Many studies describe the beneficial effect of probiotic bacteria of the gastrointestinal tract and their immunomodulatory effects. The aim of this diploma thesis was to evaluate the effect of probiotics, which are commonly used in human and animal nutrition, on the viability of the cells of the immune system. For the experiment were selected blood monocytes, which were isolated from porcine and human blood by dextran sedimentation. Monocyte population was co-cultivated in vitro with probiotic strains Bifidobacterium bifidum, Lactobacillus rhamnosus and Enterococcus faecium for 2 and 4 hours. The percentage of apoptosis and necrosis of monocytes was analysed by flow cytometry. The results showed statistically significant differences in the proportion of apoptotic and necrotic of porcine and human monocytes. It is obvious that the selected strains of probiotics have immunomodulatory effect on immune cells. Enterococcus faecium and Lactobacillus rhamnosus induce cell death of porcine and human monocytes during short-term cultivation in vitro.

Assessment of semen quality (sperm viability) is an essential for the efficient honeybee semen storage. The motility of spermatozoa is not the ultimate parameter of sperm viability. Instead, the viability of spermatozoa is assessed by testing their structural or functional plasma membrane integrity. These assays are based primarily on the technique of microscopy evaluation of either fluorescent DNA dyes or hypo-osmotic "tail-coiling" reaction of spermatozoa. However, the technique of microscopy evaluation is time-consuming, subjective and often is not accurate. On the other hand, the technique of flow cytometry is rapid, objective and precise. Nowadays, this technique is affordable for many institutions, as the price of "benchtop" flow cytometer is relatively low. To best of our knowledge, there is currently no flow cytometry assay available for evaluation of functional integrity of honeybee spermatozoa. Here we report the establishing of novel flow cytometry assay for simultaneous evaluation of structural and functional plasma membrane integrity. DRAQ7, the new fluorescent DNA dye was used to test the structural integrity of plasma membranes. The hypo-osmotic "tail-coiling" reaction was used for flow cytometric assessment of functional integrity of spermatozoa plasma membranes. The combination of both is a valuable tool for rapid and precise evaluation of sperm viability in honey bees.

A practical technique for enzymatic dissociation and isolation of pikeperch (Sander lucioperca) (Percidae, Teleostei) early stage germ cells (eGC), including spermatogonia and spermatocytes, is reported in this study. Their potential to differentiate into functional gametes, and transmit genetic information to the next generations makes them suitable for cryopreservation and surrogate reproduction studies by germline chimera. Two different age groups (14 month old and 18 month old) of pikeperch were used to adjust the correct stage of eGC isolation. Finally the 18 month old samples were selected due to their high average gonadal volume (0.513 g). 10 ml PBS + 0.3% trypsin (304 mOsm/kg, pH 8) were used for enzymatic dissociation of testicular cells and they were sorted by centrifugation in Percoll density gradient. eGCs were identified on the basis of their ploidy level by CYSTAIN DNA 1 steps kit (PARTEC) and morphological characteristics trough by light microscopy. Cell counting was performed on histological sections and Percoll gradient layers whit the method of random square counting. The method of isolation enriched the number of eGC from 41.3% to 84.7%, obtained from the 33% of Percoll concentration.

This thesis focuses on the study of domestic and foreign literature on the determination of sulfur dioxide in wine. A detailed description of the standard and alternative methods, their analysis, identification of possible shortcomings or disadvantages and specifying strengths and advantages. Theory obtained by studying literature is verified by a several samples of wine are provided different forms of sulfur dioxide with different methods . Subsequently, the results obtained by various methods are compared in terms of accuracy and applicability in the viticultural practice. Everything should be processed in order that the thesis could become an additional study material for university students.

Calamagrostis phragmitoides belongs to taxonomically complex C. purpurea aggregate with circumpolar distribution area. The relations between morphologically similar taxa of this aggregate are not clear. Populations of C. purpurea agg. from Central Europe, Scandinavia and Siberia (including one plant from the North America) were genetically studied (cpDNA, ITS, ploidy level by FCM). Allozyme and morfological variability of populations C. phragmitoides and morphologically similar C. canescens was also studied in the Czech Republic. This study was supported by grant SGA PrF JU in 2014.

No description available.

Barbora Hadámková Determination the mechanism of entry F. tularensis into B lymphocytes Diploma thesis Charles University, Faculty Of Pharmacy in Hradec Králové Study program: Pharmacy Background: Besides processing the research with basics knowledge of the problem, the main aim of the study was the analysis of mechanism of entrance of intracellular bacteria Francisella tularensis into B cells. Methods: The B cells, which we obtained through peritoneal lavage from mice Balb/c, we blocked using antibodies individual complement receptors, B cell receptor and Fcƴ receptor. The population of the cells was infected by bacteria F. tularensis LVS/GFP opsonized by complement and/or by antibodies. Using flow cytometry we measured the percentage of infection of individual subpopulations of B cell B1a, B1b and B2 and we evaluated the influence of blocking and opsonization on the infection. Results: From the measured data, we can say that the percentage of infected B cells after infection by F. tularensis opsonized by complement is increased. This increase was more distinct in subtype of B cells B1b and B2. On the other hand, the opsonization F. tularensis by antibodies did not affect the infection. We also found out, that blocking of Fcƴ receptor has decrease the infection, if we used for infection of B cells...

Monocytes are a population of mononuclear lekocytes. The aim of this thesis was to select a suitable methodology for the isolation of monocytes and their cultivation into dendritic cells. For the experiment, bovine blood was taken from vena jugularis externa. This blood was used to isolate monocytes based on the density gradient of OptiPrep solution and Histopaque solution. The results showed that the method of isolation based on the density gradient of the Histopaque solution and the magnetic separation appeared to be a more efficient method. The monocyte population was cultured for 72 hours. Cells were analyzed by light microscopy and CD14 positive cells appeared to be transformed into dendritic cells. The purity of these cell population was determined by flow cytometry. Dendritic cells have a wide range of utility mainly in immunotherapy of various diseases and also play an important role in cancer therapy.

Although the genus Nymphaea (waterlily) includes only two native species (N. alba and N. candida) in Central Europe, it poses a great challenge to taxonomy and biosystematics. The determination of both species is hampered by their phenotypic similarities, and species boundaries can be further blurred by interspecific hybridization. In addition, ornamental cultivars of different parentage often escape from cultivation and make the situation even more complex. To get insight into the caryological and phenotypic variability of czech waterlilies, the DNA flow cytometry and both distance-based and geometric morphometrics were used. Collections showed two different groups of fluorescence intensities, corresponding to N. alba and N. candida, respectively. In addition, intermediate values of nuclear DNA amount were found in some plants from South Bohemia, indicating their hybrid origin. Surprisingly, ornamental cultivars possessed the smallest genome sizes. The amount of nuclear DNA therefore seems to be a promising species-specific marker that enables not only native species but also cultivars to be distinguished. Cytometrically-proven individuals have been subjected to multivariate morphometrics, and high differenciation was discovered especially between native species. More complicated seems the distinction...

[Abstract] This thesis is aimed at better understanding of cytotype co-existence in mixed- ploidy populations with an emphasis on a microevolutionary processes behind it. Our past knowledge was based on a few thoroughly investigated model taxa like Chamerion angustifolium and Heuchera grossulariifolia, but some generalizations seem to be premature in the light of new findings. A detailed research of other taxa included in the thesis showed that polyploid complexes can vary dramatically in their ability to cope with the co-existence of cytotypes in mixed-ploidy popu- lations. Whereas mixed-ploidy populations are virtually lacking in some species (an example being Vicia cracca, Paper III.), ploidy-heterogeneous populations are very common in others, maintained by free mating interactions and the absence of reproductive isolation among cytotypes (e.g. Pilosella echioides, Paper II.). The strenght and cumulative effect of various breeding barriers (both pre- or post- zygotic) govern the position of a particular multi-ploidy complex between these two extremes and co-determine the type of cytotype co-existence in its mixed- ploidy populations. Despite the fact that the number of studies revealing cytotype co-existence has been increasing rapidly, evolutionary background and consequences of such co-...