Thesis deals with problematic of raising digital image spacial resolution, mainly thermal image. There are mentioned methods of interpolation, panorama and super-resolution. Main topic of this thesis is super-resolution which is detailly described during the thesis. Finally there is a description of algorithm implementation and problems that may occur during the implemetation.

The demand for high optical resolution has brought researchers to explore the use of beam shaping diffractive optical elements (DOEs) for improving performance of high numerical aperture (NA) optical systems. DOEs can be designed to modulate the amplitude, phase and/or polarization of a laser beam such that it focuses into a targeted irradiance distribution, or point spread function (PSF). The focused PSF can be reshaped in both the transverse focal plane and along the optical axis. Optical lithography, microscopy and direct laser writing are but a few of the many applications in which a properly designed DOE can significantly improve optical performance of the system. Designing DOEs for use in high-NA applications is complicated by electric field depolarization that occurs with tight focusing. The linear polarization of off-axis rays is tilted upon refraction towards the focal point, generating additional transverse and longitudinal polarization components. These additional field components contribute significantly to the shape of the PSF under tight focusing and cannot be neglected as in scalar diffraction theory. The PSF can be modeled more rigorously using the electromagnetic diffraction integrals derived by Wolf, which account for the full vector character of the field. In this work, optimization algorithms based on vector diffraction theory were developed for designing DOEs that reshape the PSF of a 1.4-NA objective lens. The optimization techniques include simple exhaustive search, iterative optimization (Method of Generalized Projections), and evolutionary computation (Particle Swarm Optimization). DOE designs were obtained that can reshape either the transverse PSF or the irradiance distribution along the optical axis. In one example of transverse beam shaping, all polarization components were simultaneously reshaped so their vector addition generates a focused flat-top square irradiance pattern. Other designs were obtained that can be used to narrow the axial irradiance distribution, giving a focused beam that is superresolved relative to the diffraction limit. In addition to theory, experimental studies were undertaken that include (1) fabricating an axially superresolving DOE, (2) incorporating the DOE into the optical setup, (3) imaging the focused PSF, and (4) measuring aberrations in the objective lens to study how these affect performance of the DOE.

Digitization of physical artwork is usually done using image scanning devices in order to ensure that the output is accurate in terms of color and is of sufficiently high resolution, usually over 300 pixels per inch, however the usage of such a device is in some cases unfeasible due to medium or size constraints. Photography of the artwork is another method of artwork digitization, however such methods often produce results containing camera artifacts such as shadows, reflections or low resolution. This thesis project explores the possibility of creating an alternative to image scanners using smartphone photography and machine learning-based methods. Due to the very high memory requirement for enhancing images at very high resolutions, this is done in a two-stage process. The first stage uses an unpaired image style transfer model to remove shadows and highlights. The second stage uses a superresolution model to increase the resolution of the image. The results are evaluated on a small set of paired images using objective metrics and subjective metrics in the form of a user study. In some cases the method removed camera artifacts in the form of reflection and color accuracy, however the best results were achieved when the input data did not contain any major camera artifacts. Based on this it seems likely that style transfer models are not applicable for problems with a wide range of expected input and output. The use of super-resolution seems to be a crucial component of high-resolution image enhancement and the current state-of-the-art methods are able to convincingly increase the resolution of images provided that the input is of a sufficiently high resolution. The subjective evaluation shows that commonly used metrics such as structural similarity and Fréchet Inception Distance are applicable for this type of problem when analyzing the full image, however for smaller details other evaluation methods are required. / Digitalisering av fysiska konstverk görs vanligtvis med bildskannrar för att försäkra att den digitaliserade bilden är färgnoggrann och att upplösningen är tillräckligt hög, vanligtvis över 300 pixlar per tum. Dock är användandet av bildskannrar ibland svårt på grund av konstverkets material eller storlek. Fotografi av konstverk är en annan metod för digitalisering, men denna metod producerar ofta kameraartefakter i form av skuggor, reflektioner och låg upplösning. Detta examensarbete utforskar möjligheten att skapa ett alternativ till bildskannrar genom att använda smartphonefotografi och maskininlärningsbaserade metoder. På grund av de höga minneskraven för bildförbättring med mycket höga upplösningar görs detta i en tvåstegsprocess. Det första steget använder oparad bildstilöversättning för att eliminera skuggor och ljuspunkter. Det andra steget använder en superupplösningsmodell för att öka bildens upplösning. Resultaten utvärderas på en liten mängd parade bilder med objektiva jämförelser och subjektiva jämförelser i form av en användarstudie. I vissa fall reducerade metoden kameraartefakter i form av reflektioner och förbättrade färgexakthet, dock skedde dessa resultat i fall där indatan inte innehöll några större kameraartefakter. Baserat på detta är det sannolikt att stilöversättningsmodeller inte är applicerbara för problem med ett brett omfång av möjliga indata och utdata. Användandet av superupplösning verkar vara en viktig komponent av högupplöst bildförbättring och de bäst presenterande metoderna kan övertygande öka upplösningen av bilder i fall där indatan är av tillräckligt hög upplösning. Den subjektiva utvärderingen visar att vanligt använda utvärderingsmetoder som Fréchet-Inception-avstånd och strukturell likhet är applicerbara för denna typ av problem när de används för att analysera en hel bild, men för mindre detaljer behövs alternativa utvärderingsmetoder.

CD4 co-receptor of main T cell receptor (TCR) is essential for proper development of T lymphocytes and their function in adaptive immune responses. It is believed that CD4 stabilizes the interaction of TCR with antigenic ligand, peptide-MHC, and thereby improves T cell-dependent responses during immune reaction. CD4 is transmembrane glycoprotein with a number of structural motifs in its intracellular domain which do not dramatically affect its sorting to the plasma membrane but can influence its local organization at nanoscale. CD4 was shown to transiently accumulate in the immunological synapse formed between T cell and antigen-presenting cell. Such accumulation is rapidly followed by its internalization and/or delocalization outside the synapse. This is in contrast with TCR which accumulates strongly in the immunological synapse and is later found enriched in the central area of this structure. It is therefore unclear how TCR and its CD4 co-receptor function together when binding to their common ligand during the initiation of signaling in T cells. We aim to study localization of CD4 at nanoscale using advanced fluorescence microscopy techniques achieving significant improvements in resolution. In this work, CD4 and its mutant variants, potentially causing its different localization at the...

La microscopie optique est un outil indispensable pour la recherche de la neurobiologie et médecine qui permet l’étude des cellules dans leur environnement natif. Les processus sous-cellulaires restent néanmoins cachés derrière les limites de la résolution optique, ce qui rend la résolution des structures plus petites que ~300nm impossible. Récemment, les techniques de la localisation des molécules individuelles (SML) ont permis le suivi des protéines de l’échelle nanométrique grâce à l’ajustement des molécules uniques à la réponse impulsionnelle du système optique. Ce processus dépend de la quantité de lumière recueilli et rend ces techniques très sensibles aux imperfections de la voie d’imagerie, nommé des aberrations, qui limitent l’application de SML aux cultures cellulaires sur les lamelles de verre. Un système commercial d’optiques adaptatives est implémenté pour compenser les aberrations du microscope, et un flux de travail est défini pour corriger les aberrations dépendant de la profondeur qui rend la 3D SML possible dans les milieux biologiques complexes. Une nouvelle méthode de SML est présentée qui utilise deux objectifs pour détecter le spectre d’émission des molécules individuelles pour des applications du suivi des particules uniques dans 5 dimensions (x,y,z,t,λ) sans compromis ni de la résolution spatiotemporelle ni du champ de vue. Pour faciliter les analyses de manière quantitative des Go de données générés, le développement des outils biochimiques, numériques et optiques est présenté. Ensemble, ces approches ont le but d’amener l’imagerie quantitative des molécules uniques dans les échantillons biologiques complexes / Optical microscopy is an indispensable tool for research in neurobiology and medicine, enabling studies of cells in their native environment. However, subcellular processes remain hidden behind the resolution limits of diffraction-limited optics which makes structures smaller than ~300nm impossible to resolve. Recently, single molecule localization (SML) and tracking has revolutionized the field, giving nanometer-scale insight into protein organization and dynamics by fitting individual fluorescent molecules to the known point spread function of the optical imaging system. This fitting process depends critically on the amount of collected light and renders SML techniques extremely sensitive to imperfections in the imaging path, called aberrations, that have limited SML to cell cultures on glass coverslips. A commercially available adaptive optics system is implemented to compensate for aberrations inherent to the microscope, and a workflow is defined for depth-dependent aberration correction that enables 3D SML in complex biological environments. A new SML technique is presented that employs a dual-objective approach to detect the emission spectrum of single molecules, enabling 5-dimensional single particle imaging and tracking (x,y,z,t,λ) without compromising spatiotemporal resolution or field of view. These acquisitions generate ~GBs of data, containing a wealth of information about the localization and environment of individual proteins. To facilitate quantitative acquisition and data analysis, the development of biochemical, software and hardware tools are presented. Together, these approaches aim to enable quantitative SML in complex biological samples.

Over the past three decades in cellular neuroscience there has been a shift towards the view of the 'tripartite synapse', where, astrocytes -- as well as the pre-synapse and post-synapse -- are involved in synaptic signalling. The migration of astrocytes to form branched networks in the brain is, therefore, of great interest in understanding brain development and neuronal function. Migration is a complex interplay between cytoskeletal reorganisation and cell mechanical stiffness. In order to improve understanding of this process, correlative measurements of cytoskeletal organisation and mechanical stiffness are required. To investigate astrocyte migration a technique combining atomic force microscopy (AFM) with stimulated emission depletion (STED) microscopy was developed. First a custom STED microscope was developed. To facilitate the design of this system the theoretical performance of a range of STED techniques (cw-STED, time-gated STED, pulsed STED and RESOLFT) were compared, identifying that pulsed STED theoretically has the highest photon efficiency. A pulsed STED microscope, which uses adaptive optics, was then designed, developed and characterised. The microscope was found to achieve resolutions below 50 nm. The STED microscope was combined with a commercial AFM to study live cells. Using the recently developed SiR-actin and SiR-tubulin dyes and AFM probes optimised for live cell mechanical property studies, images of the actin and tubulin cytoskeleton were correlated with AFM topography and mechanical stiffness measurements. It was found that, in astrocytes, actin contributes significantly both to astrocyte stiffness and topography. Investigations of migrating cells showed differences in actin organisation and mechanical stiffness between the basis and leading edge of migration. A further study was performed, investigating the effects of the gap-junction protein connexin30, which is expressed during the early stages of brain development, on migration. This protein was found to inhibit the actin reorganisation and mechanical stiffness changes observed in basal conditions. Overall the combination of mechanosensitive AFM measurements with advanced microscopy, such as super-resolution, on live cells is a promising approach which will enable a range of investigations, for instance when studying cell structural remodeling during brain development or tumorigenesis.

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京都大学 / 新制・課程博士 / 博士(生命科学) / 甲第24983号 / 生博第512号 / 新制||生||68(附属図書館) / 京都大学大学院生命科学研究科統合生命科学専攻 / (主査)教授 荒木 崇, 教授 鈴木 淳, 教授 谷口 雄一 / 学位規則第4条第1項該当 / Doctor of Philosophy in Life Sciences / Kyoto University / DFAM

This thesis exploits so called “Optical Eigenmodes” (OEi) in the focal plane of an optical system. The concept of OEi is introduced and the OEi operator approach is outlined, for which quadratic measures of the light field are expressed as real eigenvalues of an Hermitian operator. As an example, the latter is employed to locally minimise the width of a focal spot. The limitations of implementing these spots with state of the art spatial beam shaping technique are explored and a selected spot with a by 40 % decreased core width is used to confocally scan an in focus pair of holes, delivering a two-point resolution enhanced by a factor of 1.3. As a second application, OEi are utilised for fullfield imaging. Therefore they are projected onto an object and for each mode a complex coupling coefficient describing the light-sample interaction is determined. The superposition of the OEi weighted with these coefficients delivers an image of the object. Compared to a point-by-point scan of the sample with the same number of probes, i.e. scanning points, the OEi image features higher spatial resolution and localisation of object features, rendering OEi imaging a compressive imaging modality. With respect to a raster scan a compression by a factor four is achieved. Compared to ghost imaging as another fullfield imaging method, 2-3 orders of magnitude less probes are required to obtain similar images. The application of OEi for imaging in transmission as well as for fluorescence and (surface enhanced) Raman spectroscopy is demonstrated. Finally, the applicability of the OEi concept for the coherent control of nanostructures is shown. For this, OEi are generated with respect to elements on a nanostructure, such as nanoantennas or nanopads. The OEi can be superimposed in order to generate an illumination of choice, for example to address one or multiple nanoelements with a defined intensity. It is shown that, compared to addressing such elements just with a focussed beam, the OEi concept reduces illumination crosstalk in addressing individual nanoelements by up to 70 %. Furthermore, a fullfield aberration correction is inherent to experimentally determined OEi, hence enabling addressing of nanoelements through turbid media.

No description available.