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Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Radka Vaňková Supervisor: doc. PharmDr. Martin Beránek, Ph.D. Title of diploma thesis: Influence of Carrier Molecule Addition on Plasma Free Circulating Nucleic Acid Extraction Outline: The aim of this thesis was to compare the yields of cell-free DNA (cfDNA) extracted from blood plasma specimens with the addition of commercially available carrier molecules. According to the real-time polymerase chain reaction (qPCR) results, we chose the most optimal carrier molecules for routine isolation of cfDNA from blood plasma samples. Methods: cfDNA was isolated from aliquots of a pooled blood plasma by spin column extraction method NucleoSpin Plasma XS (Macherey-Nagel). We used 7 different types of carrier molecules as RNA carrier (Qiagen), Glycogen (Invitrogen), Poly (A) Polyadenylic acid (Roche), Linear Acrylamide (Invitrogen), Yeast transfer RNA (Ambion), Salmon sperm DNA (Invitrogen), Herring sperm DNA (Promega). After the extraction, plasmatic DNA with the addition of carrier molecules was quantified by spectrophotometry, fluorimetry and qPCR. Results: The best results were achieved with the addition of polyadenylic acid with final concentration of 1.55 μg/ml. The addition of carrier RNA and...

Diplomová práce se zabývá tuhostní optimalizací nosiče satelitů, tzv. Dispenseru. První kapitola uvádí přehled evropských vesmírných aktivit a poskytuje nezbytné technické pozadí týkající se nosných raket. Druhá kapitola se věnuje popisu tvorby výpočetního modelu, neboť veškeré výpočty, včetně optimalizace popsané v kapitole třetí, jsou založeny na metodě konečných prvků. Pro optimalizovanou variantu je ve čtvrté kapitole provedena základní pevnostní kontrola.

The aim of this diploma thesis is a design of hydraulic and pneumatic systems of a communal vehicle. This thesis includes a theoretical part where hydraulic and pneumatic systems of a communal vehicle are described; it also refers to a design of a pneumatic suspension system, a hydro-pneumatic breaking system, a hydraulic system for hook carrier containers including a design and a calculation of hydraulic motors and the volume of a hydraulic tank and a presentation of possible construction alternatives.

This thesis deals with the means of mechanization for maintenance of permanent grassland by mowing without collection or mulching. The means of mechanization belongs to category of the one axle tool carrier and the zero turn mower. The aim of the study is practical comparison of the means of mechanization and their suitability for different grassland. The practical part of study deals of functional and operational aspects. Futher, the machines were compared in terms of operating cost. The machines are assigned to suitable operators, on the basis of this analysis.

The aim of the thesis is focused on the proposal of a new technological procedure of the blade carrier for use in its own production. Currently, the carrier is manufactured by the German company B.Maier Zerkleinerungstechnik GmbH. The work includes several parts. The first part deals with the component itself, its function and importance, as well as its function in the assembly and the whole machine. In the next part the technological procedure of the blade carrier production is created. Fleet of machines including the necessary tools is also designed. In a separate chapter there is an economic evaluation of the proposed solution in two different variants, a subsequent evaluation and a proposal for implementation.

This master’s thesis is focused on preparation and analysis of ultrathin CaF2 (calcium fluoride) films on a graphene substrate. CaF2 was deposited in UHV conditions at substrate temperatures in the range from 20 °C to 400 °C. The material was deposited on Si(111) with a native SiO2 layer and on a substrate with a CVD graphene layer. The deposited films were analyzed by XPS, AFM and SEM. Moreover four different sample holders for deposition purposes were designed in this work.

In this diploma thesis we are discussing about using 3D bioprinting in tissue engineering. We are discribing using biomaterials for construction scaffolder and aplication stem cells in 3D bioprinting. Last section of theoretical part deals with very often used techniques of 3D bioprinting and we are focused on extrusion technique. In the practical part we propose a method for print vasculars structures. We realized prototype of print head, her design and 3D printing of individual parts. To mechanical part we create a control system for printing control. At the end we visualize the organization of the cells using program modules.

The work was comprehensively focused on the development of adhesive forms of probiotics in the form of a biofilm on combined carriers with a prebiotic component. The second part dealed with the influence of food on the multiplication and survival of selected types of probiotic bacteria. Subsequently, the effect of individualized probiotic supplements on changes in the human intestinal microbiome was monitored. Suitable adherent probiotic strains for biofilm formation were selected and tested. Methods have been introduced and different variants of carriers for culturing and binding bacteria have been tested. In vitro experiments verified the stability of biofilm stucture and its resistance to low pH, bile and antibiotics in comparison with the planktonic cell form. The antimicrobial effect of probiotic strains in the form of a biofilm was studied. The cultivation of the multispecies biofilm on the combined carrier was optimized and the stability of the biofilm and the final viability of probiotic bacteria were confirmed. Furthermore, the influence of various foods and beverages on the viability of probiotic bacteria was evaluated with emphasis on the simulation of passage through the gastrointestinal tract. Both models, solutions with standardised concentrations of alcohol, sugar, salts, proteins or different pH and different types of real foods and beverages were tested. The effect of food and beverages was tested on monocultures of Lactobacillus acidophilus, Bifidobacterium breve and on probiotic capsules containing a mixed culture of probiotic microorganisms. The survival of probiotics in various food matrices in the simulated gastrointestinal tract was quantitatively different. We managed to define foods suitable for supporting the multiplication of probiotic bacteria. A separate part of the work was focused on the targeted modulation of the intestinal microbiome by individualized probiotics that were prepared on the basis of molecular biological analyzes of the intestinal microbiome aimed at detecting the percentage of lactobacilli, bifidobacteria and phylum Firmicutes and Bacteroidetes. Personalized probiotic supplementation confirmed the positive effect of this approach on microbiome changes, especially on the increase of the content of lactobacilli, bifidobacteria and the overall diversity of the microbiome.

This diploma thesis brings new data about design, synthesis, physico-chemical characterisation and biological efficacy of the novel star-like HPMA-based conjugates intended for treatment of solid tumors. Recently, many different water-soluble drug delivery systems based on N-(2- hydroxypropyl)methacrylamide (HPMA) copolymers have been described. Here, we report synthesis and physico-chemical characterisation of high molecular weight star-like HPMA- based polymer carriers with low polydispersity prepared by controlled grafting of HPMA copolymers onto PAMAM dendrimer core. With the aim to keep the polydispersity of drug delivery system as low as possible, reversible Addition-Fragmentation Chain Transfer (RAFT) polymerisation was used for HPMA-based polymer precursor preparation. The end groups of the polymer presursors was afterwards used for grafting using carbodidimide condensation reaction or copper free click chemistry on polyamidoamine (PAMAM) dendrimers resulting in a formation of star-like high-molecular-weight (HMW) drug carriers. Described synthetic procedure provided preparation of star-like HMW drug carriers with Mw between 1.105 - 3.105 g/mol and narrow distribution of Mw. The model drug, doxorubicin (Dox), was attached to the hydrazide group containing polymer cariers by pH- sensitive...