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Plasmonic techniques for viral membrane characterizationFeizpour, Amin 08 November 2017 (has links)
The lipid bilayer membrane of enveloped viruses, such as human immunodeficiency virus type 1 (HIV-1), plays an important role in key steps of the infection, including cell binding and uptake. Phosphatidylserine (PS) and monosialotetrahexosylganglioside (GM1) are examples of two host-derived lipids in the membrane of enveloped virus particles that are known to contribute to virus attachment, uptake, and ultimately dissemination. A quantitative characterization of their contribution to the functionality of the virus requires information about their relative concentrations in the viral membrane. In this dissertation, a gold nanoparticle (NP) binding assay for probing relative PS and GM1 lipid concentrations in the outer leaflet of different virus-like particles (VLPs) using small sample sizes is introduced. The assay evaluates both scattering intensity and resonance wavelength and determines relative NP densities through plasmon coupling as a measure for the target lipid concentrations in the NP-labeled VLP membrane. The performed studies reveal significant differences in the membrane of HIV-1 and Ebola VLPs that assemble at different intracellular sites and pave the way to an optical quantification of lipid concentration in virus particles at physiological titers. In addition, this technique was used in another application to improve the understanding of the relationship between the membrane PS lipid and the infectivity of HIV-2 and murine leukemia virus (MLV).
The composition of the membrane, in particular the cholesterol (chol) content, determines its fluidity. As differences in the membrane composition of individual virus particles can lead to different intracellular fates, biophysical tools capable of probing the membrane fluidity on the single-virus level are required. In this dissertation, we demonstrate that fluctuations in the polarization of light scattered off gold or silver nanoparticle (NP)-labeled virus-like-particles (VLPs) encode information about the membrane fluidity of individual VLPs. We developed a plasmonic polarization fluctuation tracking microscopy (PFTM) which facilitated, for the first time, the investigation of the effect of chol content on the membrane fluidity and its dependence on temperature on the single-VLP level. Chol extraction studies with different methyl-β-cyclodextrin (MβCD) concentrations yielded a gradual decrease in polarization fluctuations as function of time. The PFTM revealed chol content and fluidity heterogeneities of an HIV-1 VLP population.
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In vitro pseudomonas aeruginosa biofilms : improved confocal imaging and co-treatment with dispersion agents and antibioticsRoss, Stacy Sommerfeld 01 May 2013 (has links)
Pseudomonas aeruginosa bacterial biofilms are the leading cause of mortality among cystic fibrosis (CF) patients. Biofilms contain bacteria attached to a surface and encased in a protective matrix. Since bacteria within a biofilm are less susceptible to antibiotics, a new approach is to use dispersion compounds that cause the biofilms to release free-swimming bacteria. Our approach has focused on combining nutrient dispersion compounds with antibiotics to increase eradication of bacteria within biofilms. This approach takes advantage of the enhanced susceptibility of free-swimming bacteria to antibiotics, compared to bacteria within biofilms. Ultimately, this research will guide the development of an aerosol therapy containing both antibiotic and dispersion compounds to treat bacterial biofilm infections.
To study the effect of antibiotic and dispersion compound treatments on biofilm eradication, a high-throughput screening assay was used to assess the effect on young Pseudomonas aeruginosa biofilms. In addition, a Lab-Tek chambered coverglass system imaged via confocal microscopy was used to assess the effect on mature Pseudomonas aeruginosa biofilms. Seven antibiotics (amikacin disulfate, tobramycin sulfate, colistin sulfate, colistin methanesulfonate (CMS), polymyxinB sulfate, erythromycin, and ciprofloxacin hydrochloride) were tested alone or in combination with four nutrient dispersion compounds (sodium citrate, succinic acid, xylitol, and glutamic acid) to assess the level of eradication of bacteria within biofilms. For young biofilms, 15 of 24 combinations significantly eliminated more live bacteria within the biofilms (measured in colony forming units per milliliter) compared to antibiotics alone. In the more mature biofilm system, only 3 out of 26 combinations resulted in a higher percentage of live biofilm bacteria being eliminated compared to antibiotics alone, showing the importance of biofilm age in the effectiveness of these potential combination therapies.
To aid in confocal microscopic analysis of biofilms, an automated quantification program called STAINIFICATION was developed. This new program can be used to simultaneously investigate connected-biofilm bacteria, unconnected bacteria (dispersed bacteria), the biofilm protective matrix, and a growth surface upon which bacteria are grown in confocal images. The program contains novel algorithms for the assessment of bacterial viability and for the quantification of bacteria grown on uneven surfaces, such as tissue. The utility of the viability assessments were demonstrated with confocal images of Pseudomonas aeruginosa biofilms. The utility of the uneven surface algorithms were demonstrated with confocal images of Staphylococcus aureus biofilms grown on cultured human airway epithelial cells and Neisseria gonorrhoeae biofilms grown on transformed cervical epithelial cells.
Finally, a proof-of-concept study demonstrated that dry powder aerosols containing both antibiotic and nutrient dispersion compounds could be developed with properties optimized for efficient deposition in the lungs. A design of experiments study showed that solution concentration was the most significant parameter affecting aerosol yield, particle size, and in vitro deposition profiles.
Collectively this work demonstrated that bacterial dispersion from biofilms can enhance antibiotic susceptibility and can be better quantified using the new STAINIFICATION software. Formulation of dispersion compounds and antibiotics into a dry powder aerosol could enable more effective treatment of biofilm infections in the lungs.
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Contributions à la discretisation des contraintes de mesurabilité pour les problèmes d'optimisation stochastiqueBarty, Kengy 25 June 2004 (has links) (PDF)
Nous nous sommes penchés sur différents aspects des problèmes<br />d'optimisation stochastique qui, à notre connaissance, ont été peu<br />étudiés. Ainsi, nous nous sommes intéressés au problème de l'effet dual,<br />puis à la discrétisation des contraintes de mesurabilité, à la<br />résolution numérique de problèmes avec contraintes en information statique et enfin,<br />nous avons étudié les conditions d'optimalité d'un problème<br />d'optimisation stochastique, le but recherché étant de mieux<br />comprendre comment intervient la contrainte de mesurabilité dans la<br />caractérisation de la (ou des) solution(s) optimale(s). Notre approche<br />numérique du problème est originale de deux points de vue :<br />Elle utilise les topologies sur l'espace des sigma-algèbres<br /> pour mesurer la perte d'information due à la discrétisation de la<br /> contrainte de mesurabilité. L'étude de cet espace nous a permis entre<br /> autres d'apporter de nouveaux résultats qui constituent des éléments<br /> essentiels dans notre étude~;<br />Nous montrons que l'erreur de discrétisation provient de la<br /> contribution de deux termes d'erreur : une erreur issue de la <br /> discrétisation de la contrainte de mesurabilité et une autre erreur<br /> issue de l'approximation de l'espérance.<br /><br />Nous donnons dans ce mémoire des résultats asymptotiques de<br />convergence d'une suite de problèmes discrets vers le problème<br />d'origine. Nous avons également, sur des problèmes particuliers, des<br />résultats de type Lipschitz sur la fonction valeur. Par ailleurs,<br />l'étude des conditions d'optimalité nous a permis d'obtenir deux<br />possibilités différentes d'approche d'un problème de commande optimale<br />stochastique.
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Compression d'images complexes avec pertes: application à l'imagerie RadarValade, Cédric 12 1900 (has links) (PDF)
Installé à bord d'aéronefs ou de satellites, le Radar à Synthèse d'Ouverture (RSO) permet d'imager des zones à haute résolution. Les volumes de données générés sont très importants et les capacités limitées des canaux de transmission et/ou de stockage forment un sous-ensemble critique des systèmes d'observation. De ce fait, la solution classiquement retenue est de recourir à la compression des données. Le système radar cohérent génère des données de forte dynamique où les pixels issus de la formation de l'image sont des données complexes au sens mathématique du terme, c'est-à-dire constitués d'une valeur d'amplitude et de phase. La faible corrélation spatiale des pixels rend les techniques de compression répandues dans le domaine optique beaucoup moins efficaces. L'objectif de cette étude consiste à proposer un système de compression avec pertes adapté aux images RSO SLC (Single Look Complex) et prenant en compte le caractère complexe des pixels. Nous suggérons différents schémas de compression dépendants de la répresentation du signal dans le plan complexe (amplitude/phase ou partie réelle/imaginaire). Nous nous focalisons essentiellement sur des techniques de type multirésolution comme la transformée en ondelettes et paquets d'ondelettes discrètes. Un ensemble de modèles statistiques spécifiques à la caractérisation des images RSO permettra d'adapter diverses méthodes de quantification polaire à la problématique de la compression d'images complexes. Enfin, la dernière partie de cette étude sera consacrée à la présentation des résultats à traver l'utilisation d'un panel de critères objectifs avec notamment une évaluation de la qualité des images décompressées.
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The influence of tense in adverbial quantificationEndriss, Cornelia, Hinterwimmer, Stefan January 2004 (has links)
We argue that there is a crucial difference between determiner and adverbial
quantification. Following Herburger [2000] and von Fintel [1994],
we assume that determiner quantifiers quantify over individuals and adverbial
quantifiers over eventualities. While it is usually assumed that
the semantics of sentences with determiner quantifiers and those with
adverbial quantifiers basically come out the same, we will show by way
of new data that quantification over events is more restricted than quantification
over individuals. This is because eventualities in contrast to
individuals have to be located in time which is done using contextual information
according to a pragmatic resolution strategy. If the contextual
information and the tense information given in the respective sentence
contradict each other, the sentence is uninterpretable. We conclude that
this is the reason why in these cases adverbial quantification, i.e. quantification
over eventualities, is impossible whereas quantification over
individuals is fine.
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The interpretation of Universally Quantified DPs and singular definites in adverbially quantified sentencesHinterwimmer, Stefan January 2007 (has links)
This paper deals with the conditions under which singular definites, on the one hand, and universally quantified DPs, on the other hand, receive interpretations according to which the sets denoted by the NP-complements of the respective determiner vary with the situations quantified over by a Q-adverb. I show that in both cases such interpretations depend on the availability of situation predicates that are compatible with the presuppositions associated with the respective determiner, as co-variation in both cases comes about via the binding
of a covert situation variable that is contained within the NP-complement of the respective determiner. Secondly, I offer an account for the observation that the availability of a co-varying interpretation is more constrained in the case of universally quantified DPs than in the case of singular definites, as far as word order is concerned. This is shown to follow from the fact that co-varying
definites in contrast to universally quantified DPs are inherently focus-marked.
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Real Time PCR Protocol Development for Rapid and Low Cost Quantification of Baculovirus and for Monitoring Progression of InfectionGeorge, Steve January 2010 (has links)
The work presented in this thesis aims to further the understanding and implementation of the Baculovirus Expression Vector System (BEVS) for varied uses such as protein production and viral vector production. To this end, three projects have been presented, two of which deal with methods to quantify baculovirus titres and the last deals with tracking baculovirus transcripts in infected insect cells.
The first project examined assumption-free analysis as a method for data analysis of Real Time PCR data in order to enable direct comparison of baculovirus titres between samples, without the need for a traditional standard curve. It concluded that assumption-free analysis was well suited for this purpose and fold differences of baculovirus titres of different samples obtained using this method corresponded to real differences in sample titres.
The second project aimed to develop a cheap and reliable method for sample preparation for Real Time PCR which would remove the need for the use of commercially available extraction kits. Samples were subjected to various combinations of Triton X-100 at different concentrations and different numbers of freeze/thaw cycles in order to determine the combination which would provide the best baculovirus genome exposure. One of these combinations was found to be at least as good as commercially available kits in reliably extracting baculovirus DNA and providing baculovirus titres that are at least as accurate.
The third project was a preliminary study examining the effects of multiplicity of infection on the levels of baculovirus Gp-64 transcript in insect cell culture. The study concludes that at high multiplicities of infection, there seems to be no increase in baculovirus transcripts when the multiplicity of infection is further increased. This study served to allow for familiarization with tracking transcript levels, and the principles and techniques demonstrated here will form the basis for an exhaustive future study on the same subject.
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Investigating Functional Breast Image Quality and Quantification with a Dedicated SPECT-CT SystemPerez, Kristy Lynn January 2011 (has links)
<p>This work investigates phantom and subject positioning as well as collecting data with a variety of angular sampling and acquisition trajectories. The overall goal of this work has been to utilize the dedicated, breast SPECT-CT system to acquire the best possible images. A large portion of this work has been to apply corrections to the system for quantitative imaging. The system has been shown to provide high quality images with minimal out-of-field signal contribution. Additionally, the quantification procedure has been shown to be within 10% of the known activity concentration present at the time of imaging for both VAOR and PROJSINE trajectories.</p> / Dissertation
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Flow Imaging Using MRI: Quantification and AnalysisJiraraksopakun, Yuttapong 2009 May 1900 (has links)
A complex and challenging problem in flow study is to obtain quantitative flow information in opaque systems, for example, blood flow in biological systems and flow channels in chemical reactors. In this regard, MRI is superior to the conventional optical flow imaging or ultrasonic Doppler imaging. However, for high speed flows, complex flow behaviors and turbulences make it difficult to image and analyze the flows.
In MR flow imaging, MR tagging technique has demonstrated its ability to simultaneously visualize motion in a sequence of images. Moreover, a quantification method, namely HARmonic Phase (HARP) analysis, can extract a dense velocity field from tagged MR image sequence with minimal manual intervention. In this work, we developed and validated two new MRI methods for quantification of very rapid flows. First, HARP was integrated with a fast MRI imaging method called SEA (Single Echo Acquisition) to image and analyze high velocity flows. Second, an improved HARP method was developed to deal with tag fading and data noise in the raw MRI data. Specifically, a regularization method that incorporates the law of flow dynamics in the HARP analysis was developed. Finally, the methods were validated using results from the computational fluid dynamics (CFD) and the conventional optimal flow imaging based on particle image velocimetry (PIV). The results demonstrated the improvement from the quantification using solely the conventional HARP method.
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Investigation of quantitative absolute concentrations of in vivo proton magnetic resonance spectroscopyLiang, Deng-hao 11 July 2006 (has links)
Magnetic resonance spectroscopy has been widely used in medical applications, rendering precise evaluation and diagnosis in clinics. As the development of various tools for automatic spectra analysis, providing objective quantification of metabolites, absolute concentrations has been playing an important role in clinical studies and applications as well.
In this study, we investigate the reliability and accuracy of absolute concentration quantified by LCModel. Ten healthy subjects were included. We compared the resultant concentrations calculated by internal water scaling and phantom calibration, both of which are provided by LCModel. Partial volume effect was also taken into account to improve the accuracy of absolute concentrations. Automatic segmentation was applied to volume of interest in order to separate gray matter and white matter, which will facilitate the further partial volume correction and thus better accuracy of absolute quantification.
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