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Development of Biophysical Interpretation of Quantitative Phase Image Data
Křížová, Aneta ; Jákl, Petr (referee) ; Vomastek, Tomáš (referee) ; Chmelík, Radim (advisor)
This doctoral thesis deals with biophysical interpretation of quantitative phase imaging (QPI) gained with coherence-controlled holographic microscope (CCHM). In the first part methods evaluating information from QPI such as analysis of shape and dynamical characteristics of segmented objects as well as evaluation of the phase information itself are described. In addition, a method of dynamic phase differences (DPD) is designed to allow more detailed monitoring of cell mass translocations. All of these methods are used in biological applications. In an extensive study of various types of cell death, QPI information is compared with flow cytometry data, and preferably a combination of QPI and fluorescence microscopy is used. The DPD method is used to study mass translocations inside the cell during osmotic events. The simplified DPD method is applied to investigate the mechanism of tumor cell movement in collagen gels.
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Coherence Controlled Holographic Microscope with the digital optics
Vavřinová, Jana ; Jákl, Petr (referee) ; Dostál, Zbyněk (advisor)
The Digital Micromirror Device (DMD) technology has been developed especially for Digital Light Processing projectors, which allow the image projection. After this succesful implementation, and thanks to the commercial availibility and low initial cost of the DMD chip, a wide range of other applications became possible. Besides, it may be used in microscopy as a spatial light modulator. For example in Coherence-Controlled Holographic Microscope (CCHM) that finds its use especially for imaging and measurement of live-cell dynamic processes. The DMD chip placed in the illumination part of CCHM allows for broadening the application possibilities. Namely it could be different illumination mode experiments or tomographic applications. The master's thesis deals with the optical design of CCHM with digital optics, i. e. DMD chip. The selection of optical elements for CCHM, the experimental verification of the imaging setup and the process of designing the illumination part are described in detail. In the end, the analysis of different designs for illumination setup with the digital optics in object arm is carried out and the results are compared.
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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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Imaging via multimode optical fiber: recovery of a transmission matrix using internal references
Šiler, Martin ; Jákl, Petr ; Traegaardh, Johanna ; Ježek, Jan ; Uhlířová, Hana ; Tučková, Tereza ; Zemánek, Pavel ; Čižmár, Tomáš
Current research of life shows a great desire to study the mechanics of biological processes\ndirectly within the complexity of living organisms. However, majority of practical techniques\nused nowadays for tissue visualization can only reach depths of a few tens of micrometres as\nthe issue obscures deep imaging due to the random light scattering. Several imaging\ntechniques deal with this problems from different angels, such as optical coherence\ntomography, light sheet microscopy or structured light illumination A different and promising strategy to overcome the turbid nature of scattering tissues is to employ multimode optical fibers (MMF) as minimally invasive light guides or endoscopes to provide optical access inside. Although the theoretical description of light propagation through such fibers has been developed a long time ago it is frequently considered inadequate to describe real MMF. The inherent randomization of light propagating through MMFs is typically attributed to undetectable deviations from the ideal fiber structure. It is a commonly believed that this\nadditional chaos is unpredictable and that its influence grows with the length of the fiber.\nDespite this, light transport through MMFs remains deterministic and can be characterized by a transmission matrix (TM) which connects the intensity and phase patterns on the fiber input and output facets. Once the TM is known it can be used to create focus in any desired 3D\ncoordinates beyond the distal fiber facet, see figure 1, and perform e.g. fluorescence based\nlaser scanning microscopy or optical trapping.
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Orbital motion from optical spin: the extraordinary momentum of circularly polarized light beams
Svak, Vojtěch ; Brzobohatý, Oto ; Šiler, Martin ; Jákl, Petr ; Zemánek, Pavel ; Simpson, Stephen Hugh
We provide a vivid demonstration of the mechanical effect of transverse spin momentum in an\noptical beam in free space. This component of the Poynting momentum was previously thought\nto be virtual, and unmeasurable. Here, its effect is revealed in the inertial motion of a probe\nparticle in a circularly polarized Gaussian trap, in vacuum. Transverse spin forces combine with\nthermal fluctuations to induce a striking range of non-equilibrium phenomena. With increasing\nbeam power we observe (i) growing departures from energy equipartition, (ii) the formation of\ncoherent, thermally excited orbits and, ultimately, (iii) the ejection of the particle from the trap.\nOur results complement and corroborate recent measurements of spin momentum in evanescent\nwaves, and extend them to a new geometry, in free space. In doing so, we exhibit fundamental,\ngeneric features of the mechanical interaction of circularly polarized light with matter. The work\nalso shows how observations of the under-damped motion of probe particles can provide detailed\ninformation about the nature and morphology of momentum flows in arbitrarily structured light\nfields as well as providing a test bed for elementary non-equilibrium statistical mechanics.
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Automated Procedures for Coherence Controlled Holographic Microscope
Dostál, Zbyněk ; Štarha, Pavel (referee) ; Jákl, Petr (referee) ; Chmelík, Radim (advisor)
Coherence-Controlled Holographic Microscope (CCHM) and a Fluorescence Holographic Microscope (FHM) were developed particularly for quantitative phase imaging and measurement of live cell dynamics, which used to be a subject of digital holographic microscopy (DHM). CCHM and FHM in low-coherence mode extend capabilities of DHM in the study of living cells. However, this advantage following from the use of low coherence is accompanied by increased sensitivity of the system to its correct alignment. Therefore, the introduction of an automatic self-correcting system is inevitable. Accordingly, in the thesis, the theory of a suitable control system is derived and the design of an automated alignment system for both microscopes is proposed and experimentally proved. The holographic signal was identified as a significant variable for guiding the alignment procedures. On this basis the original basic realignment algorithms were proposed, which encompasses the processes for initial and advanced alignment as well as for long-term maintenance of the microscope aligned state. Automated procedures were implemented in both microscopes unique set of robotic mechanisms designed and built within the frame of the thesis work. All of the procedures described in the thesis were in real experimentally proved at real microscopes in the experimental biophotonics laboratory. In addition, the control software, which contains the needed automated procedures, was developed for FHM.
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Correlation and Spiral Microscopy using a Spatial Light Modulation
Bouchal, Petr ; Čižmár,, Tomáš (referee) ; Jákl, Petr (referee) ; Petráček, Jiří (advisor)
Dizertační práce je uceleným shrnutím výsledků dosažených v průběhu doktorského studia. V úvodní části práce je představena motivace, odborné a technické zázemí a grantová podpora realizovaného výzkumu. Popsány jsou také dosažené výsledky a jejich význam pro skupinu Experimentální biofotoniky, Ústavu fyzikálního inženýrství, Vysokého učení technického v Brně. Vědecká část práce je rozdělena do dvou hlavních bloků, které se postupně zabývají návrhem nových zobrazovacích koncepcí a technickou modifikací stávajících zobrazovacích systému v praktických aplikacích. Dosažené vědecké výsledky podporují vývoj v oblastech korelační a spirální mikroskopie s prostorovou modulací světla. V části zabývající se návrhem nových zobrazovacích koncepcí je provedena studie korelačního zobrazení v podmínkách proměnné časové a prostorové koherence. Následně jsou zkoumány možnosti praktického využití vírových a nedifrakčních svazků v oblastech korelační, holografické a optické mikroskopie. Interference vírových svazků a samozobrazení nedifrakčních svazků je postupně využito k dosažení 3D zobrazení s hranovým kontrastem a rotující bodovou rozptylovou funkcí. Pokročilé zobrazovací metody jsou úspěšně zavedeny optickou cestou ale i digitální modifikací holografických záznamů. Výsledky teoretických modelů a numerických simulací jsou doprovázeny praktickým vyhodnocením navržených zobrazovacích principů. V technicky zaměřené části jsou navrženy nové způsoby zavedení prostorové modulace světla, které umožňují rozšíření zorného pole v experimentech korelačního zobrazení a dosažení achromatizace při zobrazení pomocí programovatelných difraktivních prvků. Rozšíření zorného pole v korelačních experimentech umožňuje přizpůsobovací optický systém vložený do standardní zobrazovací sestavy. Achromatizace difraktivního zobrazení je zajištěna použitím speciálně navrženého refraktivního korektoru. V navazující části je navržena nová metoda krokování fáze, která pracuje s dvojlomností kapalných krystalů využívaných v systémech pro prostorovou modulaci světla. Použití metody je experimentálně demonstrováno v polarizačně modifikovaném Mirau interferometru. Získané technické zkušenosti jsou využity v praktickém návrhu a realizaci multimodálního zobrazovacího systému s prostorovou modulací světla.
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