This diploma thesis deals with hyphenation of liquid chromatography with mass spectrometric detection based on microfluidic chip. Firstly, a miniaturized ion source for atmospheric-pressure chemical ionization (APCI), and atmospheric-pressure photoionization (APPI) was constructed. The main component of this source was a glass microfluidic chip. Geometry and the working conditions of the chip were optimized. Since both ion sources work under the same conditions, possible advantages resulting from APCI/APPI combination were investigated. Furthermore, the performance characteristics of the sources were evaluated, and compared to the conventional high flow-rate sources. The best performing source, APCI, was then hyphenated with HPLC using low flow-rate. A method for separation of fatty acids methyl esters using Supelco 37 standard was developed. The separation conditions were as follows: C18 reversed stationary phase, and acetonitrile containing 0.1 % formic acids was used as the mobile phase. A temperature gradient was used in order to enhance the speed of the separation. The limits of detection and quantitation of for selected analytes using the chip micro-APCI were calculated, and compared to the ones obtained using a commercially available micro-APCI source. The method was used for separation of...

In this diploma thesis we will study the luminescence properties of nanocrystals. We will summarize the basic division according to size and standard method of theoretical description of semiconductor and metal nanocrystals. We will describe the luminescence properties of nanocrystals and the influence of the surrounding environment. In the se- cond part of the thesis we will follow up the construction of an apparatus for imaging luminescence spectroscopy intended for the measurement of individual nanoobjects in a liquid environment. Finally, we will study luminescence properties of organixally passi- vated metal clusters in a liquid environment with changes in temperature and excitation intensity. We compare the luminescence of gold nanocrystals with and without long po- lyethylene glycol chains bound on the surface. 1

This diploma thesis deals with cavitation flow in the microscale, which remains an area with a lack of sufficient description of this phenomenon. At the same time, microfluidics is a field experiencing a dramatic rise in numerous biochemical applications, which underlines the relevance of researches of this type. In theoretical part of the thesis, cavitation was described in detail. In the practical part, a microfluidic device with a cavitation orifice was designed and manufactured. The ANSYS program was used for this design. An experiment was performed with the designed microchip, the aim of which was to observe a cavitating flow on the orifice. This measurement took place at the microfluidic laboratory at Victor Kaplan Department of Fluid Engineering. Due to the failure of the experiment, a CFD model of two-phase cavitation flow was built. The conclusions of the thesis were compiled from the findings of measurement and the results of modeling.

Microarray technology provides powerful tools in the field of biomedicalresearch because it can measure molecular interactions in a highly parallelfashion. It has uses in protein, DNA or cell research, in both discovery anddiagnostic applications. Microfluidics, on the other hand, provides thenecessary tools to rapidly transport and mix small volumes of sample to amicro-sensor area. Bridging these two technologies has the potential todevelop a miniaturized, automated and ease-of-use toolbox for biologicalanalysis. However, the integration of microfluidics with microarrays is notstraightforward, as if a robust and leak-tight seal between the microarray andthe microfluidic channels. Current sealing methods are either impractical,such as mechanical clamping, or not compatible with proteins, such as heat orplasma bonding or gluing. Moreover the former methods create a permanentseal that, once applied prevents the microfluidic structure to be removed later.This work focuses on developing a microfluidic add-ons ("Biosticker") that canbe robustly sealed with protein microarrays with maintained biologicalactivity, but at the same time easily removed to allow for multiple stickersapplied in a sequence or scanning of the microarray in a standard reader. Thefeatures of the novel Biostickers are made possible by the use ofOff-stoichiometry thiol-ene-epoxy (OSTEmer) polymers. In this thesis, wedesign and fabricate Biostickers for rapid integration with pre-spottedmicroarrays and experimentally verify how these micropatterned Biostickerscan be used to significantly facilitate multiplexed assays, by avoiding the useof beads. / Microarray-tekniken är ett kraftfullt verktyg inom biomedicinsk forskningeftersom den kan mäta miljontals molekylära interaktioner parallellt. Den haranvändningsområden i protein-, DNA- eller cellforskning, både i forskningoch diagnostik. Mikrofluidik, å andra sidan, ger de nödvändiga verktygen föratt snabbt transportera och blanda små provvolymer till en sensoryta. Genomatt kombinera dessa två teknologier finns potential att utveckla enminiatyriserad, automatiserad och lättanvänd verktygslåda för biologiskanalys. Emellertid är integrationen av mikrofluidik med mikroarrayer inteenkel, då ytorna är känsliga, kanalerna mycket små men tätningen måste varaperfekt. De vanligast förekommande förseglingsmetoder är antingenopraktiska, som mekaniskt tryck eller så är de inte kompatibla med proteiner,som t.ex. värme- eller plasmabondning. Dessutom syftar de flestaförseglingsmetoder mot att skapa en permanent försegling som vidanvändning förhindrar mikrofluidikstrukturer från att tas bort i ett senareskede, tex. vid avläsning i en skanner. Detta arbete fokuserar på att utvecklamikrostrukturerade plastartiklar ("Biosticker") innehållande kanaler ochkaviteter. Dessa Biosticker kan på ett robust och läckfritt sätt kansammanfogas med proteinmikroarrayer utan att påverka den biologiskaaktivitetet men samtidigt kunna avlägsnas för att tillåta flera Biostickersapplicerade i en sekvens eller scanning i en mikroarrayläsare. Dessafunktioner möjliggörs genom av så kallade icke-stökiometrska-tiol-ene-epoxipolymerer (OSTEmer) används som material. I den här avhandlingenutvecklas och tillverkas Biostickers för snabb integrering medproteinmikroarrayer. Det verifieras även experimentellt hur dessa Biostickerskan användas för att underlätta genomförandet av sk. multiplexadeprotinanalyser.

This master thesis deals with the use of microfluidics for the purpose of microfluidic enzymatic reactor for drug screening. At first it considers the issue from a theoretical point of view – describes microfluidics as a newly developing and promising field of production of microfluidic devices, materials, biomedical applications and advantages and disadvantages of microfluidics overall. Furthermore, it focuses on an area of analytical utilization of enzymes within enzyme reactors. In the first part of the experimental section, conditions for the testing of enzymes of xenobiotics metabolism in the liver were optimized, namely the model of coumarin metabolism via the spectrofluorimetry method. The second part of the experimental work dealt with optimization of the fabrication conditions of microfluidic chips from OSTE (off-stoichiometry Thiol Ene) via the soft lithography method. Subsequently, the functionality of the produced chips was tested. Based on the results of both parts of the experimental work, an evaluation was carried out to assess the suitability of their interconnection for future research – screening of microsomal enzyme activity and model biotransformation of drugs within the channels of the fabricated devices.

In this doctoral thesis we study novel analogues based on R06 aptamers and targeting TAR hairpins of the HIV virus by means of surface plasmon resonance biosensor, which allows for sensitive and real-time monitoring of molecular interactions. We investigate seven different modifications placed at nine different positions on the R06 aptamer in order to find out their applicability in the construction of efficient and stable anti-TAR oligonucleotides. We also determine which positions are suitable for substitutions with a modification and interpret the results in the context of the local nucleotide geometries and interactions in the TAR/anti-TAR complex. In this doctoral thesis we further develop a new fluidic system. This fluidic system eliminates sample dispersion and intermixing effects and thus enables accurate monitoring of molecular interactions on the surface of an SPR chip. We also characterize experimental conditions on the surface of an oligonucleotide chip and their relations towards bio-molecular assays. Specifically, we study the shielding effect of monovalent and divalent cations, which are crucial for the interaction of negatively charged oligonucleotides.

The goal of submitted thesis is to perform a computer simulation of bubble creation in T-channel. In the first section of the paper, the theoretical applications of microfluidic bubble, micromachines and droplet formation are described. In the second part of the text, author uses cross flowing method for simulation od bubble creation. Furthermore, several settings in computer simulation software Fluent are mentioned. In addition, the influence of velocity at the T-channel inlet on surface tension and on bubble length is presented.

Microfluidic devices are more frequently being used in medicine as they operate with small amounts of test samples, such as blood or reagent chemicals. To work with such substances, effective mixing of the solution is usually required, which emerged as the most challenging problem in microfluidic systems. Due to the minor dimensions of the devices only laminar flow occurs, thus the turbulent eddies do not contribute to the mixing, but only the molecular diffusivity.

Předložená dizertační práce se zabývá kombinací optických mikromanipulací s mikrospektroskopickými metodami. Využili jsme laserovou pinzetu pro transport a třídění živých mikroorganismů, například jednobuněčných řas, či kvasinek. Ramanovskou spektroskopií jsme analyzovali chemické složení jednotlivých buněk a tyto informace jsme využili k automatické selekci buněk s vybranými vlastnostmi. Zkombinovali jsme pulsní amplitudově modulovanou fluorescenční mikrospektroskopii, optické mikromanipulace a jiné techniky ke zmapování stresové odpovědi opticky zachycených buněk při různých časech působení, vlnových délkách a intenzitách chytacího laseru. Vyrobili jsme různé typy mikrofluidních čipů a zkonstruovali jsme Ramanovu pinzetu pro třídění mikro-objektů, především živých buněk, v mikrofluidním prostředí.

This dissertation concerns the development, fabrication and integration in a gas sensing microdevice of a novel 3-dimensional (3D) nanostructured metal-oxide semiconducting film that effectively merges the benefits of inorganic nanomaterials with the simplicity offered by non-lithographic electrochemistry-based preparation techniques. The film is synthesized via the porous-anodic-alumina-assisted anodizing of an Al/Nb metal bilayer sputter-deposited on a SiO2/Si substrate and is basically composed of a 200 nm thick NbO2 layer holding an array of upright-standing spatially separated Nb2O5 nanocolumns, being 50 nm wide, up to 900 nm long and of 8109 cm2 population density. The nanocolumns work as semiconducting nano-channels, whose resistivity is greatly impacted by the surface and interface reactions. Either Pt or Au patterned electrodes are prepared on the top of the nanocolumn array using an innovative sensor design realized by means of microfabrication technology or via a direct original point electrodeposition technique, followed by selective dissolution of the alumina overlayer. For gas-sensing tests the film is mounted on a standard TO-8 package using the wire-bonding technique. Electrical characterization of the 3D niobium-oxide nanofilm reveals asymmetric electron transport properties due to a Schottky barrier that forms at the Au/Nb2O5 or Pt/Nb2O5 interface. Effects of the active film morphology, structure and composition on the electrical and gas-sensing performance focusing on sensitivity, selectivity, detection limits and response/recovery rates are explored in experimental detection of hydrogen gas and ammonia. The fast and intensive response to H2 confirms the potential of the 3D niobium-oxide nanofilm as highly appropriate active layer for sensing application. A computer-aided microfluidics simulation of gas diffusion in the 3D nanofilm predicts a possibility to substantially improve the gas-sensing performance through the formation of a perforated top electrode, optimizing the film morphology, altering the crystal structure and by introducing certain innovations in the electrode design. Preliminary experiments show that a 3D nanofilm synthesized from an alternative Al/W metal bilayer is another promising candidate for advanced sensor applications. The techniques and materials employed in this work are advantageous for developing technically simple, cost-effective and environmentally friendly solutions for practical micro- and nanodevices, where the well-defined nano-channels for charge carriers and surface reactions may bring unprecedented benefits.