This diploma thesis deals with analysing of modern methods for cell detection, visualization and quantification in 3D space. The first section deals with optical methods for cells detection. There is detailed discussion about cell labeling and detection on confocal microscopy. There is also description about developed algorithm for whole cell volume quantification from microscopy images. This could made a comparsion of fluorescence signal according to time of cell labeling and according to cell shapes. There was also optimalization of handmade tissue phantoms visualization. It could be compared a possibilities of cell detections in these phantoms by confocal microscopy and OCT. It was also implemented algorithm for quantification of cells from OCT images. Besides confocal microscopy and OCT cells are also analyzed by other methods. The last part is the Conclusion of results and comparison of used methods.

This thesis focuses on cell tracking in image sequences acquired using a multimodal holographic microscope (MHM). The principles of holographic microscopy are described together with the application in cells acquisition. The main part of the thesis describes a complete approach for segmentation and tracking of single cells in acquired in long-term sequences. The approach is designed based on parametric active contour models with specific modifications to achieve reasonable precision and robustness. The implemented method is described in detail, including the evaluation and demonstration of results.

Two-photon laser scanning microscopy is a modern method of in vivo neurophysiological research, capable of imaging up to hundreds of neurons at once. However, this method produces a large amount of data, difficult to process and analyze manually. This thesis presents Two-Photon Processor, a new toolkit for complex processing of data from two-photon microscope. During the work on this thesis, we designed the SeNeCA segmentation algorithm for detection of neurons in full-frame recording from a two-photon microscope. SeNeCA combines high speed and high quality of segmentation and, according to our evaluation, it currently is the best algorithm for segmentation of neurons in in vivo data. Two- Photon Processor is already routinely used in the Institute of Experimental Medicine of the ASCR, Department of Auditory Neuroscience, and it was published in the Journal of Neurophysiology.

This thesis deals with the topic of 3D image processing for digital holographic microscopy - numerical refocusing. This method allows to perform mathematically accurate defocus correction on image of a sample captured away from the sample plane and it was applicable only for images that were made made using coherent illumination source. It has been generalized to a form in which it is also applicable to devices that use incoherent (non-monochromatic or extended) illumination sources. Another presented achievement concerns hologram processing. The advanced hologram processing method enables obtaining more data mainly concerning precision of quantities from one hologram — normally, one would have to capture multiple holograms to get those. Both methods have been verified experimentally.

This master thesis deals with possibilities of using nanoparticles of different metals as markers for immune labeling in the low voltage electron microscope. It presents the low voltage elektron microscope LVEM 5 (Delong Instruments, Brno, Czech Republic) as a new type of a table-size microscope specially designed for observation of specimens composed from elements with low atomic numbers.

The topic of this master’s thesis is the design of a sample manipulator for TEM. Regarding which the segment of the sample manipulator used to move the sample in the direction of the x-axis (so-called x-stage) was designed. The introductory theoretical part is concerned with a brief description of transmission electron microscopy focused on the imaging methods used in TEM and the procedures of preparing and loading TEM samples. The theoretical part follows with the design of precise mechanisms, especially with respect to the inaccuracies of their operation and the technological aspect of the design. In the practical part, a detailed analysis of the design of the x-stage is presented. Kinematics and dynamics of the mechanism is analyzed in depth, following with the description of the overall design of the mechanism and some of its parts in detail.

This graduation theises deal with influence of an additional grating possitioned berween the sample and the ionisation detektor of the environmental scannin elektron microscope on signal detection. Signal level is measured for changing pressure, at different voltages on the aditional grating, at different sample - additional electrode distance and at different sample - detektor distance. Influence of the additional grating is simulated for real working conditions on PC.

This thesis deals with scanning electron microscope working at higher pressure in the specimen chamber. The main goal was to study the voltage contrast on the PN junction of the transistor under suitable working conditions for using environmental scanning microscope. The observation of sample was enabled by a scintillation detector designed for observation of high pressure.

Master‘s thesis deals with the methods and system for measurement of concentration of intracellular calcium in LabVIEW. The thesis covers the theory of fluorescence principles, fluorescence methods and indicators for measurement of concentration of intracellular calcium. The procedure of measurement by means of fluorescence microscope, camera and indicators is also mentioned. The theory dealing with confocal microscopy is presented and two techniques are compared. Literary solution is presented. The practical part describes the program with a graphical user interface used for acquisition and analysis of data measuring dynamics of intracellular calcium concentration. The experimental part describes the preparation and measurement of cells and evaluation of experiments.

The goal of this thesis is to describe the possibility of using a microscope for documentation defects and innovation of electrical machines. I used an electron microscope to document carbon brushes and nanomaterials for possible upgrade of the sliding contact. Use microscopes gives us detailed information about the structure of materials, at the largest stress of the electrical machine. Based on the collected data can be further analyzed and innovation of the carbon brush.