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Driscoll, Peter F. "Bioanalytical Applications of Chemically Modified Surfaces." Digital WPI, 2009. https://digitalcommons.wpi.edu/etd-dissertations/465.
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"The design and development of chemically modified surfaces for bioanalytical applications is presented. Chemical surface modification is demonstrated to be a method to control surface properties on the molecular level by selecting the appropriate substrate, linking chemistry, and terminal group functionality. These systems utilize spontaneous interactions between individual molecules that allow them to self-assemble into larger, supramolecular constructs with a predictable structure and a high degree of order. Applications investigated in this thesis include: surface patterning, switchable surface wettability, and biological sensor devices that combine surface based molecular recognition, electrochemical detection methods, and microfluidics. A multilayered approach to complex surface patterning is described that combines self-assembly, photolabile protecting groups, and multilayered films. A photolabile protecting group has been incorporated into molecular level films that when cleaved leaves a reactive surface site that can be further functionalized. Surface patterns are created by using a photomask and then further functionalizing the irradiated area through covalent coupling. Fluorophores were attached to the deprotected regions, providing visual evidence of surface patterning. This approach is universal to bind moieties containing free amine groups at defined regions across a surface, allowing for the development of films with complex chemical and physico-chemical properties. Systems with photoswitchable wettability were developed by fabricating multilayered films that include a photoisomerizable moiety, cis-/trans- dicarboxystilbene. When this functionality was incorporated into a multilayered film using non-covalent interactions, irradiation with light of the appropriate wavelength resulted in a conformational change that consequently changed the hydrophobicity of the substrate. Methods were investigated to increase the reversibility of the photoswitching process by creating surface space between the stilbene ligands. Utilizing mixed monolayers for spacing resulted in complete isomerization for one cycle, while the use of SAMs with photolabile groups produced surfaces that underwent isomerization for three complete cycles. A microfluidic device platform for ion sensing applications has been developed. The platform contains components to deliver small volumes of analyte to a surface based microelectrode array and measure changes in analyte concentration electrochemically in an analogous method to that used in conventional electrochemical cells. Crown ether derivatives that bind alkali metal ions have been synthesized and tested as ionophores for a multi-analyte device of this type, and the sensing platform was demonstrated to measure physiological relevant concentrations of potassium ions. Advantages of this design include: high sensitivity (uM to mM), small sample volumes (less than 0.1 mL), multi-analyte capabilities (multiple working electrodes), continuous monitoring (a flow through system), and the ability to be calibrated (the system is reusable). The self-assembled systems described here are platform technologies that can be combined and used in molecular level devices. Current and future work includes: photopatterning of gold and glass substrates for directed cell adhesion and growth, the design and synthesis of selective ion sensors for biological samples, multi-analyte detection in microfluidic devices, and incorporating optical as well as electrochemical transduction methods into sensor devices to allow for greater sensitivity and self-calibration."
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Anazia, Oge. "Chemistry of Zirconia and Its Bioanalytical Applications." TopSCHOLAR®, 2009. http://digitalcommons.wku.edu/theses/127.
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This research studies the chemical nature of zirconia and the complex surface chemistry of zirconia in order to better comprehend its behavior under chromatographic conditions. This research shows how the physical and chemical properties of zirconia depend strongly on the thermal treatment during synthesis. The morphology of the samples was also studied. The absorption capability of Adenosine Triphosphate (ATP) on zirconia was also monitored and spectrally characterized.
The results of this research showed how the properties of zirconia vary with thermal treatment. It was observed that the zirconia prepared at a higher temperature had lower surface area, lower pore size and pore volume as compared to the zirconia prepared at a lower temperature. The morphology studies showed the porosity of the zirconia. The results from the absorption experiments showed that zirconia prepared at a higher temperature absorbed more ATP than the zirconia prepared at a lower temperature. Significant changes were also observed on the pellets of zirconia pre and post absorption experiments. I hope that this research sheds more light on the complex properties of zirconia’s surface chemistry and the results of this study could better help in the application and use of zirconia in chromatography to separate proteins.
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Pereiro, Iago. "Microfluidic magnetic fluidized bed for bioanalytical applications." Thesis, Paris 6, 2016. http://www.theses.fr/2016PA066089/document.
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Des phénomènes de fluidisation de billes magnétiques apparaissent à l'échelle micrométrique au sein du système de lit fluidisé microfluidique. On obtient un fonctionnement en flux continu à basse pression de travail avec un étroit contact liquide/solide et une recirculation constante des billes, des caractéristiques avantageuses pour des applications dédiées à la pré-concentration de cibles biologiques. La caractérisation du système physique a montré l'influence de paramètres tels que la géométrie de la chambre ou la distribution du champ magnétique, leur optimisation étant nécessaire pour obtenir des phénomènes de fluidisation à cette échelle et améliorer le mélange et la distribution des billes. De plus, le potentiel du lit fluidisé comme plateforme pour des bio-essais analytiques a été exploré avec succès lors d'applications biologiques: 1) la pré-concentration de bio-markers de la maladie d'Alzheimer et leur marquage in situ pour un future couplage avec des techniques de détection sensibles; 2) la détection de bactéries sans besoin de marquage préalable à travers une immuno-capture suivie d'une culture donnant lieu à des changements physiques du support fluidisé; 3) l'extraction d'ADN contenant un gène cible et son ultérieur amplification enzymatique sur la surface des billes, suivie d'une détection multiplexée des mutations présentes par un système de microarray. Ainsi, le lit fluidisé magnétique rend possible des applications au de-là d'un simple système de pré-concentration, permettant son utilisation comme une plateforme efficace de biologie moléculaire allant jusqu'à l'utilisation des propriétés autorégulatrices inhérentes au système comme mécanisme de détectionWith the use of an external magnetic field and magnetic microbeads, the microfluidic magnetic fluidized bed system enables fluidization phenomena at the microscale. This results in flow-through operations at low driving pressures with intimate liquid/solid contact and a continuous beads recirculation, interesting for efficient biological target preconcentration applications. The physical system has been characterized, showing the importance of chamber angle of aperture and height confinement as well as magnetic field distribution parameters, to obtain fluidization and further enhance mixing and maximize beads density. Further, the potential of the fluidized bed as a platform for analytical bioassays has been successfully explored with a series of biologically relevant applications: (1) the preconcentration of rare Alzheimer’s biomarkers together with their in situ fluorescence labeling for future enhanced detection with hyphenated techniques; (2) the label-free sensitive detection of bacteria in liquid food samples through the specific immunocapture and on-chip culture of these microorganisms and the resulting physical changes induced in the fluidized support; (3) the gene-specific extraction of DNA and its subsequent enzymatic amplification on the surface of the beads, coupled to a microarray detection system for a multiplexed detection of cancer-inducing mutations. These results show that the applications of the magnetic fluidized bed go beyond its initial conception as a dynamical affinity-based concentrator, serving as an efficient platform for molecular biology protocols and even making use of its inherent auto-regulating properties as a detection mechanism
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Selegård, Robert. "Polypeptide functionalized gold nanoparticles for bioanalytical applications." Doctoral thesis, Linköpings universitet, Molekylär fysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-106719.
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Detection strategies that allow for simple, rapid, cost efficient and sensitive monitoring of proteins and their interactions with biomolecules are of great importance in drug development and diagnostics. This thesis describes the development of bioanalytical applications based on the tunable self-assembly of gold nanoparticles functionalized with a de novo designed polypeptide. Strategies for protein affinity sensing and for detection of several fundamentally important biological processes have been investigated, including Zn2+-mediated coordination between polypeptides and low molecular weight chelants and protease and phosphatase activity. A Zn2+ responsive synthetic polypeptide designed to fold into a helix-loop-helix motif and dimerize into a four-helix bundle has been used to control the stability and self-assembly of gold nanoparticles. This polypeptide has a high negative net charge at neutral pH as a consequence of its many glutamic acid residues, efficiently preventing folding and dimerization due to charge repulsion. Zn2+ coordination provides a means to trigger folding and dimerization at neutral pH. The polypeptide can be readily attached to gold nanoparticles via a cysteine residue in the loop region, retaining its folding properties and responsiveness to Zn2+. The polypeptide functionalized gold nanoparticles display excellent colloidal stability but aggregate reversibly after addition of millimolar concentrations of Zn2+. Aggregates are dense with a defined interparticle distance corresponding to the size of the four-helix bundle, resulting in a distinct red shift of the localized surface plasmon resonance band. Three completely different strategies for colorimetric biosensing have been developed, all being based on the same responsive hybrid nanomaterial. In the first strategy a synthetic receptor was co-immobilized on the gold nanoparticles together with the Zn2+ responsive polypeptide. Protein analyte binding to the receptor could be detected as this interaction sterically prevented aggregation induced by Zn2+. In the second strategy the reduction in colloidal stability caused by specific proteolytic cleavage of the immobilized polypeptide was exploited to monitor the enzymatic activity. The third strategy utilized the sensitivity of the system to small variations in Zn2+ concentration. The presence of low molecular weight chelants was found to influence the mode of aggregation, both by sequestering Zn2+ and through the formation of ternary complexes involving the polypeptides, which prevented dimerization and thus aggregation. This approach was further developed into a generic concept for phosphatase detection exploiting the different affinity of enzyme substrates and reaction products for Zn2+. The flexibility of the different detection schemes enables detection of a large number of analytes by exploiting the tunable stability of the nanoparticles and the possibilities to effectively decouple the recognition event and the nanoparticle stability modulation.
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Cannan, Susan. "Microelectrode methods for bioanalytical and biophysical applications." Thesis, University of Warwick, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.397013.
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Dolatmoradi, Ata. "Thermally-Assisted Acoustofluidic Separation for Bioanalytical Applications." FIU Digital Commons, 2017. http://digitalcommons.fiu.edu/etd/3371.
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Changes in the biomechanical properties of cells accompanying the development of various pathological conditions have been increasingly reported as biomarkers for various diseases and as a predictor of disease progression stages. For instance, cancer cells have been found to be less stiff compared to their healthy counterparts due to the proteomic and lipidomic dysregulations conferred by the underlying pathology. The separation and selective recovery of cells or extracellular vesicles secreted from such cells that have undergone these changes have been suggested to be of diagnostic and prognostic value.
This dissertation first describes the implementation of a stiffness-based separation of phosphatidylcholine-based vesicles using a method first introduced based on the research in this work and was dubbed thermally-assisted acoustophoresis, or thermo-acoustophoresis. By tuning the temperature, we achieved the separation of vesicles of the same size, shape, and charge but with different stiffness values. It was observed that at a specific transition point, the acoustic contrast factor of vesicles changed sign from positive to negative. This change was mainly due to change in the compressibility of the vesicles, which is inversely proportional to stiffness. The acoustic contrast temperature (Tϕ), corresponding to the temperature at which the contrast factor switches sign, was determined to be unique to the composition of the vesicles. This unique temperature signature allowed us to develop this separation method of vesicles with distinct membrane stiffness with target outlet purities exceeding 95%.
We have further explored the functionality of this method by experimenting with cholesterol-containing vesicles. In cells, the cholesterol content plays a crucial role in determining stiffness. Changes in the cholesterol content in cellular membranes can be an indication of pathological disorders. We evaluated the Tϕ of vesicles at different cholesterol molar ratios (Xchol) and developed a multi-stage lab-on-a-chip method to accomplish for the first time the separation of a three-vesicle mixture. Using Xchol = 0.1, 0.2, and 0.3 vesicles, we obtained efficiencies exceeding 93%. The simplicity, rapidity, and label-free nature of this approach holds promise as a diagnostic and separation tool for cells affected by diseases that affect the stiffness and extracellular vesicles such as exosomes and microvesicles.
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Gallagher, Jane. "Protein nanoparticle conjugates for use in bioanalytical applications." Thesis, University of Strathclyde, 2011. http://oleg.lib.strath.ac.uk:80/R/?func=dbin-jump-full&object_id=17065.
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Sentic, Milica. "Electrogenerated chemiluminescence : from mechanistic insights to bioanalytical applications." Thesis, Bordeaux, 2015. http://www.theses.fr/2015BORD0307/document.
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La chimiluminescence électrogénérée (ECL) est une technique analytique puissante exploitée pour la détection autant au niveau industriel que dans le domaine de la recherche scientifique ou du diagnostic clinique. La sensibilité élevée et la bonne sélectivité de cette technique font de l'ECL une méthode analytique de choix pour un large éventail d'applications, dont la plus importante est son utilisation commerciale dans un grand nombre de tests immunologiques à base de billes fonctionnalisées. Dans cette thèse, nous avons cherché à étudier le phénomène ECL et son application pour le développement de nouvelles techniques analytiques.Dans la première partie de ce travail, nous utilisons les techniques d'imagerie pour étudier les mécanismes ECL se produisant sur les billes utilisées pour les tests immunologiques. La cartographie de la réactivité au niveau d'une seule microparticule fonctionnalisée avec un complexe de ruthénium fournit une nouvelle stratégie visant à tester l'efficacité du co-réactif et montre des effets optiques associés de focalisation.Dans la deuxième partie, la conception d'un test immunologique pour la détection de l'anti-transglutaminase pour le diagnostic de la maladie coeliaque est présentée en utilisant des ensembles de nanoélectrodes comme plates-formes bioélectroanalytiques. Nous avons également étudié les caractéristiques de l'ECL générée par des réseaux de nanoélectrodes dopées au bore-diamant en tant que matériaux prometteurs pour des applications biologiques ainsi que l'efficacité ECL de deux co-réactifs sur ces réseaux.L'électrochimie bipolaire est un processus sans contact que nous avons exploité pour contrôler le mouvement d'objets conducteurs exposés à un champ électrique en l'absence de contact ohmique direct. Dans la troisième partie de ma thèse, nous présentons l'ECL couplée à l'électrochimie bipolaire pour le suivi d’objets autonomes luminescents. Nous avons élargi ce concept à la détection enzymatique dynamique de glucose en utilisant l'émission de lumière ECL comme signal analytiqueElectrogenerated chemiluminescence (ECL) is a powerful analytical technique exploited for clinical, industrial and research applications. The high sensitivity and good selectivity, makes ECL a tool-of-choice analytical method for a broad range of assays, most importantly for a large number of commercialized bead-based immunoassays. In the present thesis, we aimed to study the ECL phenomenon and its application in development of new analytical methods.In the first part of this work, we used an imaging technique to investigate the ECL mechanisms operating in bead-based assays. Spatial reactivity mapping at the level of a single functionalised bead provides a new strategy to test the co-reactant efficiency and shows associated optical focusing effects.In the second part, the design of a novel anti-transglutaminase ECL immunoassay for celiac disease diagnostic is shown using nanoelectrode ensembles as bioelectroanalytical platforms. We also studied the characteristics of ECL generated by arrays of boron-doped-diamond nanoelectrodes (BDD NEAs) as a promising materials for bioapplications. The ECL efficiency of two co-reactants at BDD NEAs was investigated.Finally, bipolar electrochemistry is a ‘‘wireless’’ process that was exploited for the controlled motion of conductive objects exposed to an electric field in the absence of direct ohmic contact. In the third part of the thesis, we report ECL coupled to bipolar electrochemistry for tracking the autonomous trajectories of swimmers by light emission. We further expanded this concept for dynamic enzymatic sensing of glucose concentration gradient using ECL light emission as an analytical readout
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Gruenhagen, Jason Alan. "Bioanalytical Applications of Real-Time ATP Imaging Via Bioluminescence." Washington, D.C. : Oak Ridge, Tenn. : United States. Dept. of Energy. Office of Science ; distributed by the Office of Scientific and Technical Information, U.S. Dept. of Energy, 2003. http://www.osti.gov/servlets/purl/822057-FTilZ3/native/.
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Thesis (Ph.D.); Submitted to Iowa State Univ., Ames, IA (US); 12 Dec 2003.Published through the Information Bridge: DOE Scientific and Technical Information. "IS-T 2604" Jason Alan Gruenhagen. 12/12/2003. Report is also available in paper and microfiche from NTIS.
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Wimalasena, Rohan Lalith. "Preparation and characterization of immunochemical reagents for bioanalytical applications." Diss., The University of Arizona, 1991. http://hdl.handle.net/10150/185457.
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Immunological reagents were prepared and characterized for the development of analytical methodology in bioanalytical research. Monoclonal antibodies to glucose oxidase (E.C. 1.1.3.4) from Aspergillus niger were prepared with apoenzyme as the antigen. Five of these antibodies, all of the IgG, subisotype, were further characterized. The carbohydrate moiety of the enzyme is not immunogenic. Binding of the five antibodies to the enzyme had no detectable effect on its catalytic properties. All the antibodies are shown to be directed towards segmental epitopes of the enzyme, not involving the carbohydrate moiety. Each enzyme subunit has more than one non-overlapping epitope. All five antibodies bound enzyme in a non-native conformation when coated on ELISA plates in preference to the native solution conformation. The importance of having a solution phase screening procedure for monoclonal antibodies is demonstrated. Factors affecting the specific activity of immobilized antibodies and their biologically active fragments were studied with goat anti-mouse and goat anti-human IgG. Antibodies were immobilized on HW 65 polymeric support matrix activated with carbonyldiimidazole, hydrazide and iodoacetic acid. The most significant factors influencing the specific activity of stochastic coupling of antibodies are multisite attachment, multiple orientations, and steric hindrance imposed by crowding of antibody and the size of the antigen. With oriented immobilization the specific activity is affected only by steric hindrance. The specific activity of immunosorbents prepared by immobilization of F(ab') fragments can be improved to almost 100% by limiting the amount of protein immobilization and the size of the antigen. The present study shows the protocols for optimizing immobilized antibody performance. Preparation of fragments of immunoglobulin were studied. Within the same species different antibodies showed different sensitivities to proteolytic cleavage by pepsin. A rapid, simple, high performance size exclusion chromatographic method was developed to monitor the reaction progress. Conditions must be optimized for each antibody in the preparation of F(ab')₂. Preparation of F(ab') from F(ab')₂ shows that 10-15% of goat anti-mouse F(ab')₂ was resistant to reduction. The procedure causes reduction of disulfide bonds other than the inter-heavy chain disulfide bonds.
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Johannesson, Nina. "Column Development in Capillary Electrophoresis and Electrochromatography for Bioanalytical Applications." Doctoral thesis, Uppsala universitet, Analytisk kemi, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7124.
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Analysis of biological samples can be a difficult task. This thesis covers a broad aspect of the analytical areas of capillary electrophoresis (CE) and capillary electrochromatography (CEC) in combination with mass spectrometry (MS) that are of great importance for achieving fast, accurate and sensitive bioanalyses. A significantly time reduced and automated system for sample cleanup was developed to greatly simplify the pretreatment process of biological samples with a complex matrix. Desalting and preconcentration of species in urine was conducted and the limit of detection for the antidepressant escitalopram was lowered 10 times. This extraction devise was also successfully incorporated in a chip based platform for the possibility to be a part of multidimensional separation systems. The reduced risk of sample loss leads to improved detection limits, which are usually one the most challenging parts when working with bioanalyses. In the area of separation, a monomer surface with tailored hydrophobicity was developed to achieve rapid, high efficient separations of complex mixtures. Within five minutes a tryptic digest of a protein could be separated and then identified by a Mascot search. The applications addressed have been focused on medical conditions which are of highest interest for both physicians and patients. A high throughput analysis of the kynurenine metabolites with CE-MS offers a new method to rapidly examine samples from patients with neurological disorders. A screening study of possible biomarkers for the two different types of appendicitis, gangraenous and phlegmonous was conducted. Indicative patterns were found for both pre and post surgery of the two types of inflammation as well as between them. The divergences were traced back to the MS peaks obtained in the CE- and CEC-MS setups as possible biomarkers for the two forms of appendicitis. A preliminary study of polycystic ovary syndrome also offered some valuable results for future biomarker identification.
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Wicks, Arriel. "Luminescent Quantum Dot and Protein Composite Nanoparticles for Bioanalytical Applications." ScholarWorks@UNO, 2010. http://scholarworks.uno.edu/td/1149.
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The first project focused on the preparation, characterization, and application of dual emission quantum dot encoded mesoporous silica microparticles. The quantum dots were added in precisely controlled ratios and were stably encapsulated within the pores of the silica. Several experiments were performed to test the superior stability of the quantum dot-silica composites over dye-loaded silica particles. The composite particles exhibited very high fluorescence, were functionalized with antibodies, and were used as signal transducers for the detection of a protein expressed by breast cancer cells. The second project focused in more detail on the detection capabilities of the quantum dot-silica composites. Three different types of quantum dot-silica composites were prepared. Each type was loaded with a separate type of quantum dot with distinct emission wavelengths and was functionalized with separate antibodies for detection of three different breast cancer biomarkers. These three composite sensors were used together for the simultaneous detection of each of the breast cancer markers. The initial strategy utilized the direct detection method in which the antigen is nonspecifically adsorbed to a glass plate. An improved second strategy was more sensitive and used a capture antibody which was covalently bound to a glass plate to immobilize the antigen. The third project focused on the preparation and application of magnetic, fluorescent human serum albumin nanoparticle composites. A fluorescent drug analogue and iron oxide nanoparticles were encapsulated into 100 nm human serum albumin nanoparticles. The advantage of these composite particles is that they could be used as a theranostic tool which could target, detect, and treat diseased tissue in a single application. Release of the drug analogue from the nanocomposites was achieved by addition of proteolytic enzymes that are expressed or overexpressed in cancer cells. The temporal release of the fluorescent drug analogue was measured as a function of enzyme concentration. The amount of drug released was directly proportional to enzyme concentration.
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Qamar, Ahmad Zaman. "DEVELOPING WAX-ON-PLASTIC PLATFORMS FOR BIOANALYTICAL AND BIOMEDICAL APPLICATIONS." OpenSIUC, 2019. https://opensiuc.lib.siu.edu/dissertations/1759.
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Developing microdevices on flexible material attracts scientific community to explore applications in different aspects of health and point of care diagnostics. Flexible substrates offer unique characteristics such as flexibility, stretchability, portability, low-cost, and simple fabrication. Fabrication of cost-effective paper-based analytical devices by wax printing has recently become popular using cellulose filter papers. Paper-based devices need higher temperature to form hydrophobic barrier across paper substrate, rely on large working channels (≥ 500 μm) for liquid handling, and exhibit lower efficiency (~50%) of sample mobility. Such limitations confine applications of wax-based fabrication. In this dissertation, we report printability, fidelity, and applications of wax micropatterns on polyethylene terephthalate-based substrate (PET), which is a a non-cellulosic, non-fibrous, and non-porous material. Resolution, sustainability against heat and biocompatibility was tested on wax micro-features. The patterned devices were explored for variety of applications.First, wax microwells on PET showed mouse embryonic stem cell (mESC) self-renewal or direct differentiation. Second, microfluidic flow was demonstrated on wax printed microchannels on PET which was used to develop distance-based assay. Third, fluidic properties of trinucleotide repeat sequences were investigated on wax microchannels. Fourth, multilayer wax-on-plastic device was fabricated using wax printing with hand painting of conductive materials for electrochemical immunosensing.
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Cai, Xiaohan. "¿¿¿¿¿¿Development of Bioanalytical Methods for Clinical Applications and Drug Screening." Cleveland State University / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=csu1314982525.
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Kumar, Suresh. "Design, Fabrication, and Optimization of Miniaturized Devices for Bioanalytical Applications." BYU ScholarsArchive, 2015. https://scholarsarchive.byu.edu/etd/5979.
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My dissertation work integrates the techniques of microfabrication, micro/nanofluidics, and bioanalytical chemistry to develop miniaturized devices for healthcare applications. Semiconductor processing techniques including photolithography, physical and chemical vapor deposition, and wet etching are used to build these devices in silicon and polymeric materials. On-chip micro-/nanochannels, pumps, and valves are used to manipulate the flow of fluid in these devices. Analytical techniques such as size-based filtration, solid-phase extraction (SPE), sample enrichment, on-chip labeling, microchip electrophoresis (µCE), and laser induced fluorescence (LIF) are utilized to analyze biomolecules. Such miniaturized devices offer the advantages of rapid analysis, low cost, and lab-on-a-chip scale integration that can potentially be used for point-of-care applications.The first project involves construction of sieving devices on a silicon substrate, which can separate sub-100-nm biostructures based on their size. Devices consist of an array of 200 parallel nanochannels with a height step in each channel, an injection reservoir, and a waste reservoir. Height steps are used to sieve the protein mixture based on size as the protein solution flows through channels via capillary action. Proteins smaller than the height step reach the end of the channels while larger proteins stop at the height step, resulting in separation. A process is optimized to fabricate 10-100 nm tall channels with improved reliability and shorter fabrication time. Furthermore, a protocol is developed to reduce the electrostatic interaction between proteins and channel walls, which allows the study of size-selective trapping of five proteins in this system. The effects of protein size and concentration on protein trapping behavior are evaluated. A model is also developed to predict the trapping behavior of different size proteins in these devices. Additionally, the influence of buffer ionic strength, which can change the effective cross-sectional area of nanochannels and trapping of proteins at height steps, is explored in nanochannels. The ionic strength inversely correlates with electric double layer thickness. Overall, this work lays a foundation for developing nanofluidic-based sieving systems with potential applications in lipoprotein fractionation, protein aggregate studies in biopharmaceuticals, and protein preconcentration. The second project focuses on designing and developing a microfluidic-based platform for preterm birth (PTB) diagnosis. PTB is a pregnancy complication that involves delivery before 37 weeks of gestation, and causes many newborn deaths and illnesses worldwide. Several serum PTB biomarkers have recently been identified, including three peptides and six proteins. To provide rapid analysis of these PTB biomarkers, an integrated SPE and µCE device is assembled that provides sample enrichment, on-chip labeling, and separation. The integrated device is a multi-layer structure consisting of polydimethylsiloxane valves with a peristaltic pump, and a porous polymer monolith in a thermoplastic layer. The valves and pump are fabricated using soft lithography to enable pressure-based sample actuation, as an alternative to electrokinetic operation. Porous monolithic columns are synthesized in the SPE unit using UV photopolymerization of a mixture consisting of monomer, cross-linker, photoinitiator, and various porogens. The hydrophobic surface and porous structure of the monolith allow both protein retention and easy flow. I have optimized the conditions for ferritin retention, on-chip labelling, elution, and µCE in a pressure-actuated device. Overall functionality of the integrated device in terms of pressure-controlled flow, protein retention/elution, and on-chip labelling and separation is demonstrated using a PTB biomarker (ferritin). Moreover, I have developed a µCE protocol to separate four PTB biomarkers, including three peptides and one protein. In the future, an immunoaffinity extraction unit will be integrated with SPE and µCE to enable rapid, on-chip analysis of PTB biomarkers. This integrated system can be used to analyze other disease biomarkers as well.
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Kim, Youngmi. "Molecular engineering of multifunctional nucleic acid probes for bioanalytical and biomedical applications." [Gainesville, Fla.] : University of Florida, 2008. http://purl.fcla.edu/fcla/etd/UFE0023647.
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Kim, Junseok. "Bioanalytical Applications of Intramolecular H-Complexes of Near Infrared Bis(Heptamethine Cyanine) Dyes." Digital Archive @ GSU, 2008. http://digitalarchive.gsu.edu/chemistry_diss/23.
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This dissertation describes the advantages and feasibility of newly synthesized near-infrared (NIR) bis-heptamethine cyanine (BHmC) dyes for non-covalent labeling schemes. The NIR BHmCs were synthesized for biomolecule assay. The advantages of NIR BHmCs for biomolecule labeling and the instrumental advantages of the near-infrared region are also demonstrated. Chapter 1 introduces the theory and applications of dye chemistry. For bioanalysis, this chapter presents covalent and non-covalent labeling. The covalent labeling depends on the functionality of amino acids and the non-covalent labeling relies on the binding site of a protein. Due to the complicated binding process in non-covalent labeling, this chapter also discusses the binding equilibria in spectroscopic and chromatographic analyses. Chapter 2 and 3 evaluate the novel BHmCs for non-covalent labeling with human serum albumin (HSA) and report the influence of micro-environment on BHmCs. The interesting character of BHmCs in aqueous solutions is that the dyes exhibit non- or low-fluorescence compared to their monomer counterpart, RK780. It is due to their H-type closed clam-shell form in the solutions. The addition of HSA or organic solvents opens up the clam-shell form and enhances fluorescence. The binding equilibria are also examed. Chapter 4 provides a brief introduction that summaries the use of capillary electrophoresis (CE), and offers a detailed instrumentation that discusses the importance and advantage of a detector in NIR region for CE separation. Chapter 5 focuses on the use of NIR cyanine dyes with capillary electrcophoresis with near-infrared laser induce fluorescence (CE-NIR-LIF) detection. The NIR dyes with different functional groups show that RK780 is a suitable NIR dye for HSA labeling. The use of BHmCs with CE-NIR-LIF reduces signal noises that are commonly caused by the interaction between NIR cyanine dyes and negatively charged capillary wall. In addition, bovine carbonic anhydrase II (BCA II) is applied to study the influence of hydrophobicity on non-covalent labeling. Finally, chapter 6 presents the conformational dependency of BHmCs on the mobility in capillary and evaluates the further possibility of BHmCs for small molecule detection. Acridine orange (AO) is used as a sample and it breaks up the aggregate and enhances fluorescence. The inserted AO into BHmC changes the mobility in capillary, owing to the conformational changes by AO.
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Mezzanotte, Laura <1982>. "Bioanalytical applications of multicolour bioluminescence imaging: new tools for drug discovery and development." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amsdottorato.unibo.it/3536/.
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The subject of this thesis is multicolour bioluminescence analysis and how it can provide new tools for drug discovery and development.The mechanism of color tuning in bioluminescent reactions is not fully understood yet but it is object of intense research and several hypothesis have been generated. In the past decade key residues of the active site of the enzyme or in the surface surrounding the active site have been identified as responsible of different color emission. Anyway since bioluminescence reaction is strictly dependent from the interaction between the enzyme and its substrate D-luciferin, modification of the substrate can lead to a different emission spectrum too. In the recent years firefly luciferase and other luciferases underwent mutagenesis in order to obtain mutants with different emission characteristics. Thanks to these new discoveries in the bioluminescence field multicolour luciferases can be nowadays employed in bioanalysis for assay developments and imaging purposes. The use of multicolor bioluminescent enzymes expanded the potential of a range of application in vitro and in vivo. Multiple analysis and more information can be obtained from the same analytical session saving cost and time. This thesis focuses on several application of multicolour bioluminescence for high-throughput screening and in vivo imaging. Multicolor luciferases can be employed as new tools for drug discovery and developments and some examples are provided in the different chapters. New red codon optimized luciferase have been demonstrated to be improved tools for bioluminescence imaging in small animal and the possibility to combine red and green luciferases for BLI has been achieved even if some aspects of the methodology remain challenging and need further improvement. In vivo Bioluminescence imaging has known a rapid progress since its first application no more than 15 years ago. It is becoming an indispensable tool in pharmacological research. At the same time the development of more sensitive and implemented microscopes and low-light imager for a better visualization and quantification of multicolor signals would boost the research and the discoveries in life sciences in general and in drug discovery and development in particular.
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Scott, Daniel F. "DEVELOPMENT OF LUMINESCENT SENSING SYSTEMS WITH CLINICAL APPLICATIONS." UKnowledge, 2011. http://uknowledge.uky.edu/gradschool_diss/184.
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As the move towards the miniaturization of many diagnostic and detection systems continues, the need for increasingly versatile yet sensitive labels for use in these systems also grows. Luminescent reporters provide us with a solution to many of the issues at hand through their unique and favorable characteristics. Bioluminescent proteins offer detection at extremely low concentrations and no interference from physiological fluids leading to excellent detection limits, while the vast number of fluorescent proteins and molecules available allows the opportunity to select a tailored reporter for a specific task. Both provide relatively simply instrumentation requirements and have exhibited great promise with many of the miniaturized systems such as lab-on-a-chip and lab-on-a-CD designs. Herein, we describe the novel employment of luminescent reporters for four distinct purposes. First off, by combining both time and wavelength resolution we have expanded the multiplexing capabilities of the photoprotein aequorin beyond duel-analytes, demonstrating the ability to simultaneously detect three separate analytes. Three semi-synthetic aequorin proteins were genetically conjugated to three pro-inflammatory cytokines (interleukins 1, 6, and 8) resulting in aequorin labeled cytokines with differing emission maxima and half lives to allow for the simultaneous detection of all three in a single solution through the elevated physiological concentration range. Secondly a semi-synthetic aequorin variant has been genetically enhanced to serve as an immunolabel and exhibited the ability to sensitively detect the acute myeloid leukemia marker, CD33, down to the attomole level in addition to improving aequorin imaging capabilities. In the third example, the aequorin complex was rationally, genetically split into two parts and attached to the termini of the cAMP selective cAMP receptor protein (CRP) creating a genetically fused molecular switch. The conformational change experienced by CRP upon the binding of cAMP translates into a loss of bioluminescent signal from aequorin and has shown the ability to respond linearly to cAMP over several orders of magnitude. Lastly, through custom design, a reagentless, portable, fluorescent fiber optic detection system has been developed, capable of being integrated into the body through a heart catheter. The system was able to respond to changes in potassium concentration selectively, reproducibly and reversibly with a fast response time of one minute.
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Chapman, Gala M. "Spectroscopic Studies of Carbocyanine and 2,4,6- Trisubstituted Pyridine Dyes for Bioanalytical and pH Indicating Applications." Digital Archive @ GSU, 2011. http://digitalarchive.gsu.edu/chemistry_theses/47.
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In part A, the effect of varying short-chain alkyl substitution on the spectroscopic properties of cyanine dyes was examined. Molar absorptivities and quantum yields were determined for groups of pentamethine and heptamethine dyes for which the substitution of the indole nitrogen was varied. For both sets of dyes, increasing alkyl chain length did not significantly change quantum yield or molar absorptivity. These results may be useful in designing new cyanine dyes.
In part B, the effect of structure on the suitability of 2,4,6-trisubstituted pyridines as color pH indicators was studied by determining spectral effects of protonation, molar absorptivities, pKa values, and the structural origin of the spectral behavior. Good color indicating properties result from aniline substitution at the 4 position of pyridine and electron donating substitution at the 2 and 6 positions of pyridine, which provide a strong red shift in the spectra and greater red shifted peak absorptivity, respectively.
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Knittle, James Eugene. "Bioanalytical and photophysical applications of sensitive nonlinear wave-mixing spectroscopy based on laser-induced gratings /." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2004. http://wwwlib.umi.com/cr/ucsd/fullcit?p3138952.
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Debela, Mehdi Ahmed. "Bioanalytical applications of polyoxometalates using molecular tools for detection of DNA and SNPs in primer extension." Doctoral thesis, Universitat Rovira i Virgili, 2014. http://hdl.handle.net/10803/277386.
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L'objectiu general de la present tesi és el desenvolupament d'una plataforma per la detecció electroquímica de mutacions genètiques emprant un mètode d'alta especificitat basat en l'extensió de seqüències d'ADN, anomenades cebadors, disposades organitzadament sobre una superfície (APEX). Per tant el treball inclou la preparació de molècules que constitueixen marcadors electroquímics i que van ser posteriorment enllaçades de manera covalent a les quatre bases d'ADN i, a més, el desenvolupament d'una plataforma robusta resistent a altes temperatures, que són necessàries durant la incorporació per la via enzimàtica de les bases d'ADN marcades. En la primera part del present treball, es van sintetitzar dues classes de polioxometalats (anomenats Keggin and Dawson POM) i van ser covalentment enllaçats a seqüències d'ADN. A més, al costat d'altres marcadors electroquímics, van ser enllaçats covalentment a bases d'ADN individuals per a la detecció de mutacions genètiques. A la segona part es va desenvolupar un nou mètode per a la funcionalització de superfícies de carbó vitri basat en processos electroquímics consecutius d'hidrogenació i cloració. Aquestes superfícies modificades van ser utilitzades per a la immobilització de molècules tioladas, incloent ADN tiolado. Finalment, les molècules sintetitzades i les superfícies modificades van ser usades per a la detecció electroquímica de mutacions genètiques seguint la metodologia APEX.El objetivo general de la presente tesis es el desarrollo de una plataforma para la detección electroquímica de mutaciones genéticas empleando un método de alta especificidad basado en la extensión de secuencias de ADN, llamadas cebadores, dispuestas organizadamente sobre una superficie (APEX). Por tanto el trabajo incluye la preparación de moléculas que constituyen marcadores electroquímicos y que fueron posteriormente enlazadas de forma covalente a las cuatro bases de ADN y, además, el desarrollo de una plataforma robusta resistente a altas temperaturas, que son necesarias durante la incorporación por la via enzimática de las bases de ADN marcadas. En la primera parte del presente trabajo, se sintetizaron dos clases de polioxometalatos (llamados Keggin and Dawson POM) y fueron covalentemente enlazados a secuencias de ADN. Además, junto a otros marcadores electroquímicos, fueron enlazados covalentemente a bases de ADN individuales para la detección de mutaciones genéticas. En la segunda parte se desarrolló un nuevo método para la funcionalización de superficies de carbón vítreo basado en procesos electroquímicos consecutivos de hidrogenación y cloración. Estas superficies modificadas fueron utilizadas para la inmovilización de moléculas tioladas, incluyendo ADN tiolado. Finalmente, las moléculas sintetizadas y las superficies modificadas fueron usadas para la detección electroquímica de mutaciones genéticas siguiendo la metodología APEX.
This PhD work was started with the intention of developing platform for electrochemical detection of mutations using highly specific method called arrayed primer extension. So the work involves the preparation of labels to be linked with the four different DNA bases and the preparation of robust platform which can withstand the high temperature during enzymatic incorporation of the labelled DNA bases. The first part of this work deals of the preparation of compounds called polyoxometalates (Keggin and Dawson types) and their covalent attachment to DNA sequence and also, to DNA single bases along with other organic redox labels for mutation detection. In the second part while trying to achieve a thermally stable surface tethered DNA probes, a new method of surface functionalisation based on electrochemical hydrogenation followed by electrochemical chlorination of glassy carbon was developed. The so prepared surfaces were used for immobilisation of thiolated molecules including thiolated DNA. Finally the synthesised molecules and modified surfaces were used for electrochemical detection of genetic mutation following the APEX protocol.
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Atherton, Adrian Ashley. "Nonlinear wave-mixing spectroscopic methods for bioanalytical and biophysical applications with sensitive detection at the single cell level." Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2006. http://wwwlib.umi.com/cr/ucsd/fullcit?p3230037.
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Thesis (Ph. D.)--University of California, San Diego and San Diego State University, 2006.Title from first page of PDF file (viewed November 17, 2006). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references.
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Pan, Tao. "Towards early stage disease detection in microdevices : fabrication and testing of micro total analysis systems for bioanalytical applications / /." Diss., CLICK HERE for online access, 2007. http://contentdm.lib.byu.edu/ETD/image/etd1836.pdf.
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Pan, Tao. "Towards Early State Disease Detection in Microdevices: Fabrication and Testing of Micro Total Analysis Systems for Bioanalytical Applications." BYU ScholarsArchive, 2007. https://scholarsarchive.byu.edu/etd/1351.
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The past few years have seen a rapid expansion in interest in the characterization of the entire complement of proteins, or proteome. Micro total analysis systems (μTAS) are an emerging promising method, offering rapid, sensitive and low sample consumption separations. I have demonstrated microchip capillary electrophoresis (CE) devices made of CaF2. New methods have been developed for micromachining enclosed capillaries in CaF2. CE analysis of fluorescently labeled amino acids was used to illustrate bioanalytical applications of these microdevices. Initial on-chip infrared spectroscopy results for qualitative analyte identification were achieved in microfluidic CaF2 channels. I have also shown the evaluation of poly(methylmethacrylate) (PMMA) and thermoset polyester (TPE) microchips for use in protein profiling. To improve separation efficiency and reduce protein adsorption, dynamic coating and poly(ethylene glycol) (PEG) grafting using atom transfer radical polymerization (ATRP) have been used in PMMA microdevices. Proteins, peptides and protein digests have been separated electrophoretically in these PMMA microchips. My results demonstrate that PMMA microdevices should be well suited as microfluidic systems for high performance separations of complex biological mixtures. In-channel ATRP has been developed for the surface modification of TPE microdevices. Characterization indicates that PEG-modified microchannels have much lower and more pH-stable electroosmotic flow, more hydrophilic surfaces and reduced nonspecific protein adsorption. CE of amino acid and peptide mixtures in these PEG-modified TPE microchips had good reproducibility. Phosducin-like protein and phosphorylated phosducin-like protein were also separated to measure the phosphorylation efficiency. My results show that PEG-grafted TPE microchips have broad potential application in biomolecular analysis. Cancer marker analysis is important for medical research and applications. I report a method that can covalently attach appropriately oriented antibodies of interest on monolith surfaces. To reduce nonspecific adsorption, protein solutions were used to effectively block the monolith surface. Selective preconcentration and elution of human chorionic gonadotropin have been performed in my affinity columns, demonstrating that this type of system should have promising applications in cancer marker detection.
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Gamez, Gerardo. "Advances in analytical spectrochemistry with ionized gases. I. Improved fundamental understanding through laser based techniques. II. Novel bioanalytical applications." [Bloomington, Ind.] : Indiana University, 2006. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3223049.
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Thesis (Ph. D.)--Indiana University, Dept. of Chemistry, 2006."Title from dissertation home page (viewed June 28, 2007)." Source: Dissertation Abstracts International, Volume: 67-06, Section: B, page: 3105. Adviser: Gary M. Hieftje.
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Tanase, Otilia Maya <1982>. "Development and applications of hollow fiber flow field-flow fractionation in the bioanalytical field. Studies of aggregation phenomena in complex protein samples." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6277/.
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Recent advances in the fast growing area of therapeutic/diagnostic proteins and antibodies - novel and highly specific drugs - as well as the progress in the field of functional proteomics regarding the correlation between the aggregation of damaged proteins and (immuno) senescence or aging-related pathologies, underline the need for adequate analytical methods for the detection, separation, characterization and quantification of protein aggregates, regardless of the their origin or formation mechanism.
Hollow fiber flow field-flow fractionation (HF5), the miniaturized version of FlowFFF and integral part of the Eclipse DUALTEC FFF separation system, was the focus of this research; this flow-based separation technique proved to be uniquely suited for the hydrodynamic size-based separation of proteins and protein aggregates in a very broad size and molecular weight (MW) range, often present at trace levels. HF5 has shown to be (a) highly selective in terms of protein diffusion coefficients, (b) versatile in terms of bio-compatible carrier solution choice, (c) able to preserve the biophysical properties/molecular conformation of the proteins/protein aggregates and (d) able to discriminate between different types of protein aggregates.
Thanks to the miniaturization advantages and the online coupling with highly sensitive detection techniques (UV/Vis, intrinsic fluorescence and multi-angle light scattering), HF5 had very low detection/quantification limits for protein aggregates. Compared to size-exclusion chromatography (SEC), HF5 demonstrated superior selectivity and potential as orthogonal analytical method in the extended characterization assays, often required by therapeutic protein formulations. In addition, the developed HF5 methods have proven to be rapid, highly selective, sensitive and repeatable. HF5 was ideally suitable as first dimension of separation of aging-related protein aggregates from whole cell lysates (proteome pre-fractionation method) and, by HF5-(UV)-MALS online coupling, important biophysical information on the fractionated proteins and protein aggregates was gathered: size (rms radius and hydrodynamic radius), absolute MW and conformation.
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Souza, Glalci Alves de. "Estudo mecanístico do sistema peroxioxalato com diferentes catalisadores." Universidade de São Paulo, 2017. http://www.teses.usp.br/teses/disponiveis/46/46136/tde-20092017-104722/.
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A reação peroxioxalato é o sistema quimiluminescente não enzimático de maior eficiência, alcançando rendimentos quânticos de até 50%. A quimiluminescência deste sistema vem sendo amplamente utilizada em aplicações analíticas e bioanalíticas visando a detecção de analitos de interesses biológicos e medicinais. O presente trabalho consistiu em se estudar a reação peroxioxalato com diferentes catalisadores e determinação de seu mecanismo em condições experimentais distintas das que vinham sendo estudadas até o momento, visando sua adaptação a meios aquosos para futuras aplicações. Foi estudada a utilização do salicilato de sódio como catalisador deste sistema em meio puramente orgânico, em substituição ao imidazol, e verificou-se a atuação do salicilato como catalisador básico geral e/ou específico. O rendimento quântico máximo desta transformação foi da ordem de 10-3 E mol-1. Ensaios cinéticos de emissão também foram realizados com ésteres oxálicos de diferentes reatividades em um sistema binário composto por 1,2-dimetoxietano/água contendo tampão fosfato como catalisador, em diferentes valores de pH. Nestas condições se observou a ocorrência de catálise ácida geral e catálise básica geral, uma vez que as constantes de velocidade se mostraram dependentes da concentração de tampão no meio reacional. Além disso, também se utilizou salicilato de sódio como catalisador no sistema binário 1,2-dimetoxietano/água, o qual também apresentou um papel catalítico importante no percurso desta transformação com diferentes ésteres oxálicos. Verificou-se nestes estudos a boa reprodutibilidade da reação peroxioxalato com salicilato de sódio como catalisador em meios parcialmente aquosos, que pode ampliar a utilização deste sistema quimiluminescente em aplicações analíticas e bioanalíticas. Quando se realizou os ensaios em um meio majoritariamente aquoso, o salicilato não se mostrou um catalisador eficiente, porém, mesmo sem catalisador, a reação peroxioxalato se mostrou reprodutível nestas condiçõesThe peroxyoxalate reaction is the non-enzymatic chemiluminescence system with the highest efficiency, achieving quantum yields of up to 50%. The chemiluminescence of this system has been widely used in analytical and bionanalytical applications in order to detect analytes of biological and medicinal interests. The present work consisted in a study of the peroxyoxalate reaction with different catalysts and the determination of its mechanism in experimental conditions different from those studied before, aiming its adaptation to aqueous media for future applications. The use of sodium salicylate as base catalyst for this system in pure organic medium, in substitution to imidazole, was studied and shown that salicylate acts as a general and/or specific base catalyst. The maximum quantum yield obtained for the transformation in these conditions was in the order of 10-3 E mol-1. Emission experiments were also performed in a binary solvent system composed of 1,2- dimethoxyethane/aqueous phosphate buffer at different pH values as catalyst, using oxalic esters with different reactivities. In these conditions the occurrence of general acid catalysis and general base catalysis was observed, since the rate constants proved to be dependent on the buffer concentration. In addition, sodium salicylate was also used as catalyst in the binary 1,2-dimethoxyethane/water system with different oxalic esters, indicating its important catalytic role in the transformation. These studies allowed it to establish a reproducible peroxyoxalate system in partially aqueous media using sodium salicylate as base catalyst, which may increase the use of the chemiluminescence of this system in analytical and bioanalytical applications. However, when the experiments were performed in a medium containing mostly water, salicylate did not act as an efficient catalyst in these conditions. Even in the absence of catalyst, the reaction proved to be reproducible in the medium containing mostly water as solvent
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Rainville, Paul. "The investigation and application of alternative strategies for LC/MS based bioanalytical research." Thesis, King's College London (University of London), 2013. https://kclpure.kcl.ac.uk/portal/en/theses/the-investigation-and-application-of-alternative-strategies-for-lcms-based-bioanalytical-research(9e73e438-c941-4e4f-8502-69f6e4decef9).html.
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The work presented here demonstrates two unique approaches in the application of LC/MS for the analysis of drugs and endogenous compounds in biofluids. First, the use of basic aqueous mobile phases in conjunction with both methanol and acetonitrile by LC/MS operating in electrospray positive ionization mode was investigated. Second, the design and development of a prototype ceramic micro-fluidic device (CMFD) and optimized MS source was carried out. The CMFD was packed with sub 2 μm bridged ethyl hybrid (BEH) chromatographic particles that could withstand operating pressures of up to 12,000 psi. The MS source was built to operate in both positive and negative electrospray ionization mode with the operating flow rates corresponding to the 300 um i.d. of the CMFD. The results generated from studies utilizing the addition of a base, such as ammonium hydroxide to modify the aqueous mobile phase, showed significant benefits for LC/MS/MS based bioanalytical assays when analyzed with electrospray positive ionization mode. Increases in the signal-to-noise values were observed for twenty-two out of twenty-four of the probe pharmaceuticals tested. Increase in the chromatographic retention of poorly retained compounds was also observed however, this increase in analyte retention did not occur for many highly polar compounds that eluted in the column void when chromatographed with the basic mobile phase conditions. The effect of the pH of the mobile phase further showed that the phospholipid fraction, present in protein precipitated plasma, was only slightly affected by the change in mobile phase pH. It provided complementary data to that obtained with traditional acidic based mobile phases and results in an increased number of ions detected overall for metabonomic studies. Further it was observed that by changing the pH to a basic system it was possible to resolve compounds that had previously co-eluted on traditional acidic reversed-phase LC/MS. A 0.3 x 100 mm i.d. CMFD and compatible MS source was successfully designed for the analysis of biological samples. The device showed average chromatographic efficiencies of 9038 plates compared to 10219 plates for standard silica capillary columns. Gradient performance utilizing a diverse mix of compounds yielded a peak capacity of 55 as the average peak widths for all analytes was 0.11 minutes for a 6 minute separation. Resolution of the probe pharmaceutical alprazolam and associated hydroxylated metabolite was maintained between 1.2 and 1.5 for four different devices. Testing of the device with plasma samples prepared by protein precipitation resulted in over 1000 injections being carried out over approximately a one week period. The system was however unable to withstand the high pH (10.5) utilized in the previous study but could operate with a mobile phase pH of 10. The resulting MS source built to operate under these conditions and with the flow rates required to operate the 0.3 x 100 mm CMFD yielded signal to noise increase in the range of 8.3 to 38.6 for the molecules tested. The CMFD/MS system was successful in the analysis of biofluid samples in both ESI + and ESI – ionization modes and was shown to allow for the analysis of small sample injection volumes from plasma prepared by protein precipitation, and dried blood spots. This device was successfully utilized for the profiling of metabolites from the beta blocker drug, propranolol. Further separation of complex biofluid samples derived from axenic rats and bile from dogs again illustrated the separation and sensitivity capabilities offered by the CMFD/MS system.
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Römsing, Susanne. "Development and Validation of Bioanalytical Methods : Application to Melatonin and Selected Anti-Infective Drugs." Doctoral thesis, Uppsala universitet, Analytisk kemi, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-131519.
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This thesis describes bioanalytical methods for measuring melatonin and some anti-infective drugs in biological fluids. Solid-phase extraction (SPE) or protein precipitation was used for enrichment and purification of the analytes and Liquid Chromatography (LC) was used to analyze the samples. Developed methods were validated according to international guidelines. Melatonin is a hormone secreted by the pineal gland with a robust circadian rhythm. Bioanalytical methods for determination of melatonin in plasma and saliva have been developed which were used for monitoring melatonin levels in volunteers and patients suffering from sleep related diseases. Eflornithine (DFMO) is a chiral drug used for the treatment of human African trypanosomiasis. A bioanalytical method for determination of the DFMO enantiomers in plasma, after precolumn derivatization with o-phtalaldehyde and N-acetyl-L-cystein has been developed. The method has been used to study the L- and D-DFMO pharmacokinetics, in order to investigate the possible development of an oral treatment of DFMO. A method for simultaneous determination of three antiretroviral drugs i.e. Lamivudine (3TC), Zidovudine (AZT) and Nevirapine (NVP) in dried blood spots (DBS) was developed. The method was used for drug determination in two subjects after receiving standard antiretroviral treatment. The method seemed well suitable for the determination of 3TC and NVP and in some extent for AZT. Lumefantrine (LF) is one of the active components in a new fixed drug combination recommended by the WHO as a replacement to older drugs that has lost their effect. A method for the determination of LF in DBS was developed. The method is suitable for monitoring of drug treatment in rural settings. Tafenoquine is a new promising antimalarial drug under development. A method for the determination of Tafenoquine in plasma and in DBS is described. The method may be useful in future clinical studies in laboratory environment as well as in rural settings.Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 703
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Öhman, Daniel. "Bioanalytical development for application in therapeutic drug monitoring : focus on drugs used in psychiatry /." Linköping : Univ, 2003. http://www.bibl.liu.se/liupubl/disp/disp2003/med775s.pdf.
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Mak, Wing Cheung. "The applications of layer-by-layer technology in bioengineering and bioanalytics /." View abstract or full-text, 2004. http://library.ust.hk/cgi/db/thesis.pl?BIEN%202004%20MAK.
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Benabou, Zdaou Sanae. "Application of analytical and chemometric methodologies to study complex bioanalytical processes involving DNA i-motif structures." Doctoral thesis, Universitat de Barcelona, 2018. http://hdl.handle.net/10803/663796.
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The i-motif is a DNA structure formed by cytosine-rich sequences that consists of parallel- stranded duplexes held together by intercalated base pairs. The in vitro formation of this structure in DNA sequences corresponding to the promoter regions of several oncogenes, such as c-kit, c-myc or bcl-2, has been demonstrated. Recently, the first direct evidence for its in vivo presence in human cells and control regulatory functions has been proven. This structure is not only interesting from a biophysical and biomedical point of view, but also for their potential application in Analytical Chemistry or Nanotechnology.
The present Doctoral Thesis deals with the application of analytical and chemometric methodologies to study complex bioanalytical processes involving DNA i-motif structures. The sequences studied correspond to those found at cytosine-rich regions found near the promoter regions of the nmyc and SMARCA4 genes. On the one hand, the stability of the i- motif structures formed by these sequences according to variations of pH, temperature, ionic strength, or presence of ligands in steady-state conditions has been studied. On the other hand, the potential of ultrafast spectroscopies for the study of fast kinetic processes triggered by light has been evaluated. Through the Thesis, curve resolution methods, either based on soft-, hard- or hybrid-modelling have been used extensively to model the biochemical processes of interest.
The steady-state studies have demonstrated that the stability against pH or temperature variations of the three different kinds of cytosine-rich sequences mentioned above is strongly dependent on the number of the C·C+ base pairs, but also on the contribution of other factors, such as the base composition and length of the loops and the presence of additional stabilising structures (hairpins) in the DNA sequence. The studies performed at ultrafast time scales have revealed that the photochemical process induced by UV-lamp irradiation and monitored by rapid-scan FTIR involves the formation of dimeric photoproducts in folded and unfolded sequences. The study of processes monitored by time-resolved fluorescence in the scale of picoseconds has shown that i-motif relaxation is detected by the presence of fast lifetimes in the pH range between 4 and 6, associated with intrinsic conformational changes at the fluorescent site. In this last study, one or two
different i-motif structures have been detected in the nmyc and in the shortest DNA sequences studied, respectively.
Finally, the application of multivariate resolution methods, based either on hard- or soft- modelling, has allowed the recovery of valuable chemical information from evolutionary processes of DNA. Besides, the adaptation and application of hybrid hard- and soft- modelling has been shown to be a useful approach to detect intermediate temperature- dependent conformational transitions and to avoid the effect of baseline drifts in the estimation of the melting temperature, as well to retrieve rate constants from the kinetic information present in rapid-scan FTIR difference spectra.La estructura de l’ADN coneguda com “i-motif” es forma en seqüències riques en bases citosina (C). L’esquelet de l’i-motif està format per parells de bases C·C+ intercalats i estabilitzats per ponts d’hidrogen. S'ha demostrat la formació in vitro d'aquesta estructura en seqüències d'ADN corresponents a les regions promotores de diversos oncògens, com el c-kit, el c-myc o el bcl-2. Recentment, s'ha demostrat la primera evidència de la seva presència in vivo. La present Tesi Doctoral tracta de l'aplicació de metodologies analítiques i quimiomètriques per estudiar processos bioanalítics complexos que en els que intervenen aquestes estructures. Les seqüències estudiades corresponen a les regions promotores dels gens nmyc i SMARCA4. D'una banda, s'ha estudiat l'estabilitat de les estructures formades per aquestes seqüències segons variacions de pH, temperatura, força iònica o presència de lligands en condicions d'estat estacionari. D'altra banda, s'ha avaluat el potencial d'espectroscòpia ultraràpida per a l'estudi de processos cinètics ràpids provocats per la llum. Al llarg de la Tesi, els mètodes de resolució multivariant, ja sigui basats en models flexibles, rígids o híbrids, s'han utilitzat àmpliament per modelitzar els processos d'interès. Els estudis d'estat estacionari han demostrat que l'estabilitat davant el pH o les variacions de temperatura de les tres diferents seqüències riques en citosina esmentades anteriorment depèn molt del nombre de parells de bases de C·C+, però també de la contribució d'altres factors, com ara la composició de la base i la longitud dels bucles. A partir d'estudis ultraràpids, el procés fotoquímic induït per la irradiació de llum UV i l'IR d'escaneig ràpid s'ha relacionat amb la formació de fotoproductes dimèrics en seqüències plegades i desplegades. L'estudi dels processos seguits mitjançant fluorescència resolta en el temps ha demostrat l’existència de més d’una espècie associada amb l’estructura i-motif en el cas de les seqüencies curtes. Finalment, l'aplicació de mètodes de resolució multivariant, basats tant en models rígids o flexibles, han permès extreure informació química valuosa dels processos evolutius de l'ADN. A més, s'ha demostrat que l'adaptació i l'aplicació del modelatge híbrid ha permet calcular les constants cinètiques i detectar transicions dependents de la temperatura.
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Stumbaum, Mihaela [Verfasser]. "Development and bioanalytical application of affinity-mass spectrometry for identification and structural characterisation of protein-ligand interactions / Mihaela Stumbaum." Konstanz : Bibliothek der Universität Konstanz, 2014. http://d-nb.info/1048015424/34.
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Ali, Mohsin [Verfasser]. "Bioanalytical method performance verification concept for cardiovascular research in pediatrics: From development to application in clinical trials / Mohsin Ali." Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2020. http://d-nb.info/1205544135/34.
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Chan, Yannie Ka Yan. "Evaporation-induced 3-dimensional diblock copolymer micelles micropattern : applications as templated polymeric microwells for cell culture scaffold, bioanalytic arrays and micro-silver networks /." View abstract or full-text, 2004. http://library.ust.hk/cgi/db/thesis.pl?BIEN%202004%20CHAN.
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Thesis (M.Phil.)--Hong Kong University of Science and Technology, 2004.Includes bibliographical references (leaves 122-133). Also available in electronic version. Access restricted to campus users.
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Zhou, Mowei. "Incorporation of Surface Induced Dissociation into a Commercial Ion Mobility - Tandem Mass Spectrometer and Application of Mass Spectrometry Methods for Structural Analysis of Non-covalent Protein Complexes." The Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1373977026.
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Vaněčková, Tereza. "Luminiscenční nanočástice pro bioanalytické aplikace =:Luminescent nanoparticles for bioanalytical applications /." Doctoral thesis, 2019. http://www.nusl.cz/ntk/nusl-426259.
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The dissertation thesis entitled Luminescence nanoparticles for bioanalytical applications deals with the use of optical nanomaterials in life sciences. An overview of the commonly used luminescent nanoprobes is provided together with their advantages over commonly used organic dyes or fluorescence proteins. Next, surface modifications and biofunctionalization of nanoparticles with targeting moieties are discussed. Molecularly imprinted polymers are introduced as an alternative surface modification enabling biorecognition. Finally, theoretical part is concluded with recent examples of the luminescent nanoparticles in bioanalytical and imaging applications. The scientific results of the Ph.D. candidate are presented in the form of 2 review articles and 3 research articles in the peer reviewed journals.
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Dong, Rui. "Microfluidic devices for bioanalytical applications /." 2006. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3250234.
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Thesis (Ph. D.)--University of Illinois at Urbana-Champaign, 2006.Source: Dissertation Abstracts International, Volume: 68-02, Section: B, page: 0936. Adviser: Ralph G. Nuzzo. Includes bibliographical references. Available on microfilm from Pro Quest Information and Learning.
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Jian, Ting-Rui, and 簡廷叡. "Bioanalytical Applications of CuO/Cu2O/Ppy Composites." Thesis, 2017. http://ndltd.ncl.edu.tw/handle/qzpnrq.
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碩士國立臺灣大學
化學研究所
105
Linen texture (LT) paper functionalized with CuO/Cu2O/Ppy composite has been prepared and used as an electrode for quantitation of glucose, β-galactosidase, and Escherichia coli (E. coli). The low-cost CuO/Cu2O/Ppy LT paper is prepared through a two-step process; pyrrole adsorbed on an LT paper reacts with Cu2+ in the presence of ascorbic acid and fructose to form Cu2O/Ppy that then reacts with Cu2+ in the presence of cetyltrimethylammonium bromide. The CuO/Cu2O/Ppy LT paper electrode possessing glucose oxidase-like activity catalyzes the oxidation of glucose, allowing quantitation of glucose down to 0.04 mM. Furthermore, functionalized paper size was decreased to 0.12 cm2 to fabricate the CuO/Cu2O/Ppy LT paper microelectrodes. The microelectrode enhances the mass transfer efficiency and reduces the sample volume. By taking the specificity of β-galactosidase to hydrolyze lactose to form glucose, the functionalized paper microelectrode has been employed for the quantitation of β-galactosidase, and detection of E. coli that express β-galactosidase as low as to 0.034 U mL-1 and 120 CFU mL-1, respectively. In addition, this assay is powerful for screening activators/inhibitors of β-galactosidase, and the antibiotic activities toward E. coli. Our results clearly show the merits such as low-costs, high sensitivity, specificity, and superior stability of CuO/Cu2O/Ppy LT paper (micro) electrodes for blood glucose monitor, as well as for quantitation of β-galactosidase activity and E. coli.
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Koehne, Jessica E. "Applications of carbon nanotubes in bioanalytical chemistry research." Diss., 2010. http://proquest.umi.com/pqdweb?did=2065739081&sid=1&Fmt=2&clientId=48051&RQT=309&VName=PQD.
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Berggren, William Travis. "Mass spectrometry as a bioanalytical technique - applications and advancements." 2002. http://www.library.wisc.edu/databases/connect/dissertations.html.
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Lewis, Erica. "Spectroscopic Studies of Cyanine Dyes and Serum Albumins for Bioanalytical Applications." 2015. http://scholarworks.gsu.edu/chemistry_theses/72.
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The use of cyanine dyes in bioanalytical applications has become a widely explored topic of interest in chemistry. Their ability to absorb and fluoresce in the UV-visible and near-infrared region of the electromagnetic spectrum benefits their use as imaging probes and fluorescent labels due to the reduced auto-fluorescence from biological molecules. The behavior of these dyes lies in their structure which consists of two nitrogen containing heterocycles joined by an electron deficient polymethine bridge that allows specific energy transitions to occur. The first portion of this work aims to explore dye functionality for analytical applications regarding the non-covalent labeling of bovine serum albumin. The second portion of the work explores dye interactions with human serum albumin in biological membrane mimetic environments using the ternary system of sodium dioctyl sulfosuccinate (AOT) in water and n-heptane.
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Špirková, Zuzana. "Macroporous indium tin oxide as a potential platform for bioanalytical applications." Master's thesis, 2015. http://www.nusl.cz/ntk/nusl-348656.
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Charles Univesity in Prague, Faculty of Pharmcy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Control Candidate: Zuzana Špirková Supervisor: Prof. PharmDr. Martin Doležal, Ph.D. Consultant: Assist. Prof. Gulnara Safina, Ph.D. Title of thesis: Macroporous indium tin oxide as a potential platform for bioanalytical applications Blood glucose monitoring is an essential tool in diabetes mellitus diagnosis. Therefore new strategies have been developed to improve the performance of glucose sensing devices. In this thesis the suitability of macroporous indium tin oxide (ITO) electrodes for construction of third-generation glucose biosensors was investigated. As a biosensing part in sensor cellobiose dehydrogenase from Corynascus thermophilus (CtCDH) immobilized onto ITO platform was used. Several immobilization strategies based on physical adsorption, electrostatic bindings of the enzyme to the surface functionalized with polythyleneimine (PEI) and cross-linking with glutaraldehyde (GA) were studied in order to achieve reasonable sensitivity and stability of the biosensor. The morphology/topography and elemental composition of the enzyme modified surface were examined by scanning electron microscopy (SEM), energy-dispersive X- ray spectroscopy (EDX), X-ray photoelectron spectroscopy...
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Bird, Susan Mary. "Elemental speciation for bioanalytical applications: HPLC-ICP-MS and CE-ICP-MS." 1998. https://scholarworks.umass.edu/dissertations/AAI9909148.
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Advances in knowledge of toxicology, clinical chemistry, and nutrition have made the mere determination of an element in a sample insufficient. Both the amount of an element and the forms in which it is present need to be determined. Inductively coupled plasma mass spectrometry (ICP-MS) is a powerful element selective detection method for elemental speciation studies, and was employed as a detection method for high performance liquid chromatography (HPLC) and capillary electrophoresis (CE) separations. An ICP-MS ion pair HPLC method for the separation of selenoamino acid standards was developed. The method was used to identify and quantify selenoamino acids present in water extracts of selenium enriched allium and yeast samples which have been shown to have cancer preventive effects in animal or human studies. Commercially available selenium nutritional supplements were also evaluated. Selenium speciation was shown to vary with level of selenium enrichment and among samples. The method was shown to be compatible with standard ICP-MS operating conditions and electrospray ionization mass spectrometry (ESI-MS). The developed method was also shown to be applicable for sulfur and tellurium speciation studies. The oxidation and reduction of selenoamino acids was investigated by HPLC-ICP-MS. Selenoamino acids were shown to be readily oxidized, and the formation of oxidation products were monitored with time. The presence of oxidized or reduced selenoamino acids in extracts of selenium enriched samples was investigated. The reversibility of the oxidation of some selenoamino acids was demonstrated, and identification of the oxidation products was attempted by ESI-MS. A preliminary evaluation of a CE-ICP-MS interface for metalloprotein speciation was carried out using model proteins and human serum, and its effects on the separation characterized. The adsorption of lead to CE capillaries was investigated. The use of commercially coated and dynamically coated capillaries resulted in reduced lead adsorption.
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Pennington, Christopher Lane. "Bioanalytical applications of electrospray ionization mass spectrometry for proteome and biomarker analysis." 2008. http://proquest.umi.com/pqdweb?did=1542114881&sid=4&Fmt=2&clientId=39334&RQT=309&VName=PQD.
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Thesis (Ph.D.)--State University of New York at Buffalo, 2008.Title from PDF title page (viewed on Nov. 26, 2008) Available through UMI ProQuest Digital Dissertations. Thesis adviser: Wood, Troy D. Includes bibliographical references.
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Chen, Tse-Hsien, and 陳澤賢. "Macrocyclic Polyamine Bonded Phase and Gold Nanoparticles Self-assembled Column for Bioanalytical Applications." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/65889027822530593308.
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博士國立臺灣大學
化學研究所
95
Two macrocyclic polyamines, [32]ane-N8 and [28]ane-N6O2 were prepared and employed as a selective modifier for the capillary electrochromatographic separation. Under the same coating process, it has been observed that [28]ane-N6O2 coated column showed higher electroosmotic flow (e.g. μeo = -9.67 × 10-4 cm2V-1s-1) than that of [32]ane-N8 (μeo = -6.19 × 10-4 cm2V-1s-1) at pH 4. It is noteworthy to mention that μeo values increase if pyridine is added into the solution of coating process. As for example, μeo increases from -1.31 × 10-4 to -4.43 × 10-4 cm2V-1s-1 at pH 6, indicating that a greater amount of the bonded ligands were found with the addition of pyridine. With [32]ane-N8 coated column, the optimum conditions for the separation of aminothiols were at -20 kV using an acetate buffer (30 mM, pH 4.0), hydrodynamic injection and detection wavelength at 214 nm. A basic mixture (pH 7.76) of sulfur-containing biomolecules was baseline separated in less than 15 min. In the similar experimental conditions, the [28]ane-N6O2 coated capillary column showed only five peaks and no characteristic splitting for reduced and oxidized form of glutathione. With pH gradient mode, the baseline separation of the acidic mixture (pH 2.73) was demonstrated using an acetate buffer (30 mM) with inlet buffer pH 4 and outlet buffer pH 3 (pH 4-3). But ethanol was needed in the mobile phase for the separation of acidic mixture (pH 1.89). A mixture of angiotensin (I、II and ST)、β-casomorphin (bovine and human)、oxytocin acetate、tocinoic acid、vasopressin and FMRF amide could be separated using phosphate buffer (pH 7, 30 mM). Column efficiency was found with the average theoretical plate numbers of 70000 m-1 and relative standard deviation (RSD) of < 1 % (n = 6) The prepared column can be used with injections more than 600 with the EOF increase only 5.91 %. [Met5]-enkephalin and [Leu5]-enkephalin could also be separated with the bonded phase in 30 mM acetate using pH gradient method (pH 4-3). The [32]ane-N8 coated column was also applied to separate of mono- and disaccharides. The reductive aminated product was carried out with p-aminobenzoic acid. A synergic effect was observed for the separation of borate complexes with the bonded phase. A complete separation of all compounds as well as the excess derivatizing agent by using borate buffer (pH 9.0) in a mode of concentration gradient (60 mM inlet side and 70 mM outlet side) could be achieved. The RSD of the migration time measured for each sample was less than 4 % in six continuous runs, suggesting that the bonded phase along with the gradient formed inside the column was quite stable. The results indicated that with the mixing modes of anion coordination, anion exchange, and shape discrimination, the interaction adequately accomplishes the separation of carbohydrates which are epimers or have different glycosidic linkage, although the electrophoretic migration is also involved in the separation mechanism. The gold nanoparticles(Au NPs) was immobilized in fused-silica capillary column wall by (3-mercaptopropyl)trimethoxysilane for the separation of sulfur-containing biomolecules. We used high concentration buffer (80 ~ 120 mM) to avoid bubble formation. Baseline separation of basic mixture was obtained at 20 kV, using an acetate buffer (80 mM, pH 4). The results suggested that the interaction between analytes and the bonded groups on the wall is ligand exchange. The Au NPs solutions were added into both 32- and 28-membered macrocyclic polyamines for realizing the status of NPs in the presence of these ligands. Spectral changes and TEM pictures demonstrate that the macrocyclic polyamines, which possess positive charges, trigger aggregation of negatively charged citrate-capped nanoparticles. Then the Au NPs self-assembled in a [32]ane-N8 coated column wall. The good separation efficiency was procured not only sulfur-containing biomolecules but also proteins include six ovalbumin peaks are observed.
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Botelho, Julianne Cook. "Bioanalytical applications of mass spectrometry to identify proteins and glycans from complex biological samples." 2007. http://purl.galileo.usg.edu/uga%5Fetd/botelho%5Fjulianne%5Fm%5F200712%5Fphd.
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Thesis (Ph. D.)--University of Georgia, 2007.Directed by Ron Orlando. Includes articles submitted to Journal of proteome research and International journal of mass spectrometry. Includes bibliographical references.
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Johnson, Jeremi David. "Bioanalytical applications of liquid chromatography mass spectrometry protein identification, enzyme function, carbohydrate analysis, post-translational modifications, and targeted proteomics /." 2003. http://purl.galileo.usg.edu/uga%5Fetd/johnson%5Fjeremi%5Fd%5F200308%5Fphd.
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Thesis (Ph. D.)--University of Georgia, 2003.Directed by Ron Orlando. Includes articles submitted to Journal of allergy and clinical immunology, Journal of the American Society for Mass Spectrometry, and European journal of mass spectrometry. Includes bibliographical references.
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Warren, Maria Rosario Esteban. "Bioanalytical applications of mass spectrometry development and evaluation of modified probe surfaces for enhanced MALDI-TOF-MS and examination of the substrate specificity and glycosylation of A. niger pectin methylesterase by ESI-MS /." 2001. http://purl.galileo.usg.edu/uga%5Fetd/warren%5Fmaria%5Fr%5F200108%5Fphd.
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Thesis (Ph. D.)--University of Georgia, 2001.Directed by Ronald Carl Orlando. Includes articles published in Analytical chemistry, and Biochemical journal, and articles submitted to Rapid communications in mass spectrometry, and Biochemical journal. Includes bibliographical references.