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Optimization of optical tweezers for multimodal holographic microscope
Vaverka, Jan ; Antoš, Martin (referee) ; Dostál, Zbyněk (advisor)
This bachelor's thesis has described the principle of operation of interference microscopes with closer analysis of multimodal holograpfic microscope (MHM), which has been constructed at the Institut of Physical Engeneering (IPE) of Brno University of Technology. It has also analysed principles of optical trapping and has closely descirebed force effects on the trapped object with characteristic lenght bigger than wavelenght of trapping beam. It has designed optimization of existing optical tweezers module so that the module is connectable to the prototype of MHM, located at IPE. Simulation and optimization of the optical assembly of module have been performed, the mechanical design of module with the drawings has been designed. The module has been entered into production.
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Features for the analysis and classification of cells in holographic microscope images
Navrátilová, Markéta ; Kolář, Radim (referee) ; Vičar, Tomáš (advisor)
This thesis deals with features used for analysis and classification of cell images captured by holographic microscope. Distinctive features are described together with tools for their classification. Features are extracted on provided segmented cells with use of Matlab programming environment. Based on extracted features the cells are classified by SVM classificator. With use of clustering methods and dimensionality reduction different cell types are analyzed. Reliabity of each feature is tested.
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Confocal module for the Coherence Controlled Holographic Microscope
Kubátová, Eva ; Kozubek, Michal (referee) ; Dostál, Zbyněk (advisor)
The Coherence Controlled Holographic Microscope (CCHM) was developed at BUT Brno for a quantitative phase imaging of living cells. Nowadays it ocurres that its imaging properties are enhanced by the use of additional modules. In the present the microscope is equipped with the epifluorescence module, which allows observation of fluorescently marked living cells. This thesis is going to follow up on the development of this module and is going to extend its options by confocal imaging. The disadvantage of current multi-channel confocal microscopes is a mechanical rotation of the Nipkow discs, which causes undesired mechanical vibrations. That is why in this thesis it is replaced by Digital Micromirror Device. With its use was developed optical system of the whole confocal model, whose correct funcion was simulated in optical CAD. The experimentally verified prototype serves to test the imaging properties. On this basis is designed an application idea of the fluorescence confocal module, which will be possible to connect to the CCHM microscope.
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The adjusting collimator for the Fluorescent holographic microscope
Hlaváčová, Kateřina ; Jákl, Petr (referee) ; Dostál, Zbyněk (advisor)
For the proper function of the Fluorescence olographic microscope, it is necessary to adjust all the optical components of the microscope. Furthermore, the precise adjustment is the very critical condition for proper imaging of the Coherence-controlled holographic microscope. Therefore, it is necessary to create a sight collimator for these microscopes for their adjustment. The fluorescence holographic microscope is based on an interference and holographic principles, whose history is mentioned in the theoretical part of the thesis. The existing state of the art of laser sight collimators and their use in practice is also mentioned. The optical and mechanical design of the laser sight collimator and its realization are described in the next part of the thesis. The software for detecting the black sight cross was created for the use of the laser sight collimator in practice. The software is necessary to evaluate the correctness of the alignment of the adjusted microscope. The descriptions of the adjustment procedures for the laser sight collimator and for the Fluorescence holographic microscope are mentioned in the last part of the thesis. These procedures are necessary for proper manipulation and use with the proposed laser sight collimator.
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Measurement of the distribution of an equivalent of cell dry mass using quantitative phase contrast of the CCHM
Kovářová, Klára ; Čolláková, Jana (referee) ; Křížová, Aneta (advisor)
Subject of this bachelor's thesis is to create an overview of conversions of phase profile to the values of dry mass and to realize experiments verifying the importance of dry mass. In the theoretical part, the history of the holographic microscopy and the development of the holographic microscopy on IPE FME BUT in Brno are summarized. The transmission holographic microscope is also described. This thesis covers its construction description and basic working instructions. The theoretical part also deals with the importance of image phase. Two approaches are presented for the interpretation of the phase delay and there is research on phase conversions to dry cell mass (according to H. G. Davies and M. H. F. Wilkins, G. Popescue, G. Dunn and D. Zicha). In the experimental part cultivation of cells, preparation of observation sample and processing of data are described. This part of thesis is also focused on laboratory experiments. Cells K2 (full name LW13K2) were observed in all experiments. First experiments deal with nutritional deprivation of cells and later with cell growth.
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Coherence-gate assisted three-dimensional imaging by holographic microscope
Maršíková, Barbora ; Heintzmann, Rainer (referee) ; Chmelík, Radim (advisor)
Tato diplomová práce se zabývá výzkumem na téma vlivu prostorové koherence osvětlení. Účelem je určit schopnost osové lokalizace při zobrazení Koherencí řízeným holografickým mikroskopem (CCHM) v závislosti na různé prostorové koherenci světelného zdroje. Osová lokalizace je v tomto případě zkoumána jako kvalita rozlišení drobných detailů trojrozměrného vzorku, umístěných nad sebou. Teorie zobrazení holografickým mikroskopem a teorie rozptylu v nehomogenních prostředích je shrnuta v první části práce, v rozsahu nutném pro pochopení části praktické. Základní princip fungování mikroskopu a přesný popis jeho uspořádání je zde podrobně popsán. Proběhl mechanický návrh stavební úpravy mikroskopu tak, aby bylo možno využívat kondenzorovou optiku s vysokou numerickou aperturou a omezenými optickými vadami. Několik různých přístupů, které by mohly vést ke zlepšení zobrazovacích vlastností mikroskopu, bylo navrženo a vyzkoušeno a jsou zde popsány i s jejich výhodami a nevýhodami. Pro experimentální část práce byl vyroben modelový vzorek. Závislost osové lokalizace na prostorové koherenci osvětlení byla demonstrována pomocí simulace a následně ověřena experimentálně, pozorováním vyrobeného modelového vzorku. Experimentální výsledky potvrzují základní principy vycházející ze zmíněné teorie. Na závěr jsou navržena možná vylepšení, pro budoucí zpřesnění výsledků.
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Fiber guided illumination module
Kropáč, Vlastimil ; Kvasnica, Lukáš (referee) ; Dostál, Zbyněk (advisor)
This diploma thesis describes the design of the illumination system for a Coherence--Controlled Holographic Microscope (CCHM). The theoretical part mentions the history of microscopy, the principle of holography and individual types of interference microscopy. To get closer to the topic, individual light sources and an overview of current illumination systems are mentioned. The diploma thesis also describes the procedure of designing a fiber-optic illumination module for microscopy from optical design through design of construction to the last step, which is assembly and testing of the module.
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Stabilization of the mutual position of the focal plane and the observed object in the holographic microscope
Ha Trinh, Hung Son ; Brada, Michal (referee) ; Dostál, Zbyněk (advisor)
Koherencí řízený holografický mikroskop (CCHM) umožňuje studium nativních biologických vzorků v reálném čase. Při dlouhodobých pozorováních ovšem dochází k rozostřování sledovaných objektů. Tento nežádoucí jev je pravděpodobně způsoben změnami vzájemné polohy předmětové roviny a pozorovaného předmětu. Předpokládá se, že za nestálostí vzájemných poloh stojí tepelné změny uvnitř CCHM. Vznikla proto potřeba vyvinout hardwarový autofokusační systém, který by doplňoval v současnosti implementovaný softwarový autofokus a mohl by vykompenzovat jeho nedostatky. V práci je nejprve stručně popsána optika CCHM. Poté je vysvětlen princip aktuálního řešení doostřování. Posléze je představeno několik metod detekce a monitorování posunutí, kterých lze využít ve zpětnovazebné smyčce k udržování definované vzájemné polohy. Po zvážení výhod a nevýhod je pro praktickou část práce vybrána právě jedna metoda, na základě které je navrženo, sestrojeno a experimentálně otestováno zařízení zajišťující stabilizaci vzájemné polohy předmětové roviny a pozorovaného předmětu.
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Reconstruction methods in holographic microscopy
Findejsová, Anna ; Odstrčilík, Jan (referee) ; Kolář, Radim (advisor)
This master’s thesis focuses on the image reconstruction from holographic microscope. The first part of the thesis summarizes problems of holography, describes its principle and application particularly in live cell biology. In the second part the description of several methods used for off-axis hologram reconstruction is provided. The last part describes the implementation of the basic step in holographic reconstruction – the elimination of autocorrelation and twin image – and then also the reconstruction of 3D information of the sample.
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Influence of imaging sensors properties on the quality of the quantitative phase image reconstruction
Šulc, Václav ; Antoš, Martin (referee) ; Dostál, Zbyněk (advisor)
This bachelor thesis is focused on imaging sensors noise characteristics and their impact on quantitative phase imaging. From available sensors the darkframes were acquired and analyzed. Computed data of noise standard deviation were used to simulate synthetic holograms for reconstruction of quantitative phase images. A camera Basler acA2040-180km was found to be a most suitable device for holography. The design, fabrication and testing of an effective cooling system for such a device proved to have even better noise characteristics.
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