Structural characterization is essential to understand the formation mechanisms of the nanostructures in thin absorber layers in third generation solar cells and amyloid protein aggregates. Since to the dimension of the precipitated structures is in nanometer scale, electron tomography technique in transmission electron microscopy (TEM) has been applied as a major tool to analyze the 3D order and distribution of precipitates using the electron tomography technique.  Silicon rich silicon carbide (SRSC) films were deposited by plasma enhanced chemical vapor deposition (PECVD) technique and annealed in the nitrogen atmosphere for 1 hour at 1100 °C. The spectrum-imaging (SI) technique in Energy filtered TEM (EFTEM) imaging mode was used to develop electron tomography. From the reconstructed sub-volumes, the complex, three dimensional interfacial nanostructure between the precipitated NPs and their parental matrix was observed and explained in terms of thermodynamic concepts. Additionally, the feasibility of raw data 4D electron tomography has been demonstrated using the EFTEM SI dataset. The aggregation process of the human islet amyloid polypeptide (hIAPP) has a great impact on human health. In this thesis, a model system has been taken to study the ultrastructure of the hIAPP aggregates that are present in the fat body tissue surrounding the brain of Drosophila melanogaster. Electron tomography technique on TEM revealed clear crystalline structures in 3D. For the first time, the presence of a 5-fold twinned structure in biology was discovered. An intriguing finding is the filament like interconnection of hIAPP protein granules observed predominantly along the nearest neighbor directions. This suggests the existence of the directional binding forces between two nearest protein granules in addition to dipole-dipole interactions.

Organic solar cells using carbon based materials have the potential to deliver cheap solar electricity. The aim is to be able to produce solar cells with common printing techniques on flexible substrates, and as organic materials can be made soluble in various solvents, they are well adapted to such techniques. There is a large variation of organic materials produced for solar cells, both small molecules and polymers. Alterations of the molecular structure induce changes of the electrical and optical properties, such as band gap, mobility and light absorption. During the development of organic solar cells, the step of mixing of an electron donor and an electron acceptor caused a leap in power conversion efficiency improvement, due to an enhanced exciton dissociation rate. Top performing organic solar cells now exhibit a power conversion efficiency of over 10%. Currently, a mix of a conjugated polymer, or smaller molecule, and a fullerene derivative are commonly used as electron donor and acceptor. Here, the blend morphology plays an important role. Excitons formed in either of the donor or acceptor phase need to diffuse to the vicinity of the donor-acceptor interface to efficiently dissociate. Exciton diffusion lengths in organic materials are usually in the order of 5-10 nm, so the phases should not be much larger than this, for good exciton quenching. These charges must also be extracted, which implies that a network connected to the electrodes is needed. Consequently, a balance of these demands is important for the production of efficient organic solar cells. Morphology has been found to have a significant impact on the solar cell behaviour and has thus been widely studied. The aim of this work has been to visualize the morphology of active layers of organic solar cells in three dimensions by the use of electron tomography. The technique has been applied to materials consisting of conjugated polymers blended with fullerene derivatives. Though the contrast in these blends is poor, three-dimensional reconstructions have been produced, showing the phase formation in three dimensions at the scale of a few nanometres. Several material systems have been investigated and preparation techniques compared. Even if excitons are readily dissociated and paths for charge extraction exist, the low charge mobilities of many materials put a limit on film thickness. Although more light could be absorbed by increased film thickness, performance is hampered due to increased charge recombination. A large amount of light is thus reflected and not used for energy conversion. Much work has been put into increasing the light absorption without hampering the solar cell performance. Aside from improved material properties, various light trapping techniques have been studied. The aim is here to increase the optical path length in the active layer, and in this way improve the absorption without enhanced extinction coefficient. At much larger dimensions, light trapping in solar cells with folded configuration has been studied by the use of optical modelling. An advantage of these V-cells is that two materials with complementing optical properties may be used together to form a tandem solar cell, which may be connected in either serial or parallel configuration, with maintained light trapping feature. In this work optical absorption in V-cells has been modelled and compared to that of planar ones.

Electron tomography is an invaluable technique with the capability of carrying out thorough 3D structural, chemical and morphological characterization of materials at nanometer scale. Tilting range, increment and reconstruction algorithms are three of the main factors affecting the quality of tomograms. An anisotropic degradation can be observed with restricted tilting range and increment. Therefore, this study was carried out to investigate the accuracy of the reconstruction results of MgO (cube-shape) generated by FBP, SART and SIRT tomographic algorithms under various reconstruction conditions, i.e. tilting range and increment. Examining the experimental data with known morphology permits quantitative determination of the accuracy of the reconstruction results by measuring the distortion of the cube in all directions. Moreover, distortion measurements in all directions reveal the relationship between level of distortion and the alpha tilt angle. / Master of Science

Electron tomography allows visualization of objects in a form of reconstructed 3D virtual volumes with resolution power of electron microscopy. The thesis is focused primarily on biological applications of electron tomography applied on tilt series images acquired in transmission electron microscope at room temperature. Specifically, the interaction of tick-borne encephalitis virus with neural cells and 3D ultrastructure of the central electron-dense part of the flagellum 9 + 1 (Caryophyllaeides fennica) were investigated. Finally, electron tomography was combined and correlated with atomic force microscopy to allow repetitive examination of ultrathin sections on electron microscopy grids.

No description available.

Cryogenic electron tomography provides visualization of cellular complexes in situ, allowing a further understanding of cellular function. However, the projection images from this technique present a meager signal-to-noise ratio due to the limited electron dose, and the lack of projections at high tilt angles produces the 'missing-wedge' problem in the Fourier domain. These limitations in the projection data prevent traditional reconstruction techniques from achieving good reconstructions. Multiple strategies have been proposed to deal with the noise and the artifacts arising from the 'missing-wedge’ problem. For example, manually selecting subtomograms of identical structures and averaging them (subtogram averaging), data-driven approaches that intend to perform subtogram averaging automatically, and various methods for denoising tilt-series before reconstruction or denoising the volumes after reconstruction. Most of these approaches are additional pre-processing or post-processing steps independent from the reconstruction method, and the consistency of the resulting tomograms with the original projection data is lost after the modifications. We propose a GPU accelerated optimization-based reconstruction framework using proximal algorithms. Our framework integrates denoising in the reconstruction process by alternating between reconstruction and denoising, relieving the users of the need to select additional denoising algorithms and preserving the consistency between final tomograms and projection data. Thanks to the flexibility provided by proximal algorithms, various available proximal operators can be interchanged for each task, e.g., various algebraic reconstruction methods and denoising techniques. We evaluate our approach qualitatively by comparison with current reconstruction and denoising approaches, showing excellent denoising capabilities and superior visual quality of the reconstructed tomograms. We quantitatively evaluate the methods with a recently proposed synthetic dataset for scanning transmission electron microscopy, achieving superior reconstruction quality for a noisy and angle-limited synthetic dataset.

With the emergence of new imaging techniques, researchers are always eager to push the boundaries by examining objects either smaller or further away than what was previously possible. The development of image analysis techniques has greatly helped to introduce objectivity and coherence in measurements and decision making. It has become an essential tool for facilitating both large-scale quantitative studies and qualitative research. In this Thesis, methods were developed for analysis of low-resolution (in respect to the size of the imaged objects) three-dimensional (3D) images with low signal-to-noise ratios (SNR) applied to images from cryo-electron tomography (cryo-ET) and fluorescence microscopy (FM). The main focus is on methods of low complexity, that take into account both grey-level and shape information, to facilitate large-scale studies. Methods were developed to localise and represent complex macromolecules in images from cryo-ET. The methods were applied to Immunoglobulin G (IgG) antibodies and MET proteins. The low resolution and low SNR required that grey-level information was utilised to create fuzzy representations of the macromolecules. To extract structural properties, a method was developed to use grey-level-based distance measures to facilitate decomposition of the fuzzy representations into sub-domains. The structural properties of the MET protein were analysed by developing a analytical curve representation of its stalk. To facilitate large-scale analysis of structural properties of nerve cells, a method for tracing neurites in FM images using local path-finding was developed. Both theoretical and implementational details of computationally heavy approaches were examined to keep the time complexity low in the developed methods. Grey-weighted distance definitions and various aspects of their implementations were examined in detail to form guidelines on which definition to use in which setting and which implementation is the fastest. Heuristics were developed to speed up computations when calculating grey-weighted distances between two points. The methods were evaluated on both real and synthetic data and the results show that the methods provide a step towards facilitating large-scale studies of images from both cryo-ET and FM.

Bone response is one of the key determining factors in the overall success of biomaterials intended for bone regeneration and osseointegration. Understanding the formation of bone at an implant surface may lead to the improved design of biomaterials for the future. However, due to the inhomogeneity of bone tissue at an interface, two-dimensional images often lack detail on the interfacial complexity. Furthermore, the increasing use of nanotechnology in the design and production of biomaterials demands characterization techniques on a similar nano length scale. While current analysis methods, such as X-ray tomography, transmission electron microscopy, focused ion beam microscopy and scanning electron microscopy, provide a basis for analysing biomaterials and biointerfaces, they are incapable of doing so with both nanometre resolution and three-dimensional clarity. In contrast, electron tomography may be used to characterize the three-dimensional structure of biomaterials and their interfaces to bone with nanometre resolution. In this work, hydroxyapatite scaffolds, and laser-modified titanium and Ti6Al4V implants were studied in contact with human or rabbit bone. Z-contrast electron tomography revealed that the orientation of collagen in bone apposing hydroxyapatite, titanium and Ti6Al4V implants is consistently parallel to the implant surface, where the bioactive layer that precipitates on HA is oriented perpendicular to the implant surface. With this method, complete three-dimensional nanoscale osseointegration of titanium-based implants was also established. The extension of this technique from interfacial analyses to the design of biomaterials provided an understanding of the pore structure of mesoporous titania. In further investigations, the open three-dimensional pore network, as revealed by electron tomography, showed promise as a coating that improves implant osseointegration and enables site-specific drug-delivery from an implant surface. In summary, it was demonstrated that two-dimensional characterization techniques were insufficient for the investigation of nanostructured biomaterials, as well as their interfaces to bone. Visualizing biointerfaces and biomaterials with nanometre precision in three-dimensions can expose new fundamental information on materials properties and bone response, enabling better design of biomaterials for the future.

abstract: The remarkable conservation of molecular and intra-/inter-cellular pathways underpinning the fundamental aspects of sugar partitioning in two evolutionarily divergent organisms – a non-vascular moss Physcomitrella patens and a vascular cereal crop Oryza sativa (rice) – forms the basis of this manuscript. Much of our current knowledge pertaining to sugar partitioning in plants mainly comes from studies in thale cress, Arabidopsis thaliana, but how photosynthetic sugar is loaded into the phloem in a crop as important as rice is still debated. Even less is known about the mechanistic aspects of sugar movement in mosses. In plants, sugar either moves passively via intercellular channels called plasmodesmata, or through the cell wall spaces in an energy-consuming process. As such, I first investigated the structure of plasmodesmata in rice leaf minor vein using electron tomography to create as of yet unreported 3D models of these channels in both simple and branched conformations. Contrary to generally held belief, I report two different 3D morphotypes of simple plasmodesmata in rice. Furthermore, the complementary body of evidence in arabidopsis implicates plasma membrane localized Proton Pyrophosphatase (H+-PPase) in the energy-dependent movement of sugar. Within this wider purview, I studied the in situ ultrastructural localization patterns of H+-PPase orthologs in high-pressure frozen tissues of rice and physcomitrella. Were H+-PPases neo-functionalized in the vascular tissues of higher plants? Or are there evolutionarily conserved roles of this protein that transcend the phylogenetic diversity of land plants? I show that H+-PPases are distinctly expressed in the actively growing regions of both rice and physcomitrella. As expected, H+-PPases were also localized in the vascular tissues of rice. But surprisingly, H+-PPase orthologs were also prominently expressed at the gametophyte-sporophyte junction of physcomitrella. Upon immunogold labeling, H+-PPases were found to be predominantly localized at the plasma membrane of the phloem complexes of rice source leaves, and both the vacuoles and plasma membrane of the transfer cells in the physcomitrella haustorium, linking H+-PPases in active sucrose loading in both plants. As such, these findings suggest that the localization and presumably the function of H+-PPases are conserved throughout the evolutionary history of land plants. / Dissertation/Thesis / 3D MODEL OF SIMPLE PLASMODESMATA / 3D MODEL OF COMPLEX PLASMODESMATA / MODELING SIMPLE PLASMODESMATA IN IMOD / MODELING COMPLEX PLASMODESMATA IN IMOD / Doctoral Dissertation Biology 2016

The ‘missing wedge’ is a long-standing problem in electron tomography, caused by the use of slab-like flat specimens, which increase in thickness when tilted to high angles. Attempts have been made to reduce the undesirable effects caused by the missing wedge, but the problem remains, particularly for the radiation-sensitive frozen-hydrated specimens used for high resolution imaging. Specimens with cylindrical symmetry offer a way to overcome this problem, since the thickness remains constant at all viewing angles. However, while this has been suggested before, it has never been demonstrated for frozen-hydrated specimens. In this work, I present a way to make cylindrical specimens for electron cryo-tomography, using thin-walled carbon tubes as specimen holders. The tubes are made in a multi-step process, involving carbon deposition on glass micropipette templates and subsequent removal of the glass. Tube diameters are typically a few hundred nanometres, with a wall thickness of 10–20 nm. To make frozen-hydrated specimens, the tubes are filled with an aqueous sample and then plunge-frozen in liquid ethane. Electron images acquired from the tubes have equal quality at all viewing angles, with a tilt range restricted only by the physical limits of the microscope. Tomograms from specimens such as gold particles and liposomes show that the effects of the missing wedge are substantially reduced, with much improved resolution along the electron beam axis. Structural features oriented in all directions are visible in the reconstructions, in marked contrast to tomograms acquired over a more restricted angular range. These results are promising, however some technical challenges remain before this method can be used routinely.