National Repository of Grey Literature 45 records found  1 - 10nextend  jump to record: Search took 0.01 seconds. 
Changes of intracellular pH in yeast cells under stress conditions
Divín, Radek ; Plášek, Jaromír (advisor) ; Krůšek, Jan (referee)
Title: Changes of intracellular pH in yeast cells under stress conditions Author: Radek Divín Department: Institute of Physics of Charles University Supervisor: prof. RNDr. Jaromír Plášek, CSc. Abstract: Specific values of intracellular pH (pHi) can affect all biochemical processes in a cell and this phenomenon is closely connected with the degree of importance of changes in the intracellular pH under the stress conditions. In the Master Thesis, the yeast cells Saccharomyces cerevisiae were used as a model of organism eukaryotic cells. Monitoring of intracellular pH of the cells was performed by the method of synchronous fluorescence scan technique of genetically encoded fluorescent probes pHluorin which was located in the cytosol of the cells. The cells were exposed to stress conditions due to the chemical changes in the environment. Consequently, their ability to maintain a stable value of the intracellular pH in various acidic environments was studied in more detail. The attention was also focused on the impact on optimizing of glucose cytosolic pH. The work was centered on the changes in intracellular pH under the influence of the presence of KCl in suspension. Furthermore, the decrease of cytosolic pH of monitored cells by protonophore CCCP was investigated. The effect of stress environment on the...
Imaging of fluorescently labelled mitochondria using Biplane_FPALM microscopy
Dostál, Marek ; Plášek, Jaromír (advisor) ; Valenta, Jan (referee)
Title: Imaging of fluorescently labelled mitochondria using Biplane FPALM microscopy Author: Bc. Marek Dostál Department: Fyzikální ústav UK Supervisor: prof. RNDr. Jaromír Plášek, CSc. Consultants: RNDr. Petr Ježek, DrSc. Mgr. Hana Engstová, Ph.D. Abstract: In this thesis the results are presented of a search for suitable PCFP markers for a visualization inner and outer mitochondrial membranes by unique biplane FPALM microscope. We participated in debugging the measuring software for the purpose of 3D vizualization mitochondrial nets and determine their parameters. For this purpose were developed two methods. The first one can be applied to determine the parametrs of outer membranes and the second one is applicable to determine parametrs of inner membranes. On smaller statistical file we managed to confirm dependence quality of mitochondrial net on the cell culture conditions. Under stress conditions e.g. hypoxia the quality of mitochondrial net is changed. Keywords: mitochondria, Biplane FPALM, GFP, 3D visualization.
Bioapplications of novel nanostructured materials
Fučíková, Anna ; Valenta, Jan (advisor) ; Polívka, Tomáš (referee) ; Plášek, Jaromír (referee)
Title: Bioapplications of novel nanostructured materials Author: Anna Fučíková Department / Institute: Department of Chemical Physics and Optics, Faculty of Mathematics and Physics, Charles University in Prague Supervisor of the doctoral thesis: Doc. RNDr. Jan Valenta, Ph.D., Department of Chemical Physics and Optics, Faculty of Mathematics and Physics, Charles University in Prague Abstract: This work is aimed at development of new fluorescent labels based on silicon nanocrystals. Nanodiamonds and commercial CdSe quantum dots have been used as comparative materials. Silicon nanocrystals are relatively small (1-4 nm) compared to other studied nanomaterials. They are prepared by electro-chemical etching and their surface can be activated by various molecules which strongly influences luminescence properties. Luminescence quantum efficiency can be as high as 30 % and perfectly photostable even in biological environment. Si nanocrystals are biodegradable in a living organism within reasonable time scale and non-toxic. We are able to detect luminescence of single nanocrystals, even inside living cells, with use of our micro-spectroscopy apparatus. Nanodiamonds have weak luminescence; they are toxic at higher dosages and very stable in living bodies (without available technique how to remove them). Studied CdSe...
Comprehensive characterization of cellular suspensions by means of elastic light scattering
Macháň, Radek ; Štěpánek, Josef (advisor) ; Plášek, Jaromír (referee)
Studies of yeast Saccharomyces cerevisiae - a model eukaryot organism - often require determining the number of cells in a suspension. One of the methods often used for studying cell suspensions is the measuring of elastic scattering of light passing through the suspensions. Usual output of those measurements is the angular dependency of the intensity of scattered light, which bears information about the size of scatterers. This thesis studies time-dependencies of the intensity of scattered light and relationship of their statistical properties to the number of scatterers in a unit volume. For this purpose a simple model was proposed, which describes the connection between this quantity for a monodisperse suspension and the dependency of standard deviation of time-behaviour of the scattered intensity on the mean value of the same time-behaviour. An apparatus for computer controlled measurements of angular and timedependencies of scattered light intensity was completed for carrying out the experiments. Elastic scattering was measured on suspensions of yeast Saccharomyces cerevisiae and a model system (suspensions of 8m polymer particles). Experimental data were interpreted within the proposed model. Although yeast suspensions cannot be regarded as monodisperse, experimental data were in a good agreement with...
The comparison of the performace of selected carbocyanine dyes in fluorescent probing of yeast cell membrane potential.
Mudroňová, Kateřina ; Plášek, Jaromír (advisor) ; Krůšek, Jan (referee)
The membrane potential is one of the most important parameters of the living cell. It can be measured using carbocyanine fluorescent probes. In this thesis we examined parameters of several dyes of this family. For further experiments three of them were chosen - diOC3(3), diIC1(3) a diIC2(5) as a supplement to diSC3(3) and diSC3(5), which represent standard probes used at biophysical department of Institut of Physics. We compared the rates of their accumulation in S. cerevisiae cells to determine if they were MDR pumps' substrates. The other goal of this work was to decide whether the results obtained using different probes are equivalent and to determine if the presence of a probe affects the spectral characteristics of another. For this purpose we have chosen diSC3(3) and diSC3(5). With those dyes we examined the influence of the acidification on membrane potencial of the yeast S. cerevisiae. We showed that the information on depolarization obtained using both probes were matching very well.
The monitoring of intracellular ion concentrations in microbial cells
Vodáková, Adéla ; Plášek, Jaromír (advisor) ; Krůšek, Jan (referee)
The Master Thesis focuses on monitoring of intracellular ion concentrations in bacteria Escherichia coli and yeast Saccharomyces cerevisiae using genetically encoded fluorescent probes with green fluorescent protein (GFP). Aquired knowledge about this protein and its spectral characteristics is summarized in the introduction. For experimental study a pH-sensitive sensor which displays a ratio change of two excitation fluorescence peaks - pHluorin - was chosen. This probe was tested in bacteria and yeast cells. The experiments concentrated on the ability of the cell to maintain a constant cytosolic pH under various conditions like different pH values of the suspension, addition of glucose or KCl to the suspension. Another topic discussed in the thesis is the elimination of the cell autofluorescence from the GFP signal. For this purpose the synchronous fluorescence scan technique was succesfully used. I have found out that by using this method the measurements of cytosolic pH values are even more accurate thanks to the improved signal to noise ratio.
Deconvolution fluorescence microscopy of yeast cells
Štec, Tomáš ; Plášek, Jaromír (advisor) ; Heřman, Petr (referee)
Title: Deconvolution fluorescence microscopy of yeast cells Author: Tomáš Štec Department: Institute of Physics of Charles University Supervisor: prof. RNDr. Jarmoír Plášek, CSc., Institute of Physics of Charles Uni- versity Abstract: Fluorescence microscopy presents an fast and cheap alternative to more advanced imaging methods like confocal and electron microscopy, even though it is subject to heavy image distortion. It is possible to recover most of the original distortion-free image using deconvolution in computer image processing. This al- lows reconstruction of 3D structure of studied objects. Deconvolution procedure of NIS Elements AR program undergoes an thorough inspection in this diploma the- sis. It is then applied on restoration of 3D structure of calcofluor stained cell wall of budding yeast Saccharomyces cerevisiae. Changes of the structure of the cell wall during cell ageing are being examined. Cell wall of aged cells shows increased surface roughness and even ruptures at the end of cell life. Keywords: fluorescence, microscopy, deconvolution, NIS Elements AR, calcofluor, yeast, cell wall, ageing
The comparison of the performace of carbocyanine dyes disC3(3) a diSC3(5) in fluorescent probing of yeast cell membrane potential.
Matunová, Petra ; Plášek, Jaromír (advisor) ; Krůšek, Jan (referee)
Membrane potential represents a voltage across a membrane and it is an important parameter that helps to describe processes in cells. Carbocyanine fluorescent probes diS-C3(3) and diS-C3(5), for which a common short chemical name 3,3'- dipropylthiadicarbocyanine iodide is used, are suitable for monitoring membrane potential changes of cells in which microelectrodes can not be used because of a small size of the cells. These changes can be measured on the scale of mV. A spectral analysis of cell suspensions containing a fluorescent probe makes it possible to determine the ratio of extracellular and intracellular concentrations of the probe. Using it we can calculate the value of membrane potential changes which can be induced by an outer stimulus. This Bc. thesis presents a comparison of the rate of accumulation of the above mentioned fluorescent probes in yeast cells, as well as experiments aimed for studying an inuence of different substances and their various concentrations on free and bound component of the dye.

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