p> <p>This dissertation focuses on understanding microstructural evolution in Sn-In and Sn-Bi alloys during reflow. A systematic approach to characterizing the microstructure of alloys was developed, utilizing electron microscopy, non-destructive x-ray tomography and diffraction techniques, ranging from lab-scale to synchrotron experiments. The influence of In addition on microstructure was correlated with the mechanical behavior obtained using nanoindentation. The experimental understanding was further correlated with the Density Functional Theory (DFT) calculations. To study the Sn-Bi microstructures, the effect of experimental parameters, such as the cooling rate during solidification was elucidated. A 4D study was conducted, involving the analysis of 3D microstructures along with time evolution, to gain a comprehensive understanding of the solidification dynamics using synchrotron white beam tomography. For the first time, we observed a regular pyramidal morphology of Bi forming in the solder alloy. The 4D analysis provided crucial insights into morphology formation, growth kinetics, defect formation during solidification. The crystallographic analysis unraveled unique insights into the solid-liquid interface stability for semi-metals. Furthermore, the simultaneous Energy Dispersive Diffraction (EDD) analysis yielded a deeper understanding into the phase formation and lattice strain evolution. A fundamental relationship between the diffraction intensity and phase fractions, from imaging, was obtained. The experimental methodology developed in this work has the potential to be extended to investigate a wide range of alloy solidification mechanisms, enabling a deeper understanding of these materials.</p>