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Membrane potential measurement with voltage sensitive dyes
Votavová, Barbora ; Janoušek, Oto (referee) ; Čmiel, Vratislav (advisor)
The aim of this work is to realize measurements of membrane potential with voltagesensitive dye Di-4-ANEPPS and the data processed and analyzed. The work includes theoretical basis in the form of electrophysiology animal cells, explains fluorescence and describes the fluorescence microscope. The document is largely devoted to the characterization and distribution of voltage-sensitive dyes (VSD). The practical part deals with the various components necessary to perform the experiment as a pulse generator, high-speed camera and camera’s acquisition and describes experiment. The conclusion will be compared with results from theoretical assumptions.
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Membrane potential measurement with voltage sensitive dyes in confocal microscopy
Heczková, Monika ; Sekora, Jiří (referee) ; Čmiel, Vratislav (advisor)
The aim of this work is to make a literature search in the measurement of membrane voltage using the voltage-sensitive dyes and suggest a method of measuring the membrane voltage on the available cells using the voltage-sensitive dye Di-4-ANEPPS and RH237. The work contains an introduction to electrophysiology of cell, explains fluorescence and typical fluorescence characteristics. The thesis contains the description of a fluorescence microscope. The document was largely devoted to characterization and distribution of voltage-sensitive dyes. The output of a design solution is a real experiment.
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Natural drugs in cancer treatment and their cytotoxicity
Hájková, Tereza ; Raudenská, Martina (referee) ; Provazník, Ivo (advisor)
The thesis deals with the natural substances in context with the cancer disease. The natural substances have a positive effect on the human organism and they are able to influence the viability and the growth of the cancer cells. The main mechanical device is to influence the mechanisms needed to start the apoptosis of the cancer cells and stopping further proliferation. The cancer cell lines utilization in the cancer disease is discussed in the thesis too. The thesis states common methods of determining the natural substances cytotoxicity. For the experimental part of the thesis it was chosen the MTT test method and the xCELLigence system for monitoring in real time. The mechanical device of the tested substance capsaicin in application on the prostate cell lines, tumorous PC3 and nontumorous PNT1A influence will be observed within the experimental part of the thesis.
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Cytotoxicity of selected naphthoquinones on prostatic cell cultures
Mondeková, Věra ; Gumulec, Jaromír (referee) ; Provazník, Ivo (advisor)
This master´s thesis discusses cytotoxicity of selected naphthoquinones on prostatic cell cultures. The introductory part is dedicated to general characteristic of naphthoquinones with focus on their cytotoxicity, testing of cytotoxicity and mechanisms of cytotoxicity. This part is followed by chapters about cytotoxicity, characteristics and biological activities of selected naphthoquinones; plumbagin and naphthazarin. The last part of this thesis’ theoretical section speaks about fluorescence microscopy and its use in research of naphthoquinones cytotoxicity. The practical part is dedicated to evaluation of cytotoxical tests’ results and to analysation of pictures of cells obtained by fluorescence microscope. At the end of thesis, all finding are summarized and put in the context.
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Phytotoxicity of selected naphthoquinones on a selected plant model
Rucký, Jakub ; Masařík,, Michal (referee) ; Provazník, Ivo (advisor)
The introductory part of this thesis is focused on the theoretical analysis of solved problems as examining the toxicity of naphthoquinones plumbagine and juglone, especially with regard to their allelopatic action. The next section is focuses on the plant stress, caused by the action of stress factors leading to their death. There is an experimental protocol and the possibilities of determination the effect of naphthoquinones on the plant model. Experimental section discusses changing growth parameters of the plant samples in different concentrations of the naphthoquinone. There is examined cell viability and changes in the synthesis of the secondary metabolites. Data obtained by using spectrophotometric and microscopic analysis are evaluated with STATISTICA software and statistical significance are plotted.
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Membrane potential measurement with voltage sensitive dyes in fluorescence microscopy
Tkáč, Jan ; Janoušek, Oto (referee) ; Čmiel, Vratislav (advisor)
The aim of this work is to make a literature search in the measurement of membrane voltage using voltage-sensitive dyes and suggest a method for measuring the membrane voltage on the available cells using the voltage-sensitive dye di 4 ANEPPS and its further implementation. The work contains an introduction to electrophysiology of cells, and explains typical fluorescence characteristics. The thesis contains the description of a fluorescence microscope. The document presents characteristics of voltage-sensitive dyes and their distribution. A large part of the work describes the implementation and measurement of the experiment. The document also includes different methods for measuring and processing of all results.
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The penetration features of the hydrofobized hyaluronic acid – based polymeric micelles.
Mischingerová, Monika ; Pekař, Miloslav (referee) ; Muthný, Tomáš (advisor)
The aim of this thesis was to investigate the penetration features of the hydrofobized hyaluronic acid – based polymeric micelles using Nile red as carried tracer. Furthermore, to implement basic characterization of polymeric micelles for potential cosmetic applications using Coenzyme Q10 (CoQ10) as carried substance. It was found that the size of the polymeric micelles with carried CoQ10 did not exceed 100 nm. Applied delivery systems based on hydrophobic hyaluronic acid were suitable for potential topical application. Delivery systems with Nile Red as carried tracer demonstrated excellent penetration features. We assume that delivery systems with CoQ10 will exhibit similar penetration features. An issue has appeared whether the carrier breaks or proceeds along with NR to the skin. Moreover, another experiments have been designed which could also verify the penetration features of these systems.
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Use of selected fluorescence techniques to characterization of microbial cells
Vaněk, Martin ; Pekař, Miloslav (referee) ; Márová, Ivana (advisor)
Carotenoids are one of the most abundant natural pigment. Furthermore, they have great potential for wide industrial utilization. Nowadays, prefered (and cheaper) synthetic method counters progressively improved biotechnological production. Far more cheaper production should be reached via implementing methods of on-line optimization, such as flow cytometry. In this thesis the dependence of yeasts autotofluorescence intensity on carotenoids concentration in dry mass was studied. The dependence was established by means of linear regression. Moreover, the dependence between light scatter and the amount of dry mass was established by means of model utilizing forward scatter, side scatter and number of cells in unit volume. Yeasts' cells was also studied under the lenses of laser scanning confocal microscope. The goal was to localize carotenoids inside cells. Experiments undertaken suggest that they are localized inside the lipidic granules, but more experiments are required for a proof.
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GenExLib C/C++ library, Reference Manual
Kovář, Bohumil ; Schier, Jan ; Kuneš, Michal
This technical report contains reference manual for the GenExLib C++ software library. for analysis of FISH microscopy images. The purpose of this library is to detect cell nuclei and fluorescence signals contained in the images and to evaluate their shape and intensity parameters. The reference manual describes in detail the installation of the library as well as methodology of image processing for the FISH images and application of the library in the user code.
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GenEx Software library for object detection in genetical analysis evaluation: User manual
Schier, Jan ; Kovář, Bohumil ; Kuneš, Michal
This technical report contains user manual for the GenEx library for evaluation of FISH microscopy images. The purpose of the library is to find and filter objects in images from fluorescence microscope and to detect fluorescence signals contained in these objects. The library has been designed to be used for diagnostics of the Turner syndrome chromosome aneuploidies in chromosome X, in other words, to find the (roughly round) interphase nuclei and to count the associated hybridization signals. It is characterized by the following features: - separate processing channels for DAPI and for the fluorescence signals. - no restrictions on the number of the signal channels - output in the form of a CSV-file.
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