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Detection of cells in confocal microscopy images
Hubálek, Michal ; Štursa, Dominik (referee) ; Škrabánek, Pavel (advisor)
The goal of the thesis was to create an application that automatically detects healthy cardiomyocytes from images captured by a confocal microscope. The thesis was created based on the specific needs of researchers from the Slovak Academy of Sciences.The application will facilitate and increase the efficiency of their research,because until now they have to evaluate the images and search for suitable cells manually. The RetinaNet convolutional neural network is used for detection and has been implemented in a user-friendly desktop application. The application also automatically records and stores coordinates of detected cells which can be used for capturing cells in higher image quality. Another advantage of the developed application is its versatility, which allows to train detection on other data, making it applicable to other projects. The result of this work is a functional, standalone and intuitive application that is ready to be used by researchers.
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Application of carbon based electrodes for electrochemical detection in flow system
Mika, Jan ; Dejmková, Hana (advisor) ; Fischer, Jan (referee)
Carbon is due to its electrochemical properties a favourite material for construction of electrodes suitable for detection in flow system. The two most often used techniques (coulometric and amperometric) will be discussed in this Thesis. Carbon paste electrodes (CPE) represent the amperometric detectors. CPE are unstable in the medium with high content of organic solvents, which might complicate their application in HPLC, where the mobile phase contains organic component. It is interesting, that glassy carbon paste electrodes are more stable under such conditions. The change of surface and response of the electrode caused by the effect of methanol are described in this Thesis along with the possible reason of stability of GCPE. Roughing of the surface exposed to methanol is demonstrated by confocal microscopy. This effect leads to increase of height of the peak, background current and noise and to the shift of peak potential to less positive values. These changes are dependent on the time of exposure and the concentration of methanol. Coulometric detectors work with high conversion effectiveness and sensitivity, but in case of their passivation, reactivation is a relatively complicated problem, often leading to the replacement of electrode material. Answer to this problem can be given by using...
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Actuoecology of testate amoebae in fresh water and soil environment in enteraction with fungi and their analysis with new microscopic techniques
Burdíková, Zuzana ; Holcová, Katarína (advisor) ; Jansa, Jan (referee) ; Hudáčková-Hlavatá, Natália (referee)
4 Abstract The present thesis focuses on testate amoebae (TA) and their relationship to their natural environment, as well as on relevant microscopic imaging methods. The bulk of the data has been published in original scientific papers and is compiled into three separate chapters (Pt I, Pt II and Pt III), each annotated by a brief introduction. (Pt I) The methods section is devoted to specialized microscopic techniques employed to broaden the scope of the ecological analyses. In particular, precise discrimination between live and dead individuals, biomass determination inside individual tests and a multi-modal visualization of the cytoplasm and organelles enhance the data. Laser scanning confocal microscopy and two-photon microscopy are the main imaging modalities employed to study TA morphology in detail. The data have implications for taxonomy and ecophysiology, including the use of TA as bioindicators of pollution. (Pt II) An actuoecological analysis focuses on the seasonal variability of TA species composition in a freshwater ecosystem, namely the Komo any ponds in Prague, during the course of the year. The species composition variation is correlated to simultaneously recorded limnological parameters such as temperature, pH, contamination by (heavy) metals (As, Cd, Mn, Ni, Fe, Pb), polycyclic aromatic...
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Study of quantitative parameters of Norway spruce needle structure under the effect of elevated CO₂ concentration and different irradiance
Kubínová, Zuzana ; Lhotáková, Zuzana (advisor) ; Urban, Otmar (referee) ; Gebauer, Roman (referee)
Zuzana Kubínová: Study of Quantitative Parameters of Norway Spruce Needle Structure under the Effect of Elevated CO2 Concentration and Different Irradiance, Doctoral Thesis, Prague 2019 Abstract Atmospheric concentration of CO2 is increasing, while its influence on plants is still not fully elucidated. Norway spruce (Picea abies L. Karst.) is an abundant conifer tree in European temperate and boreal forests, which behave as carbon sink in the global carbon cycle. The physiological response to elevated CO2 concentration may be interconnected with changes in leaf anatomy and morphology. Needle structure is also determined by other factors in addition to CO2 concentration, irradiance being the most important one. Thus, effect of irradiance was also included in our studies. The effects of elevated CO2 concentration and irradiance on Norway spruce needle structure were studied using new applications of well-established quantitative methods and novel methods enabling effective and unbiased analysis of needle structural traits. The General Procrustes analysis showed to be effective for needle shape on cross section comparison and the disector method proved to be suitable for chloroplast number estimates. The influence of elevated CO2 concentration and different irradiance on needle structure was studied at two...
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Mechanisms of invasiveness and transcription regulation in cancer cells
Tolde, Ondřej ; Folk, Petr (advisor) ; Kovář, Jan (referee) ; Brdička, Tomáš (referee)
The mechanisms of invazivity and regulation of transcription of cancer cells Cancer originates in cells that overcome the control mechanisms of the organism. Cancer cells can be eventually released from the site of origin and spread through tissues. Cancer cells can acquire certain mechanisms that enable them to more effectively invade surrounding tissue or layers of other cells. The research on the migration of cancer cells is important for the understanding of the origin and spreading of metastases and consequently for anticancer therapy. In my Ph.D. work, I participated in the research of the properties of invasive metastatic cells. We compared non-invasive rat sarcoma cell line with a higly metastatic cell line derived from it. We showed that cells of the invasive cell line use amoeboid mode of migration, have upregulated Rho/ROCK signaling, and have accumulated actin and myosin at the leading edge. It is at the leading edge where the cells generate their traction forces. Cells of non-invasive cell line use mesenchymal mode of migration and generate forces mainly at their retracting end. We also compared two breast cancer cell lines derived from a single carcinoma. We showed that the more invasive cell line, derived from its parental line by neoplastic transformation, displayed elevated cytoskeletal...
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Determination of the area ratio of tubular and surface membranes in cardiomyocytes
Hluší, Veronika ; Čmiel, Vratislav (referee) ; Chmelíková, Larisa (advisor)
This work contains a literature search regarding the function and structure of the tubular system of cardiomyocytes. Another theory includes the basic principle of fluorescence and a description of the fluorescence and confocal microscope. Next chapter describe the principle of isolation and subsequent staining of cardiomyocytes to obtain images of the tubular system is described. Next step is the implementation of a method for segmentation of the membrane and tubular system of cardiac cells and the application of this method to the obtained data.
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Focus techniques of optical measurement of 3D features
Macháček, Jan ; Honec, Peter (referee) ; Janáková, Ilona (advisor)
This thesis deals with optical distance measurement and 3D scene measurement using focusing techniques with focus on confocal microscopy, depth from focus and depth from defocus. Theoretical part of the thesis is about different approaches to depth map generation and also about micro image defocusing technique for measuring refractive index of transparent materials. Then the camera calibration for focused techniques is described. In the next part of the thesis is described experimentally verification of depth from focus and depth from defocus techniques. For the first technique are shown results of depth map generation and for the second technique is shown comparison between measured distance values and real distance values. Finally, the discussed techniques are compared and evaluated.
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Visualization of scalar fields by back-to-front method
Gurecká, Hana ; Druckmüller, Miloslav (referee) ; Martišek, Dalibor (advisor)
Diplomová práce je zaměřena na metody zobrazování skalárních dat v pevné datové mřížce, konkrétně dat získaných užitím fluorescenčního konfokálního mikroskopu. Teoretická část textu začíná představením fungování konfokálních mikroskopů a zasazení problematiky zkoumaných grafických metod do matematického kontextu. Následující kapitola se věnuje odvození integrálu pro zobrazování objemů a z něj vyplývající back-to-front metodu. Teoretická část je zakončena představením metod vhodných pro zobrazování trojrozměrných skalárních dat při použití back-to-front algoritmu. V praktické části je pak popsán implementovaný algoritmus.
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Study of mesenchymal stem cell migration based on principles of chemotaxis
Pošustová, Veronika ; Skopalík, Josef (referee) ; Chmelíková, Larisa (advisor)
The purpose of this Master thesis is to verify migration of mesenchymal stem cells on the principle known as chemotaxis. First part of this study is focused on cell migration in order to explain the whole migration process. Next part describes various chemotaxis methods and selected studies dealing with clinical applications of mesenchymal stem cells in different medical and biomedical fields. The following step describes confocal microscopy, which is used for acquiring images of the cells. The experimental part is focused on cultivation of mesenchymal stem cells in a laboratory, which is necessary for cell vitality. Furthermore, there are designed two main experiments. Firstly there is a 2D experiment with adherent cells for chemotaxis using -Slide Chemotaxis. Secondly Transwell migration test is designed and executed. Finally, the acquired images from confocal microscope are used for image processing, which was done in Matlab R2020a programming environment. The result of this processing is evaluation of cell confluence and migration. In the end, experimental part of this study was optimized according to recommended studies. The results are summarized in the conclusion with proposal for improvements of those methods.
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Study of quantitative parameters of Norway spruce needle structure under the effect of elevated CO₂ concentration and different irradiance
Kubínová, Zuzana ; Lhotáková, Zuzana (advisor) ; Urban, Otmar (referee) ; Gebauer, Roman (referee)
Zuzana Kubínová: Study of Quantitative Parameters of Norway Spruce Needle Structure under the Effect of Elevated CO2 Concentration and Different Irradiance, Doctoral Thesis, Prague 2019 Abstract Atmospheric concentration of CO2 is increasing, while its influence on plants is still not fully elucidated. Norway spruce (Picea abies L. Karst.) is an abundant conifer tree in European temperate and boreal forests, which behave as carbon sink in the global carbon cycle. The physiological response to elevated CO2 concentration may be interconnected with changes in leaf anatomy and morphology. Needle structure is also determined by other factors in addition to CO2 concentration, irradiance being the most important one. Thus, effect of irradiance was also included in our studies. The effects of elevated CO2 concentration and irradiance on Norway spruce needle structure were studied using new applications of well-established quantitative methods and novel methods enabling effective and unbiased analysis of needle structural traits. The General Procrustes analysis showed to be effective for needle shape on cross section comparison and the disector method proved to be suitable for chloroplast number estimates. The influence of elevated CO2 concentration and different irradiance on needle structure was studied at two...
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