Дисертації: "High content of protein"

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Leuchowius, Karl-Johan. "High Content Analysis of Proteins and Protein Interactions by Proximity Ligation." Doctoral thesis, Uppsala universitet, Molekylära verktyg, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-119530.

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Fundamental to all biological processes is the interplay between biomolecules such as proteins and nucleic acids. Studies of interactions should therefore be more informative than mere detection of expressed proteins. Preferably, such studies should be performed in material that is as biologically and clinically relevant as possible, i.e. in primary cells and tissues. In addition, to be able to take into account the heterogeneity of such samples, the analyses should be performed in situ to retain information on the sub-cellular localization where the interactions occur, enabling determination of the activity status of individual cells and allowing discrimination between e.g. tumor cells and surrounding stroma. This requires assays with an utmost level of sensitivity and selectivity. Taking these issues into consideration, the in situ proximity-ligation assay (in situ PLA) was developed, providing localized detection of proteins, protein-protein interactions and post-translational modifications in fixed cells and tissues. The high sensitivity and selectivity afforded by the assay's requirement for dual target recognition in combination with powerful signal amplification enables visualization of single protein molecules in intact single cells and tissue sections. To further increase the usefulness and application of in situ PLA, the assay was adapted to high content analysis techniques such as flow cytometry and high content screening. The use of in situ PLA in flow cytometry offers the possibility for high-throughput analysis of cells in solution with the unique characteristics offered by the assay. For high content screening, it was demonstrated that in situ PLA can enable cell-based drug screening of compounds affecting post-translational modifications and protein-protein interactions in primary cells, offering superior abilities over current assays. The methods presented in this thesis provide powerful new tools to study proteins in genetically unmodified cells and tissues, and should offer exciting new possibilities for molecular biology, diagnostics and drug discovery.

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2

Tavares, Joana Formigal. "Identification of novel regulators of protein synthesis fidelity using high content genetic screens." Doctoral thesis, Universidade de Aveiro, 2018. http://hdl.handle.net/10773/22825.

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Doutoramento em Biomedicina
Protein synthesis is central to life and is being intensively studied at various levels. The exception is mRNA translational fidelity whose study has been hampered by technical difficulties in detecting amino acid misincorporations in proteins. Few genes have so far been associated to the control of protein synthesis fidelity and it is unclear how many genes control this biological process. We investigated the role of RNA modification by RNA modifying enzymes (RNAmods) in protein synthesis efficiency and accuracy. Our hypothesis was that RNAmods that modify tRNA nucleosides (tRNAmods) have a significant impact on protein synthesis through modulation of codonanticodon interactions. To address this issue, we focused our work on tRNAmods involved in the modification of tRNA anticodons. The biology of these enzymes is still poorly understood, but they are involved in RNA processing, stability and function and their deregulation is associated with cancer, neurodegenerative, metabolic and other diseases. We have set up a yeast genetic screen and used mass-spectrometry methods to determine the role of tRNAmods on proteome homeostasis. Our work identified a subgroup of yeast tRNAmods that play essential roles in protein synthesis fidelity and folding. The genes that encode insoluble proteins isolated from yeast cells lacking U34 modification were enriched in codon sites that are decoded by the hypomodified tRNAs. These aggregated proteins also participate in specific biological processes, suggesting that tRNAmods are linked to specific physiological pathways. Interestingly, we detected amino acid misincorporations at the codon sites decoded by the anticodons of the hypomodified tRNAs, demonstrating that tRNA U34 modifications control translational error rate.
A síntese proteica é central para a vida e tem sido extensivamente estudada a vários níveis. Contudo, o estudo da fidelidade da tradução do mRNA tem progredido lentamente devido a dificuldades técnicas na deteção de incorporações incorretas de aminoácidos nas proteínas. Poucos genes têm sido associados com o controlo da fidelidade da síntese proteica e não é evidente quais os genes que controlam este processo biológico. Nesta tese investigámos o papel da modificação dos nucleósidos do RNA na eficiência e precisão da síntese proteica. A nossa hipótese é que as enzimas que modificam nucleósidos do tRNA (tRNAmods) têm um impacto significativo na síntese proteica através da modulação das interações codão-anticodão. A biologia das tRNAmods e das modificações do tRNA são ainda pouco conhecidas, mas estão envolvidas na estabilidade e função do RNA e mutações nos seus genes causam doenças neurodegenerativas, metabólicas, cancro, entre outras. Neste projeto realizámos um rastreio genético em levedura com o objetivo de identificar tRNAmods que asseguram a homeostase do proteoma (proteostase) e usámos espectrometria de massa para clarificar o papel das tRNAmods na fidelidade da síntese proteica. Os resultados do estudo genético mostram que um sub-grupo de tRNAmods envolvidas na modificação de nucleósidos do anticodão do tRNA são essenciais para manter a estabilidade do proteoma. Outras tRNAmods estudadas não produziram impactos visíveis na proteostase. Os genes de proteínas agregadas que isolámos a partir de células de levedura com tRNAs hipomodificados são enriquecidos em codões descodificados por estes tRNAs. Os nossos dados mostram também que tais proteínas participam em processos biológicos específicos e têm níveis de aminoácidos errados mais elevados que as células wild-type. Estes dados mostram que certas modificações do tRNA são essenciais para a fisiologia celular, estabilidade do proteoma e fidelidade da síntese proteica.

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3

Jüttemann, Thomas. "Adding 3D-structural context to protein-protein interaction data from high-throughput experiments." Thesis, University of Edinburgh, 2011. http://hdl.handle.net/1842/5666.

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In the past decade, automatisation has led to an immense increase of data in biology. Next generation sequencing techniques will produce a vast amount of sequences across all species in the coming years. In many cases, identifying the function and biological role of a protein from its sequence can be a complicated and time-intensive task. The identification of a protein's interaction partners is a tremendous help for understanding the biological context in which it is involved. In order to fully characterise a protein-protein interaction (PPIs), it is necessary to know the three-dimensional structure of the interacting partners. Despite optimisation efforts from projects such as the Protein Structure Initivative, determining the structure of a protein through crystallography remains a time- and cost-intensive procedure. The primary aim of the research described in this dissertation was to produce a World Wide Web resource that facilitates visual exploration and validation (or questioning) of data derived from functional genomics experiments, by building upon existing structural information about direct physical PPIs. Secondary aims were (i) to demonstrate the utility of the new resource, and (ii) its application in biological research. We created a database that emphasises specifically the intersection between the PPIs-results emerging from the structural biology and functional genomics communities. The BISC database holds BInary SubComplexes and Modellable Interactions in current functional genomics databases (BICS-MI). It is publicly available at hyyp://bisc.cse.ucsc.edu. BISC is divided in three sections that deliver three types of information of interest to users seeking to investigate or browse PPIs. The template section (BISCHom and BISCHet) is devoted to those PPIs that are characterised in structural detail, i.e. binary SCs extracted from experimentally determined three-dimensional structures. BISCHom and BISCHet contain the homodimeric (13,583 records) and heterodimeric (5612 records) portions of these, respectively. Besides interactive, embedded Jmol displays emphasising the interface, standard information and links are provided, e.g. sequence information and SPOP classification for both partners, interface size and energy scores (PISA). An automated launch of the MolSurfer program enables the user to investigate electrostatic and hydrophobic correlation between the partners, at the inter-molecular interface. The modellable interactions section (BISC0MI) identifies potentially modellable interactions in three major functional genomics interaction databases (BioGRID), IntAct, HPRD). To create BISC-MI all PPIs that are amenable to automated homology modelling based on conservative similarity cut-offs and whose partner protein sequences have recrods in the UniProt database, have been extracted. The modellable interaction services (BISC-MI Services) section offers, upon user request, modelled SC-structures for any PPIs in BISC-MI. This is enabled through an untomated template-based (homology) modelling protocol using the popular MODELLER program. First, a multiple sequence alignment (MSA) is generated using MUSCLE, between the target and homologous proteins collected from UniProt (only reviewed proteins from organisms whose genome has been completely sequenced are included to find putative orthologs). Then a sequence-to-profile alignment is generated to integrate the template structure in the MSA. All models are produced upon user request to ensure that the most recent sequence data for the MSAs are used. Models generated through this protocol are expected to be more accurate generally than models offered by other automated resources that rely on pairwise alignments, e.g. ModBase. Two small studies were carried out to demonstrate the usability and utility of BISC in biological research. (1) Interaction data in functional genomics databases often suffers from insufficient experimental and reporting standards. For example, multiple protein complexes are typically recorded as an inferred set of binary interactions. Using the 20S core particle of the yeast proteasome as an example, we demonstrate how the BISC Web resource can be used as a starting point for further investigation of such inferred interactions. (2) Malaria, a mosquito-borne disease, affects 3500-500 million people worldwide. Still very little is known about the malarial parasites' genes and their protein functions. For Plasmodium falciparum, the most lethal among the malaria parasites, only one experimentally derived medium scale PPIs set is available. The validity of this set has been doubted in the the malarial biologist community. We modelled and investigated eleven binary interactions from this set using the BISC modelling pipeline. Alongside we compared the BISC models of the individual partners to those obtained from ModBase.

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4

Kattah, Michael George. "High-content protein arrays for characterizing immune responses and pathophysiology at the molecular level /." May be available electronically:, 2008. http://proquest.umi.com/login?COPT=REJTPTU1MTUmSU5UPTAmVkVSPTI=&clientId=12498.

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5

Schmiele, Marcio 1979. "Interações físicas e químicas entre isolado protéico de soja e glúten vital durante a extrusão termoplástica a alta e baixa umidade para a obtenção de análogo de carne = Physical and chemical interactions between isolated soy protein and vital gluten during thermoplastic extrusion at high and low moisture content to obtain meat analogue." [s.n.], 2014. http://repositorio.unicamp.br/jspui/handle/REPOSIP/255892.

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Orientador: Yoon Kil Chang
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos
Made available in DSpace on 2018-08-24T06:53:45Z (GMT). No. of bitstreams: 1 Schmiele_Marcio_D.pdf: 9722936 bytes, checksum: 95d9146270f349c5f3e7ad761ac0d266 (MD5) Previous issue date: 2014
Resumo: Os análogos de carne obtidos por extrusão termoplástica de proteínas vegetais são caracterizados pelo seu elevado teor proteico e estrutura semelhante às fibras da carne, envolvendo diversos tipos de ligações e/ou interações químicas entre as proteínas. O objetivo deste trabalho foi avaliar as características tecnológicas e físico-químicas de análogos de carne, à base de isolado proteico de soja, obtidos por processo de extrusão termoplástica a alta umidade (AU) e baixa umidade (BU). Para cada condição de umidade foi utilizado um Delineamento Composto Central Rotacional de três variáveis independentes (glúten vital, umidade de condicionamento e temperatura de extrusão). As variáveis dependentes avaliadas foram a textura instrumental, cor instrumental, capacidade de absorção de água, índice de solubilidade em água, capacidade de absorção de óleo, índice de dispersibilidade de proteína, energia mecânica específica e o tipo de interações proteicas. Estas interações foram avaliadas através de sete tipos de solventes específicos: (i) tampão fosfato para as proteínas no estado nativo; (ii) dodecil sulfato de sódio para as interações hidrofóbicas e iônicas; (iii) Triton 100X para as interações hidrofóbicas; (iv) ureia para as interações hidrofóbicas e pontes de hidrogênio; (v) ß-mercaptoetanol para as ligações dissulfeto; e (vi) ß-mercaptoetanol e ureia e (vii) dodecil sulfato de sódio e ureia, para avaliar o efeito sinérgico entre os sistemas. O ponto otimizado (caracterizado principalmente por promover maiores valores de L* e de capacidade de absorção de água, menores valores de índice de solubilidade em água, de capacidade de absorção de óleo, de desnaturação proteica e valores intermediários de textura instrumental e de energia mecânica específica) foi processado juntamente com uma amostra controle para ambos os processos com o intuito de validar os modelos matemáticos e avaliar as possíveis alterações na morfologia dos análogos de carne, na massa molecular das proteínas, na composição de aminoácidos totais e na desnaturação proteica. As melhores condições de processamento foram obtidos para os análogos de carne contendo de 12 e 5 % de glúten vital, 58 e 18 % de umidade de condicionamento e 135 e 100 °C para a temperatura de extrusão, para o processo AU e BU, respectivamente. As principais interações proteína-proteína encontradas nos análogos de carne foram as ligações dissulfeto e ligações de hidrogênio para o processo AU e as ligações dissulfeto e interações iônicas para o processo BU. A adição de glúten vital promoveu uma aparência mais lisa e melhor orientação na estrutura das fibras. Verificou-se que ocorreu aumento nas proteínas de baixa massa molecular e diminuição nas proteínas de alta massa molecular. No perfil de aminoácidos totais houve maior variação negativa para os aminoácidos essenciais (triptofano e treonina), semi essenciais (cisteína) e não essenciais (serina), indicando que houve redução no valor nutricional. As estruturas secundárias (a-hélice, ß-folha, ß-volta e a estrutura desordenada) mostraram alteração na sua conformação devido à desnaturação proteica e formação de novos agregados
Abstract: Meat analogue obtained by termoplastic extrusion of vegetable proteins are characterized by its high protein levels and structure similar to meat fibers, which comprises many types of chemical bonds and/or interactions between proteins. The aim of this work was to evaluate the technological and physico-chemical characteristics of meat analogue based on isolated soy protein obtained by thermoplastic extrusion process at high moisture (HM) and low moisture (LM) content. For each moisture condition was used a Central Rotational Composite Design with three independent variables (vital gluten, moisture content and extrusion temperature). The dependent variables evaluated were instrumental texture, instrumental color, water absorption capacity, water solubility index, oil absorption capacity, protein dispersibility index, specific mechanical energy, and the type of protein interactions. These interactions were evaluated using seven specific solvents types: (i) phosphate buffer for proteins in native state; (ii) sodium dodecil sulphate for hydrophobic and ionic interactions; (iii) Triton 100X for hydrophobic interactions; (iv) urea for hydrophobic interactions and hydrogen bonds; (v) ß-mercaptoethanol for dissulfide bonds; and (vi) ß-mercaptoethanol and urea and (vii) sodium dodecil sulphate and urea, for the synergistic effect between the systems. The optimized point (characterized mainly by promoting higher values for L* and water absorption capacity, lower values for water solubility index, oil absoption capacity and protein denaturation and intermediate values for instrumental texture and specific mechanical energy) was processed, together with a control sample for each processes, in order to validate the mathematical models and to evaluate possibles changes in the meat analogues morphology, in the protein molecular weight, in the total amino acid composition, and in the protein denaturation. The best processing conditions were obtained for the meat analogue containing 12 and 5 % of vital gluten, 58 and 18 % of moisture content and 135 and 100 °C of extrusion temperature, for the HM and LM processes, respectively. The main protein-protein interactions found in meat analogues were the dissulfide bonds and hydrogen bonds for the LM process and the dissulfide bonds and ionic interactions for the HM process. The addition of vital gluten promoted a smoother appearance and better orientation in the fiber structure. It was found that occured an increase in the protein with low molecular weight and a reduction in the protein with high molecular weight. There were a greater negative variation for the essential (tryptophan and threonine), semi-essential (cysteine) and nonessential (serine) amino acids in the total amino acid profile, indicating a reduction of the nutritional value. The secondary structure (a-helix, ß-sheet, ß-turn and disordered structure) showed alteration in its conformation due to the protein denaturation and formation of new aggregates
Doutorado
Tecnologia de Alimentos
Doutor em Tecnologia de Alimentos

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Tulukcuoglu, Güneri Ezgi. "Development of microfluidic device for high content analysis of circulating tumor cells." Thesis, Paris 6, 2016. http://www.theses.fr/2016PA066583/document.

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Le cancer est l'une des principales causes de décès dans le monde. D'après la société américaine contre le cancer; en 2015, un quart des décès aux Etats-Unis est du au cancer du poumon avant même les maladies cardiaques. Cette situation nous incite et bien d'autres scientifiques dans le monde à développer des moyens plus efficaces de traitement, le diagnostic et le dépistage de la maladie. Parce que près de 90% des décès par cancer sont dus à des métastases, de nombreuses études se sont concentrées sur le mécanisme de métastases et sur son impact clinique. Les cellules tumorales circulantes (CTC) sont les cellules s’échappent de tumeurs primaires ou métastatiques pour rejoindre le flux sanguin périphérique, ces cellules sont un élément de transition dans le processus métastatique et portent ainsi des informations cruciales sur ce mécanisme encore mal compris. Les CTCs ont déjà montré leur potentiel comme biomarqueur de pronostic de la progression de la maladie et de l'indicateur de l'efficacité du traitement en fonction l’augmentation ou de la diminution de leur nombre. Leur caractérisation moléculaire peut également donner des informations vis à vis de cibles thérapeutiques possibles et des mécanismes de progression de la maladie ou de la résistance aux médicaments. Leur comptage au cours du traitement combiné avec leur caractérisation moléculaire devrait améliorer la prise en charge des patients dans le cadre de la médecine personnalisée. Cependant CTCs sont extrêmement rares, 1 à 10 cellules / ml de sang parmi les 106 globules blancs et 109 globules rouges, leur capture à partir du sang reste donc un challenge analytique. Dans les dernières décennies, Une grande variété de techniques d'enrichissement et de capture a été mise au point et l'approche microfluidique est l'une des méthodes efficaces, flexibles et à haut débit. Au sein de notre équipe, un dispositif microfluidique (système Ephesia) puissant pour la capture et l'analyse des cellules tumorales circulantes a déjà été mis au point précédemment. Le principe de capture est basé sur l'auto-assemblage de billes magnétiques greffées par des anticorps, grâce aux quelles les cellules sont enrichies via l’interaction Ab- l'antigène de surface EpCAM que l'on trouve communément dans les cellules cancéreuses d'origine épithéliale. Ce système a déjà été validé avec des lignées cellulaires et des échantillons de patients. Cependant, le système n'a pas permis l'isolement / détection des sous-populations de CTCs ou d'effectuer une caractérisation moléculaire très poussée. Par conséquent, mon projet de thèse vise à améliorer encore les capacités du système sur les deux principaux aspects: le ciblage sous-populations de CTC et à l'étude des interactions des protéines à la surface des CTCs dans le Système Ephesia
Metastasis is the advanced stage of cancer progression and is the cause of 90% of deaths in cancer disease. During metastatic cascade, it is suggested that the successful metastatic initiation depends on the survival of circulating tumor cells (CTCs). CTCs are the cells that shed from the primary or secondary tumor sites into the blood circulation. it is now widely recognized as potential biomarker for companion diagnostics in which high number of CTCs in blood can indicate association with poor survival or high risk of disease progression. Besides, following the number of CTCs during the course of treatment can help to adapt the selected therapy and predict the treatment efficacy. On the other hand molecular characterization can provide patient stratification and identifying the therapeutic targets. However they are extremely rare in the bloodstream, estimated between 1-10 CTC among 6×106 leukocytes, 2×108 platelets and 4×109 erythrocytes per one mL of blood which makes their isolation very challenging. A very attractive way of isolation of CTCs is to integrate microfluidics. Microfluidics offers great advantages such as low volume of reagent consumption and short analysis times with automation as well as isolation and detection analysis can be integrated resulting in highly efficient biomedical devices for diagnostics. As parallel to state of the art, a powerful microfluidic device for circulating tumor cells capture and analysis had already been developed previously in our laboratory. The principle of capture is based on self-assembly of antibody-coated (EpCAM) magnetic beads in which the cells are enriched by EpCAM surface antigen which is found commonly in epithelial origin cancer cells. This system was already validated with cell lines and patients samples. However, the system did not allow isolation/detection of subpopulations of CTCs or performing high content molecular characterization. Therefore, my PhD project aimed at further improving the capabilities of the system on the main two aspects: targeting subpopulations of CTC and studying of protein interactions of CTCs in Ephesia System

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Skutnik, Benjamin C. "The effects of high intensity interval training on resting mean arterial pressure and C-reactive protein content in prehypertensive subjects." Thesis, Kansas State University, 2013. http://hdl.handle.net/2097/15774.

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Master of Science
Department of Kinesiology
Craig A. Harms
Subjects with prehypertension are at risk for developing hypertension (HTN). Hypertension is associated with low-grade systemic inflammation (LGSI). Aerobic exercise training (ET) is a proven means to reduce both blood pressure and LGSI in healthy and diseased subjects. Recently, high intensity interval training (HIIT) has been show to elicit similar cardiovascular and metabolic adaptations as ET in healthy and at-risk populations in a more time efficient manner. Therefore, we hypothesized that HIIT would elicit greater reductions in blood pressure and LGSI than ET. Twelve pre-hypertensive subjects (systolic blood pressure 127.0 ± 8.5 mmHg; diastolic blood pressure 86.2 ± 4.1 mmHg) were randomly assigned to an ET group (n=5) and a HIIT group (n=7). All subjects performed an incremental test to exhaustion (VO2max) on a cycle ergometer prior to, after 4 weeks, and after 8 weeks of training. Resting heart rate and blood pressure were measured prior to and three times a week during training. LGSI was measured via high-sensitivity C-reactive protein (hs-CRP) prior to, after 4 weeks and after 8 weeks of training. ET subjects performed an eight week exercise training program at 40% VO2 reserve determined from the VO2max test, while HIIT subjects performed exercise at 60% peak power determined from the VO2max test. ET group trained four days/week while HIIT trained three days/week. ET exercised for 30 minutes continuously at a constant workload and cadence of 60 rpm while HIIT performed a protocol on a 1:1 work-to-rest ratio at a constant workload and cadence of 100 rpm. Both groups showed similar (p<0.05) decreases in mean arterial (ET = -7.3%, HIIT = -4.5%), systolic (ET = -6.6%, HIIT = -8.8%), and diastolic (ET= -9.7, HIIT= -8.2%) blood pressure. HIIT decreased in LGSI (-33.7%) while ET did not change LGSI (p>0.05). VO2max increased ~25% with both HIIT and ET with no differences (p>0.05) between groups. These data suggest both HIIT and ET similarly decreased resting blood pressure and increased VO2max while HIIT was effective in decreasing LGSI in subjects who were pre-hypertensive.

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Kapor-Drezgic, Jovana. "High glucose alters mesangial cell protein kinase C activity and isoform cellular content and localization, role of the polyol pathway." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape10/PQDD_0003/MQ40808.pdf.

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Zhu, Seng. "Study of the mechanism of Tunneling nanotubes formation and their role in aggregate proteins transfer between cells." Thesis, Université Paris-Saclay (ComUE), 2017. http://www.theses.fr/2017SACLS377.

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Les Tunneling nanotubes (TNT) sont des protrusions cellulaires à base d'actine qui médient la communication cellulaire en transférant des cargos cellulaires. Les différents types de communication intercellulaires sont de plus en plus considérés comme des cibles potentielles pour le traitement de différentes maladies, telles que les maladies infectieuses liées aux virus et bactéries, les cancers ou les maladies neurodégénératives. Des études récentes ont mis en évidence un mécanisme de propagation d'agrégats protéiques ressemblant à la propagation du prion dans diverses maladies neurodégénératives non infectieuses telles que la maladie d'Alzheimer (AD), la démence frontotemporelle (FTD), la maladie de Parkinson (PD) et la maladie de Huntington. Ces maladies se caractérisent par l'accumulation de protéines mal repliées dans le cerveau des patients. Ainsi, on peut envisager de nouvelles stratégies thérapeutiques pour bloquer la propagation des protéines anormales dans tout le cerveau. Il a été démontré que les TNT pourraient jouer un rôle essentiel dans la propagation des agrégats de prions au sein du système nerveux central (SNC) et périphérique. Par conséquent, l'étude du mécanisme de la formation de TNT pourrait fournir de nouvelles idées sur le mécanisme de propagation de la maladie et de nouvelles cibles thérapeutiques. L'objectif de ma thèse était d'étudier le rôle du transfert des agrégats de protéines par les TNT entre les cellules et d'étudier le mécanisme de formation des TNT. Dans notre laboratoire, nous avons déjà montré que les TNT permettent le transfert de prions entre les cellules. Dans la première partie de mon doctorat, j'ai confirmé que les transferts d'agrégats de prions entre les cellules de CAD neuronales se faisaient par les TNT à l'intérieur de vésicules endocytiques (Zhu et al., 2015). De plus, en collaboration avec un collègue, nous avons fourni des preuves que les agrégats de prions pourraient être transférés entre des astrocytes primaires et des neurones et que ce transfert était médié par un contact cellulaire (Victoria et al., 2016). J'ai également collaboré à une autre étude où nous avons montré que les agrégats d'α-synucléine (caractéristiques de la maladie de Parkinson) peuvent être transférés entre les cellules à l'intérieur des lysosomes, et que ce transfert intercellulaire est médié par les TNT (Abounit et al., 2016). Dans mon deuxième projet, afin d'étudier le mécanisme de la formation de TNT, j'ai effectué un crible à haut débit pour les Rab GTPase. J'ai trouvé que Rab8 et Rab11 peuvent favoriser la formation des TNT, et que les cascades Rab8-VAMP3, Rab11-ERM et Rab8-Rab11 sont impliquées dans la formation des TNT. Mes données suggèrent que la polymérisation de l'actine et le trafic de membranes sont impliqués dans la formation des TNT. Ces résultats permettent d'éclairer le mécanisme de la formation des TNT et de fournir des preuves moléculaires que les Rab GTPases régulent ce processus
Tunneling nanotubes are actin-based cell protrusions that mediate cell-to-cell communication by transferring cellular cargos. The different types of intercellular communication are increasing by being considered as potential targets for the treatment of various diseases, such as infectious diseases linked to viruses and bacteria, cancers or neurodegenerative diseases. Recent studies have highlighted a prion-like mechanism of propagation of protein misfolding in a variety of common, non-infectious, neurodegenerative diseases such as Alzheimer’s disease (AD), Frontotemporal dementia (FTD), Parkinson’s disease (PD), and Polyglutamine (PolyQ) diseases, which are characterized by the accumulation of misfolded proteins in the brain of patients. Thus, new therapeutic strategies to block propagation of protein misfolding throughout the brain can be envisaged. It has been shown that TNTs might play a critical role in spreading of prion aggregates within the CNS and from the periphery. Therefore, the study of mechanism of TNT formation could provide new insights on the mechanism of disease propagation and novel therapeutic targets. The aim of my thesis was to study the role of TNT-mediate protein aggregates transfer between cells and to investigate the mechanism of TNT formation. In our lab, we already reported TNT mediate prion transfer between cells. In the first part of my PhD, I further confirmed that prion aggregates transfer between neuronal CAD cells through TNT inside endocytic vesicles (Zhu et al., 2015). Furthermore in collaboration with a colleague, we provided evidences that prion aggregates could transfer between primary astrocytes and neurons and the transfer was mediated by cell-to-cell contact (Victoria et al., 2016). I also collaborated to another study where we showed that α-synuclein aggregates (Parkinson’s disease) can transfer between cells inside lysosomes, and the intercellular transfer is mediated by TNTs (Abounit et al., 2016).In my second project, in order to investigate the mechanism of TNT formation, I performed a High-content screening of Rab GTPase. I found that Rab8 and Rab11 can promote TNT formation, that Rab8-VAMP3, Rab11-ERM and Rab8-Rab11 cascades are involved in TNT formation. My data suggests that both actin polymerization and membrane trafficking are involved in TNT formation. These results help to shed light on the mechanism of TNT formation, and provide molecular evidences that Rab GTPases regulate this process

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10

Kelly, Douglas James. "An automated fluorescence lifetime imaging multiwell plate reader : application to high content imaging of protein interactions and label free readouts of cellular metabolism." Thesis, Imperial College London, 2014. http://hdl.handle.net/10044/1/29131.

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This thesis reports on work performed in the development and application of an automated plate reading microscope implementing wide field time gated fluorescence lifetime imaging technology. High content analysis (HCA) imaging assays enabled by automated microscopy platforms allow hundreds of conditions to be tested in a single experiment. Though fluorescence lifetime imaging (FLIM) is established in life sciences applications as a method whereby quantitative information may be extracted from time-resolved fluorescence signals, FLIM has not been widely adopted in an HCA context. The FLIM plate reader developed throughout this PhD has been designed to allow HCA-FLIM experiments to be performed and has been demonstrated to be capable of recording multispectral, FLIM and bright field data from 600 fields of view in less than four hours. FLIM is commonly used as a means of reading out Förster resonance energy transfer (FRET) between fluorescent fusion proteins in cells. Using the FLIM plate reader to investigate large populations of cells per experimental condition without significant user input has allowed statistically significant results to be obtained in FRET experiments that present relatively small changes in mean fluorescent lifetime. This capability has been applied to investigations of FOXM1 SUMOylation in response to anthracycline treatment, and to studies of the spatiotemporal activation profiles of small GTPases. Furthermore, the FLIM plate reader allows FLIM-FRET to be applied to protein-protein interaction screening. The application of the instrument to screening RASSF proteins for interaction with MST1 is discussed. The FLIM plate reader was also configured to utilise ultraviolet excitation radiation and optimised for the measurement of autofluorescence lifetime for label-free assays of biological samples. Experiments investigating the autofluorescence lifetime of live cells under the influence of metabolic modulators are presented alongside the design considerations necessary when using UV excitation for HCA-FLIM.

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11

Huang, Jing. "DIFFERENTIAL ACTIVITY AND CONTENT OF HIGH-AFFINITY GLUTAMATE TRANSPORTERS, CONTENT OF THEIR REGULATORY PROTEINS, AND CAPACITY FOR GLUTAMINE AND GLUTATHIONE SYNTHESIS IN TISSUES OF FINISHED VERSUS GROWING STEERS." UKnowledge, 2017. https://uknowledge.uky.edu/animalsci_etds/81.

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Improvement of feeding regimens for production animals has been hindered by a lack of fundamental knowledge about how the capacity to regulate nutrient absorption across cell membranes affects the function of nutrient metabolizing enzymes. The objective is to determine if the activities and protein content of system X-AG glutamate transporter, its regulatory protein (GTRAP3-18 and ARL6IP1), glutamine synthetase (GS) and glutathione (GSH) content, changes in liver (Experiment 1), longissimus dorsi (LM) and subcutaneous adipose tissue (SF) (Experiment 2) as beef steers transitioned from predominantly-lean (growing) to -lipid (finished) tissue accretion phases. In liver (Experiment 1), system X-AG activity in canalicular membranes was abolished as steers developed from growing to finished stages but did not change in basolateral membranes. EAAC1 protein content in liver homogenates decreased in finished vs. growing steers, whereas GTRAP3-18 and ARL6IP1 content increased and GLT-1 content did not change. Concomitantly, hepatic GS activity decreased in finished steers whereas GS protein content did not differ. Hepatic GSH content did not differ in finished vs. growing steers. These results demonstrate a negative functional relationship between GTRAP3-18 and system X-AG activity with glutamine synthesis capacity in livers of fattened cattle. In addition to liver, skeletal muscle and adipose tissues play important roles in maintaining whole-body glutamate and nitrogen homeostasis. In Experiment 2, Western blot analysis of LM homogenates showed decreased EAAC1 and GS content, whereas GTRAP3-18 and ARL6IP1 did not differ in finished vs. growing steers. GSH content in LM was increased in finished vs. growing steers in concomitance with increased mRNA expression of GSH-synthesizing enzymes. In SF, GTRAP3-18 and ARL6IP1 content was increased, whereas EAAC1 and GS content did not differ. Concomitantly, GSH content in SF was decreased in finished vs. growing steers in parallel with decreased mRNA expression of GSH-metabolizing enzymes. These results demonstrate that the negative regulatory relationship between GTRAP3-18 and ARL6IP1 with EAAC1 and GS expression, which exists in liver, does not exist in LM and SF of fattened cattle; and antioxidant capacity in LM and SF changes and differs as steer compositional gain shifts from lean to lipid phenotype. To further explore the upstream regulatory machinery of EAAC1, transcriptome analysis (Experiment 3) was conducted to gain a greater understanding of hepatic metabolic shifts associated with the change in whole-body compositional gain of growing vs. finished beef steers. The expression of upstream regulators of EAAC1 was decreased in a manner consistent with the decreased EAAC1 activity in Experiment 1. Bioinformatic analysis found that, for amino acid metabolism, finished steers had increased capacities for ammonia, arginine, and urea production, and shunting of amino acid carbons into pyruvate. For carbohydrate metabolism, capacity for glycolysis was inhibited, whereas glycogen synthesis was stimulated in finished steers. For lipid metabolism, finished steers showed decreased capacity for fatty acid activation and desaturation, but increased capacity for fatty acid b-oxidation and lipid storage. In addition, redox capacity and inflammatory responses were decreased in finished steers. Collectively, these data describe novel regulatory relationships of system X-AG in liver and peripheral tissues, and the metabolic mechanisms that control nutrient use efficiency, as beef steers develop from lean to lipid phenotypes.

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12

Pogna, Edgar Allan. "Phosphorylation and distribution of High-Mobility Group protein HMGN1 in the context of Immediate-Early (IE) gene induction." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:5b39d964-fd81-4534-bfba-76a32bb60f4e.

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Eukaryotic genomes are highly organized and packaged into chromatin, a complex structure formed of proteins and DNA, in which the basic repeating unit is the nucleosome. Chromatin can be arranged in condensed or relaxed structures influencing accessibility of proteins that regulate transcription, replication, recombination and repair. One class of transiently chromatin-associated proteins is the High-Mobility Group (HMG) protein family. HMG proteins are subdivided into three subgroups: HMGA, HMGB and HMGN. HMGN1, the subject of this study, is a prominent member of the HMGN (High-Mobility Group Nucleosome-binding) protein family, the only HMG proteins that specifically binds to the nucleosomes. HMGNs are maintained in dynamic balance between nucleosome-associated and nucleosome-free pools. Regulation of chromatin involves several enzymatic activities that modify specific residues on chromatin proteins, which may influence these interactions. While associated with nucleosomes, HMGNs can interfere with some modifications of histone tails. Modification of HMGN1 on specific residues and post-translational modification (PTM) of histones are concomitantly regulated by the complex signalling networks associated with the induction of immediate early (IE) genes. Induction of IE genes is associated with phosphorylation of HMGN1 which has been suggested to increase the rate of dissociation of HMGN1 from the nucleosome, thus allowing access and modification of histone tails. My research has been focused on characterizing HMGN1 isoforms present in different cellular compartments and at different time-points during IE gene induction with various stimuli, including epidermal growth factor (EGF), anisomycin (An) and 12-O-tetradecanoylphorbol-13-acetate (TPA). Furthermore, I investigated the localization of HMGN1 within the nucleus and at specific IE gene loci, especially at sites where post-translationally modified histones are localised. In my analysis only the phosphorylation at serine 6 of HMGN1 shows a correlation with gene induction. Analysis of DNA sequences from chromatin immunoprecipitation (ChIP) has shown that HMGN1 is present at equal levels in active and inactive genes. It appears that HMGN1 localization on DNA is not dictated by a particular preference for any gene elements such as promoters, exons, introns or gene termination sequences.

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13

Jarvius, Malin. "Visualization of Protein Activity Status in situ Using Proximity Ligation Assays." Doctoral thesis, Uppsala universitet, Molekylära verktyg, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-131934.

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In 2001 the human proteome organization (HUPO) was created with the ambition to identify and characterize all proteins encoded in the human genome according to several criteria; their expression levels in different tissues and under different conditions; the sub-cellular localization; post-translational modifications; interactions, and if possible also the relationship between their structure and function.When the knowledge of different proteins and their potential interactions increases, so does the need for methods able to unravel the nature of molecular processes in cells and organized tissues, and ultimately for clinical use in samples obtained from patients. The in situ proximity ligation assay (in situ PLA) was developed to provide localized detection of proteins, post-translational modifications and protein-protein interactions in fixed cells and tissues. Dual recognition of the target or interacting targets is a prerequisite for the creation of a circular reporter DNA molecule, which subsequently is locally amplified for visualization of individual protein molecules in single cells. These features offer the high sensitivity and selectivity required for detection of even rare target molecules. Herein in situ PLA was first established and then employed as a tool for detection of both interactions and post-translational modifications in cultured cells and tissue samples. In situ PLA was also adapted to high content screening (HCS) for therapeutic effects, where it was applied for cell-based drug screening of inhibitors influencing post-translational modifications. This was performed using primary cells, paving the way for evaluation of drug effects on cells from patient as a diagnostic tool in personalized medicine. In conclusion, this thesis describes the development and applications of in situ PLA as a tool to study proteins, post-translational modifications and protein-protein interactions in genetically unmodified cells and tissues, and for clinical interactomics.

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14

Heidlebaugh, Nancy Marie. "Analysis of nitrogen reallocation from senescing barley leaves characterization of the influence of a high-grain protein content locus on chromosome six, and molecular cloning and heterologous expression of a serine carboxypeptidase /." Thesis, Montana State University, 2008. http://etd.lib.montana.edu/etd/2008/heidlebaugh/HeidlebaughN0508.pdf.

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15

Råvik, Mattias. "Influence of Escherichia coli feedstock properties on the performance of primary protein purification." Licentiate thesis, KTH, Bioprocessteknik, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-3941.

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Abstract The aim of the present study was to increase the understanding of how the cell surface properties affect the performance of unit operations used in primary protein purification. In particular, the purpose was to develop, set up and apply methods for studies of cell surface properties and cell interactions. A method for microbial cell surface fingerprinting using surface plasmon resonance (SPR) is suggested. Four different Escherichia coli strains were used as model cells. Cell surface fingerprints were generated by registration of the interaction between the cells and four different surfaces, with different physical and chemical properties, when a cell suspension was flown over the surface. Significant differences in fingerprint pattern between some of the strains were observed. The physical properties of the cell surfaces were determined using microelectrophoresis, contact angle measurements and aqueous two-phase partitioning and were compared with the SPR fingerprints. The generated cell surface fingerprints and the physical property data were evaluated with multivariate data analysis that showed that the cells were separated into individual groups in a similar way using principal component analysis plots (PCA). Studies of the behaviour of the model cells on stirred cell filtration and in an interaction test with different expanded bed adsorption (EBA) adsorbents were performed. It could be concluded that especially one of the strains behaved differently. Differences in the properties of the model cells were indicated by microelectrophoresis and aqueous two-phase partitioning which to some extent correlated with observed differences in behaviour during filtration and in an interaction test with EBA adsorbents. The impact of high-pressure homogenisation of E. coli cell extract was examined, with a lab scale and a pilot scale technique. The DNA-fragmentation, visualised with agarose gel electrophoresis, and the resulting change in viscosity was analysed. A short homogenisation time resulted in increased viscosity of the process solution that correlated with increased concentration of released non-fragmented DNA. With longer homogenisation time the viscosity decreased with increasing degree of DNA-fragmentation. The results show that strain dependant cell surface properties of E. coli may have an impact on several primary steps in downstream processing.
QC 20101129

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16

Råvik, Mattias. "Infuence of Escherichia coli feedstock properties on the performance of primary protein purification." Licentiate thesis, KTH, School of Biotechnology (BIO), 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-3941.

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Abstract

The aim of the present study was to increase the understanding of how the cell surface properties affect the performance of unit operations used in primary protein purification. In particular, the purpose was to develop, set up and apply methods for studies of cell surface properties and cell interactions.

A method for microbial cell surface fingerprinting using surface plasmon resonance (SPR) is suggested. Four different Escherichia coli strains were used as model cells. Cell surface fingerprints were generated by registration of the interaction between the cells and four different surfaces, with different physical and chemical properties, when a cell suspension was flown over the surface. Significant differences in fingerprint pattern between some of the strains were observed. The physical properties of the cell surfaces were determined using microelectrophoresis, contact angle measurements and aqueous two-phase partitioning and were compared with the SPR fingerprints. The generated cell surface fingerprints and the physical property data were evaluated with multivariate data analysis that showed that the cells were separated into individual groups in a similar way using principal component analysis plots (PCA).

Studies of the behaviour of the model cells on stirred cell filtration and in an interaction test with different expanded bed adsorption (EBA) adsorbents were performed. It could be concluded that especially one of the strains behaved differently. Differences in the properties of the model cells were indicated by microelectrophoresis and aqueous two-phase partitioning which to some extent correlated with observed differences in behaviour during filtration and in an interaction test with EBA adsorbents.

The impact of high-pressure homogenisation of E. coli cell extract was examined, with a lab scale and a pilot scale technique. The DNA-fragmentation, visualised with agarose gel electrophoresis, and the resulting change in viscosity was analysed. A short homogenisation time resulted in increased viscosity of the process solution that correlated with increased concentration of released non-fragmented DNA. With longer homogenisation time the viscosity decreased with increasing degree of DNA-fragmentation.

The results show that strain dependant cell surface properties of E. coli may have an impact on several primary steps in downstream processing.

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17

Benning, B. K. "Novel high water content hydrogels." Thesis, Aston University, 2000. http://publications.aston.ac.uk/9809/.

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The addition of N-isopropyl acrylamide to an acrylamide-based composition that had previously been designed to become a contact lens, produced materials that showed smart effects in that the water content showed dependence on the temperature of the hydrating solution. Such thermo-responsive materials have potential uses in drug delivery, ultrafiltration and cell culture surfaces. Proteoglycans in nature have an important role to play in structural support where a highly hydrophilic structure maintains lubricious surfaces. Certain functional groups that impart this hydrophilicity are present in certain sulphonate monomers, Bis(3-sulphopropyl ester) itaconate, dipotassium salt (SPI), 3-Sulphopropyl ester acrylate, potassium salt (SPA) and Sodium 2-(acrylamido)-2-methyl propane sulphonate (NaAMPS). These monomers were incorporated into a HEMA-based copolymer that had been designed initially as a contact lens and the resulting effects examined. Highly hydrophilic materials resulted that showed reduced protein deposition over the neutral core material. It is postulated that a sulphonate group would have a larger number of hydration shells around it than for example methacrylic acid, leading to more dynamic exchange and so reducing the adsorption of biological solutes. A cationic monomer was added to bring back the net anionic nature of the sulphonate hydrogels and the effects studied. Ionic interactions were found to cause a reduction in the water content of the resulting materials as the mobility of the network decreased, leading to stiffer but less extensible materials. The presence of a net dominant charge, whether negative or positive, appeared to act to reduce protein deposition, but increasing equivalence in the amount of both charges served to present a more 'neutral' surface and deposition subsequently increased. The grafting of hydrophilic hydrogel layers onto silicone elastomer was attempted and the results evaluated using dynamic contact angle measurements. Following plasma oxidation to reduce the surface energy barrier to aqueous grafting chemistry, it was found that the wettability of the modified elastomers could be significantly enhanced by such treatment. The SPA-grafted material in particular hinted at an osmotic drive for rehydration that may be exploited in biomaterials.

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18

Aitken, Karen S. "Genetic analysis of grain protein content in wheat." Thesis, University of East Anglia, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.357308.

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19

Feng, Tian. "High dynamic range visual content compression." Thesis, University of Sheffield, 2017. http://etheses.whiterose.ac.uk/18315/.

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This thesis addresses the research questions of High Dynamic Range (HDR) visual contents compression. The HDR representations are intended to represent the actual physical value of the light rather than exposed value. The current HDR compression schemes are the extension of legacy Low Dynamic Range (LDR) compressions, by using Tone-Mapping Operators (TMO) to reduce the dynamic range of the HDR contents. However, introducing TMO increases the overall computational complexity, and it causes the temporal artifacts. Furthermore, these compression schemes fail to compress non-salient region differently than the salient region, when Human Visual System (HVS) perceives them differently. The main contribution of this thesis is to propose a novel Mapping-free visual saliency-guided HDR content compression scheme. Firstly, the relationship of Discrete Wavelet Transform (DWT) lifting steps and TMO are explored. A novel approach to compress HDR image by Joint Photographic Experts Group (JPEG) 2000 codec while backward compatible to LDR is proposed. This approach exploits the reversibility of tone mapping and scalability of DWT. Secondly, the importance of the TMO in the HDR compression is evaluated in this thesis. A mapping-free post HDR image compression based on JPEG and JPEG2000 standard codecs for current HDR image formats is proposed. This approach exploits the structure of HDR formats. It has an equivalent compression performance and the lowest computational complexity compared to the existing HDR lossy compressions (50% lower than the state-of-the-art). Finally, the shortcomings of the current HDR visual saliency models, and HDR visual saliency-guided compression are explored in this thesis. A spatial saliency model for HDR visual content outperform others by 10% for spatial visual prediction task with 70% lower computational complexity is proposed. Furthermore, the experiment suggested more than 90% temporal saliency is predicted by the proposed spatial model. Moreover, the proposed saliency model can be used to guide the HDR compression by applying different quantization factor according to the intensity of predicted saliency map.

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20

Colin, Béatrice. "Développement d’un système rapporteur générique sensible, au double mode de lecture BRET/HCS, pour l’étude du suivi de gènes rapporteurs, de l’activité de protéase virale et des interactions protéine-protéine." Thesis, Lille 2, 2018. http://www.theses.fr/2018LIL2S050.

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De nombreux systèmes rapporteurs ont été développés pour répondre à différentes applications allant de l’étude des régions promotrices gouvernant l’expression de gènes au suivi de voies de signalisation ou encore à l’interaction de partenaires protéiques.Dans le but initial de développer un test d’activité protéase ayant une sensibilité accrue, nous avons mis au point un système rapporteur permettant l’amplification du signal en développant un système basé sur l’utilisation d’une enzyme relais hyperactive, la protéase TEV (Tobacco Etch Virus) qui vient couper une sonde protéique cible. La détection du signal se fait par deux modes de lecture compatibles avec le haut débit et conduit à un signal de type ON/OFF. Le transfert d’énergie de bioluminescence par résonance (BRET), permet de mesurer un signal à l’état basal, qui diminuera après clivage de la sonde par la protéase TEV. La microscopie de fluorescence à haut contenu ou HCS (High Content Screening) permettra d’observer le changement de localisation de la fluorescence de l’accepteur fluorescent de la sonde utilisée dans le transfert d’énergie, du noyau vers le cytoplasme des cellules, différenciant ainsi les cellules positives des négatives.Ainsi, la grande sensibilité de notre système nous a amené à valider son utilisation de façon générique, en optimisant dans un premier temps la sonde BRET/HCS à la base du test par une série de délétions dans le but d’obtenir le meilleur transfert d’énergie possible, en délétant des acides aminés à l’extrémité C-terminale de l’accepteur et N-terminale du donneur d’énergie. Le contraste entre les deux états a également été amélioré afin de faciliter la lecture en HCS, en testant différentes séquences de localisation nucléaire. Grâce à cette nouvelle sonde, plusieurs preuves de concept dans des domaines variés ont pu être démontrées telles que i) le suivi de l’infection virale avec pour modèle l’infection par le virus de l’Hépatite C (HCV) ii) le suivi de l’expression de gènes rapporteurs avec comme modèle le récepteur TGR5 couplé aux protéines G, impliqué dans le diabète ainsi que iii) la détection d’interactions protéine/protéine avec pour modèle de développement l’interaction entre les protéines FRB et FKBP12 induite par la rapamycine. Les résultats réunis lors de cette thèse ont permis le développement d’un nouveau système rapporteur d’une sensibilité accrue utilisable dans différentes applications biologiques
Several reporter systems have been developed in order to support fundamental scientific projects, from gene promoter regulation study, signaling pathway activation, or protein/protein interaction monitoring.In order to enhance the sensitivity of a protease assay, we developed a new reporter system allowing signal amplification by optimizing a system based on the use of a hyperactive relay enzyme from Tobacco Etch Virus (TEV protease). The signal is detected by two methods compatible with high-throughput and leads to an ON/OFF signal. The Bioluminescence Resonance Energy Transfer (BRET) allows us to measure a signal at basal state, which will decrease upon cleavage of the probe by the protease. High Content Screening (HCS) Microscopy also allows the differentiation between positive and negative cells by a simple shift in the fluorescence acceptor location of the probe used in energy transfer, from the nucleus to the cytoplasm of the cells.The high sensitivity of our approach now leads us to validate its use in a more generic way. This is why the project aimed at the optimization of the BRET probe at the origin of the test, by performing a serie of amino acid deletions at the N-terminus of the energy donor and the C-terminus of the energy acceptor to find the best probe in order to obtain the best energy transfer. The HCS contrast between the two states was aslo increased by testing different nuclear localization sequences. With this new probe, we tried to adapt the system to several fields of application like i) a viral infection test using the HCV protease as a proof of concept but also ii) the expression of a reporter gene with the TGR5, a G-coupled protein receptor as model and iii) the detection of protein-protein interactions. To demonstrate this concept, we used the well know interaction between FRB and FKBP12 proteins, induced by rapamycine. Results from this project will lead to the development of a new reporter system, auto-amplified and usable in a generic way, which a very high sensitivity is necessary

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21

DiÌaz, Maria Dolores FernaÌndez. "Effects of high pressure on protein protein interactions." Thesis, University of Reading, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.270415.

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22

Bartram, Christopher Gordon. "The endogenous protein content of ruminant proximal duodenal digesta." Thesis, University of Nottingham, 1987. http://eprints.nottingham.ac.uk/13952/.

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Protein arriving at the ruminant proximal duodenum consists of microbial protein, undegraded feed protein and endogenous protein. In this study, endogenous protein is defined as that fraction of the digesta derived from the animal itself (e.g. enzymes, plasma proteins, sloughed cells and mucus), not including any endogenous protein which may have been incorporated into the microorganisms. Recent feeding schemes (e.g. ARC 1980, 1984) require an accurate value of the degradability of feed in the rumen. When degradability is determined in vivo failure to account for a quantity of endogenous protein in the proximal duodenal digesta results in an underestimate of the degradability of a ration. Direct estimates of the endogenous protein content of proximal duodenal digesta are therefore required. This thesis describes the development of a method to do this and its application to sheep and cattle. The approach adopted, based on the concept of isotope dilution, involved a continuous intravenous infusion of L-[4,5-3H]-leucine. This resulted in all body protein becoming labelled. Any label detected in the duodenal digesta must therefore be derived from the animal itself. A comparison of the specific activity of duodenal digesta with that of suitable precursor proteins provided an estimate of the proportion of duodenal digesta of endogenous origin. Interestingly, the bacterial fraction of duodenal digesta was also labelled. This indicated that bacteria were utilising an endogenous source of leucine and circumstantial evidence suggested that this was derived largely from the rumen epithelium. The validity of the L-[4,5-3H]-leucine technique was investigated using three sheep fed an essentially protein free diet. Values of endogenous protein flow derived via the proposed technique (2.6 + 1.00 g N/day) were compared with those calculated by difference (2.1 + 0.92 g N/day). A possible dietary influence on the endogenous protein content of ruminant proximal duodenal digesta was examined. An estimate of 2.6 + 0.58 g N/day was derived for three sheep fed a concentrate diet and 2.1 + 0.22 g N/day for three sheep fed long hay. Thus, contrary to previous suggestions, no significant dietary effect was observed. Possible reasons for this are discussed. The proposed technique was also applied to derive estimates in cattle. A value of 11.3 + 1.73 g N/day was obtained for four steers (170 kg) fed silage supplemented with fishmeal. The continuous intravenous infusion of L-[4,5-3H]-leucine method is the first technique to provide a direct determination of the endogenous protein component of ruminant proximal duodenal digesta which can be applied to any dietary regime and used in sheep and cattle.

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23

Lewin, Erland. "Approaches to Optimizing High Content Confocal Microscopy." Licentiate thesis, KTH, Applied Physics, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-10691.

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This thesis presents two methods for improving high contentconfocal microscopy.

The author's part in the first, "Toward a confocal subcellular atlasof the human proteome" was automating image capture of foursimultaneously tagged structures in cells in 96 well plates. In total,thousands of images of hundreds of proteins in cells. The authorwas also part of deciding which imaging methods should be used tomaximize image information content and quality, given the limitedtime available per well in order to scan large numbers of wells.

The second project, "Improved water permeability measurementsbased on fluorescence normalization" involves increasing the sensitivityof measurements of protein function by normalizing with anestimate of the amount of protein in the cell - the fluorescentsignal of a co-transfected protein. This could lead to achievingsufficient confidence in measurements with fewer experiments(thus increasing the information content in each experiment). Asurprisingly high level of noise in the relationship between thefluorescent signal and the protein function was measured.

Denna avhandling presenterar två projekt för att förbättrametoder för experiment med stora informationsmängderbaserade på konfokalmikroskopi.

Författarens del i det första projektet, "Toward a ConfocalSubcellular Atlas of the Human Proteome" (Mot en konfokal,subcellulär atlas av det mänskliga proteomet) var att automatiserabildinsamlingen av fyra samtidigt inmärkta strukturer i celler iplattor med 96 brunnar. Sammanlagt togs tusentals bilder avhundratals proteiner i celler. Författaren var även del i att fastställavilka bildinsamlingsmetoder som skulle användas för att maximeramängd och kvalitet på bild-informationen givet den begränsade tidper brunn som var tillgänglig för att kunna avbilda många brunnar.

Den andra studien, "Improved water permeability measurementsbased on fluorescence normalization" (Förbättrade vattenpermeabilitetsmätningargenom normalisering av fluorescens) syftade till att ökakänsligheten hos mätningar av proteiners funktion genom attnormalisera mätningarna med signalen från fluorescensen från ettkotransfekterat protein. Det skulle kunna leda till att nå tillräckligtillförlitlighet i mätresultaten med färre experiment (därmed ökainformationsinnehållet i varje experiment). En förvånansvärt högbrusnivå i förhållandet mellan fluorescenssignalen ochproteinfunktionen uppmättes

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24

Wu, Mei. "Polymer microarrays for microbial high-content screening." Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/7664.

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Research on the interactions between microbes and polymeric materials constitutes an important part in antimicrobial identification and provides an insight into microbial response on the polymer surfaces. Herein, a high-content screening method with polymer microarray technology was developed to investigate microbe-polymer interactions, especially in studying adhesion/repellence of microbes (bacteria and parasites). Firstly, the polymer microarray approach was used to successfully identify polymers which either selectively captured or prevented the binding of major food-borne pathogen, Salmonella Typhimurium. A parallel study with a lab strain of Escherichia coli was also carried out, revealing polymers which either displayed a common binding activity or which exhibited species discrimination. Likewise, this polymer microarray technology was applied to more bacterial strains, such as Campylobacter, Clostridium, Streptococcus, Klebsiella and their cocktails to discover families of substrates that displayed strong broad-spectrum bacterial non-binding activity. These synthetic polymers represented a novel class of coating materials which can be used to prevent surface colonisation and subsequent formation of bacterial biofilms. The study of protozoan-polymer interactions was also explored in this thesis. Polymers were identified which either bound or prevented parasites (Crysporidium parvum and Giardia lamblia) binding. Material properties, including wettability, surface roughness and polymer composition were analysed to study correlation of parasite binding and the generation of polymer structure function relationships.

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25

Ekengren, Jens. "Estimating inclusion content in high performance steels." Licentiate thesis, Karlstad : Faculty of Technology and Science, Materials Engineering, Karlstads universitet, 2008. http://urn.kb.se/resolve?urn=urn:nbn:se:kau:diva-3520.

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26

Ohlsson, Simon. "IGSCC in weld with high ferrite content." Thesis, KTH, Fysik, 2019. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-252691.

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27

Jacques, Richard. "Statistical analysis of high content screening data." Thesis, University of Sheffield, 2009. http://etheses.whiterose.ac.uk/2220/.

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High throughput screening experiments are typically used within the pharmaceutical industry for the identification and evaluation of candidate drugs. Using a high throughput screen with automated imaging platform allows a large number of compounds to be tested in a biological assay in order to identify any activity inhibiting or activating a biological process. High throughput fluorescent images contain information that can be used to define fully the effects of a compound on cells. It is for this reason that florescent imaging assays have been termed high content screening (Clemons, 2004). The studies analysed in this thesis involve the use of an automated robotic system to administer compounds to cellular assays and take high content images. These images are then analysed and quantified using imaging algorithms to produce a set of variables. Each high content screen may extend to a million or more individual assays. Supervised classification methods have important applications in high content screening experiments where they are used to predict which compounds have the potential to be developed into new drugs. The use of supervised classification for high content screening data is investigated and a new classification method is proposed for batches of compounds where the rule is updated sequentially using information from the classification of previous batches. This methodology accounts for the possibility that the training data are not a representative sample of the test data and that the underlying group distributions may change as new compounds are analysed. Unsupervised classification methods are used in the analysis of high content screening experiments to evaluate potential new drugs. The study in this thesis considers clustering compounds based on their toxicological effect on the liver. Drug induced liver injury is the most common cause for non approval and withdrawal by the Food and Drug Administration (Ainscow, 2007a) and therefore this is an important stage in drug development.

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28

Gros, Robert. "Regulation of G-protein-coupled receptor function, a role for increased G-protein-coupled receptor kinase-2 protein content." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0019/NQ58133.pdf.

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29

Sadahira, Mitie Sônia 1964. "Effect of polysaccharide addition on the foaming properties of egg white protein in aqueous and high sugar contente systems = Efeito da adição de polissacarídeos nas propriedades espumantes de proteínas da clara de ovo em sistemas aquoso e com alto teor de açúcares." [s.n.], 2014. http://repositorio.unicamp.br/jspui/handle/REPOSIP/256405.

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Orientador: Flavia Maria Netto
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos
Made available in DSpace on 2018-08-25T15:30:20Z (GMT). No. of bitstreams: 1 Sadahira_MitieSonia_D.pdf: 5351794 bytes, checksum: 3b4bdc18fc4978c11662cb3791bc6fe6 (MD5) Previous issue date: 2014
Resumo: Nos confeitos aerados (marshmallow e nougat), a espuma é produzida pela aeração de xaropes de açúcares, estabilizada por proteínas tais como proteínas da clara de ovo (PCO). A pectina, polissacarídeo aniônico, pode formar complexos eletrostáticos com proteína em pH abaixo do ponto isoelétrico da proteína. A hidroxipropilmetilcelulose (HPMC) é um polissacarídeo neutro com propriedades emulsificantes. O trabalho visou estudar as propriedades espumantes (capacidade de aeração e estabilidade da espuma) da PCO na presença destes polissacarídeos em solução aquosa e sistema modelo de açúcares. Na primeira etapa, foram avaliados os efeitos das interações PCO/polissacarídeo nas propriedades espumantes em solução aquosa. Os efeitos da concentração de biopolímeros (2,0-4,0% p/p), proporção PCO:pectina (15:1-55:1) e temperatura (70-80 °C) nas propriedades espumantes no pH 3,0 foram avaliados, utilizando delineamento composto central. Na proporção PCO:pectina 15:1, os complexos eram próximos da eletroneutralidade e com tamanho médio de 95,91+ ou - 8,19 µm, conduzindo para maior estabilidade da espuma quanto à desproporção. Na proporção 55:1, os complexos não eram eletricamente neutros e com tamanho médio de 45,92+ ou - 3,47 µm, resultando em espumas com menor drenagem de líquido e coalescência. Foram avaliados os efeitos de concentração de biopolímeros (2,0-5,0% p/p), proporção PCO:HPMC (2:1-18:1) e pH (3,0-6,0) a 75 °C utilizando delineamento composto central rotacional (DCCR) e do comportamento dos biopolímeros na solução aquosa em diferentes pH nas propriedades espumantes. No pH 3,0, os biopolímeros eram compatíveis, conduzindo a melhores propriedades espumantes enquanto nos pH 4,5 e 6,0, os biopolímeros eram incompatíveis, resultando em menor estabilidade com relação a desproporção. Na segunda etapa do trabalho, foram avaliados os efeitos das interações PCO/polissacarídeo em sistema modelo de açúcares com características de marshmallow (densidade<0,50 g/mL; atividade de água<0,75). A composição da solução de açúcares (42,5% sacarose, 42,5% xarope de glicose e 15% de açúcar invertido) foi definida utilizando delineamento experimental de mistura. Os efeitos da concentração de biopolímeros (1,40¿5,60% p/p) e proporção PCO:pectina (7:1¿63:1) nas respostas foram avaliadas utilizando um DCCR, no pH 3,0. As respostas foram viscosidade aparente da mistura açúcares/PCO/pectina antes do batimento e densidade, overrun, parâmetros reológicos da amostra aerada recém-processada e após 24 horas (módulo elástico G¿, módulo viscoso G" e 'delta'). Na proporção PCO:pectina 7:1, a mistura apresentou baixa capacidade de aeração e uma espuma com característica menos sólida e baixa estabilidade. Na proporção 49:1, a mistura apresentou maior capacidade de aeração e comportamento elástico da espuma. Os efeitos da concentração de biopolímeros (1,4-5,6% p/p) e proporção clara de ovo:HPMC (2:1-18:1) nas respostas das misturas açúcar/PCO/HPMC foram avaliados, utilizando um DCCR no pH 3,0 e as mesmas respostas avaliadas no estudo com misturas açúcar/PCO/pectina. Na concentração de biopolímeros 5,0% p/p e proporção PCO:HPMC 14:1 foram realizados experimentos em diferentes pH. No pH 3,0, foram obtidos maior capacidade de aeração e comportamento elástico. No pH 4,5, a espuma apresentou melhor estabilidade comparada a espuma no pH 3,0. No pH 6,0, a espuma apresentou propriedades espumantes ruins e comportamento viscoso. Portanto, o controle das interações proteína/polissacarídeo é um fator chave para o desenvolvimento de produtos aerados com maior estabilidade física
Abstract: In aerated confectionery (marshmallow and nougat), foam is produced by aeration of sugar syrups and stabilized by proteins such as egg white protein (EW). Pectin, an anionic polysaccharide, may form electrostatic complexes with protein at pH values bellow the isoeletric point (pI) of the protein. Hydroxypropylmethylcellulose (HPMC) is a neutral polysaccharide with emulsifying properties. The study aimed at studying the foaming properties (foaming capacity and foam stability) of EW in the presence of these polysaccharides in aqueous solution and high sugar system. Firstly, the effects of EW/polysaccharide interaction on the foaming properties in aqueous solution were evaluated. The effects of biopolymer concentration (2.0-4.0% w/w), EW:pectin ratio (15:1-55:1) and temperature (70-80 °C) were evaluated at pH 3.0, using a central composite design. At EW:pectin ratio 15:1, the complexes were close to electroneutrality and with an average size of 95.91+ or - 8.19 µm, leading to greater stability related to disproportionatin. At ratio 55:1, the complexes were not electrically neutral and with an average size of 45.92+ or - 3.47 µm, resulting in a low drainage of liquid and coalescence. The effects of biopolymer concentration (2.0-5.0% w/w), EW:HPMC ratio (2:1-18:1) and pH (3.0-6.0) at 75 °C were evaluated using central composite rotatable design (CCRD) and the behavior of biopolymer in aqueous solution on the foaming properties at different pH. At pH 3.0, EW and HPMC were compatible leading to better foaming properties whereas at pH 4.5 and 6.0, EW and HPMC were incompatible resulting in lower stability related to disproportionation. In the second part of the study, the effects of EW/polysaccharide interactions on a model system of sugar with characteristics of marshmallow (density<0.50 g/mL; water activity<0.75) were evaluated. For that, a sugar solution composition (42.5% of sucrose, 42.5% of glucose syrup and 15.0% of invert sugar) was defined by a mixture experimental design. The effects of biopolymer concentration (1.40-5.60% w/w) and EW:pectin ratio (7:1-63:1) on the reponses were evaluated using CCRD, at pH 3.0. The responses were apparent viscosity of sugar/EW/pectin mixture before whipping, overrun, foam density and, rheological parameters of fresh foam and foam aged for 24 h (elastic modulus G¿, viscous modulus G" and phase angle 'delta'). At EW:pectin ratio 7:1, the mixture showed low foaming capacity and a foam with less solid character and low stability. At ratio 49:1, the mixture presented greater foaming capacity and elastic behavior of foam. The effects of biopolymer concentration (1.4-5.6% w/w) and EW:HPMC ratio (2:1-18:1) on the responses of sugar/EW/HPMC mixtures were evaluated using CCRD at pH 3.0 and the same responses evaluated in the study of sugar/EW/pectin mixtures. At biopolymer concentration 5.0% w/w and EW:HPMC ratio 14:1, experiments were carried out at different pH. At pH 3.0, the higher foaming capacity and elastic behavior were obtained. At pH 4.5, foam showed better stability than foam at pH 3.0. At pH 6.0, foam presented the poorest foaming properties and viscous behavior. Thus, the control of protein/polysaccharide interactions is a key factor for the aerated products developing with higher stability
Doutorado
Consumo e Qualidade de Alimentos
Doutora em Alimentos e Nutrição

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30

Lei, Stephen. "Economic Feasibility of Assembling Grade-A Milk by Protein Content." DigitalCommons@USU, 1988. https://digitalcommons.usu.edu/etd/4082.

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This thesis consisted of two computerized simulations of assembling milk from dairy farms and distributing it to milk plants, using TRUCKSTOPS, a commercial truck routing computer program. In the first simulation milk was assembled and delivered to the nearest available plant without regard to protein content, with the high-protein milk delivered to manufacturing plants. Doing so increased the fat and protein in milk delivered to manufacturing plants, and increased cheese production 2.6 percent. It also increased assembly costs and lowered fat and protein in milk delivered to fluid milk plants. The value of the extra cheese was less than the extra assembly costs and the value of the butterfat diverted from fluid milk to manufacturing plants, making the operation economically unfeasible.

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31

Brown, Alston Neal. "Effects of Oscillating Crude Protein Content of Dairy Cow Diets." The Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1406190341.

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32

Faupel, Thomas. "Identification of protein-protein-interactions in vitro based on high-density protein arrays." [S.l.] : [s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=972661077.

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33

Multari, Salvatore. "Plant protein isolates with optimised phenolic content to partially replace meat protein in the human diet." Thesis, University of Aberdeen, 2016. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=230929.

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The production, processing and marketing of sustainable and affordable food involve complex phenomena that affect the lives of millions of people worldwide. Due to the rapid growth of the world's population, the provision of food is a significant challenge for the agrifood industry and policy makers, as this is strictly interlinked with climate change and public health interventions. The overall aim of this research was to contribute to delivering nutritious food to feed an increasing unhealthy population. High-protein crops that can be grown sustainably in high latitude countries, including Scotland, could provide a healthy alternative to partially replace our dependency on unsustainable protein-rich foodstuffs. These include meat, the production of which is responsible for a substantial share of food-related environmental pressures. For this reason, green pea, lupin, fava bean, hemp and buckwheat were selected and analysed for their macro- and micro- nutrient content, as well as their phytochemical profile and compared to a red meat- and wheat-based meal in a human intervention trial. The crops studied were high in protein (ranging from 20 to 43% in buckwheat and lupin, respectively) and fibre (up to 25% in hemp) and also found to contain a diverse range of phenolic compounds, considered to participate in the prevention of diet-related disorders. As fava bean contained relatively high amounts of protein (approx. 22% w/w), protein fractions were isolated and further investigated to understand the contribution of the phytochemical components in terms of protein functionality and oxidative stability. Since fava bean protein isolates showed promising food applications, they were used to develop meat patties. The addition of fava bean proteins significantly decreased lipid and protein oxidation of the processed products. The results of this research could encourage a higher consumption of plant-based products, which would be favourable from both a health and environmental perspective.

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34

Ramírez, Orozco Raissel. "High dynamic range content acquisition from multiple exposures." Doctoral thesis, Universitat de Girona, 2016. http://hdl.handle.net/10803/371162.

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The limited dynamic range of digital images can be extended by composing different exposures of the same scene to produce HDR images. This thesis is composed of an overview of the state of the art techniques and three methods to tackle the image alignment and deghosting problems in the HDR imaging domain. The first method detects the areas affected by motion, registers the dynamic objects over a reference image, and combines low-dynamic range values to recover HDR values in the whole image. The second approach builds multiscopic HDR images from LDR multi-exposure images. It is based on a patch match algorithm which was adapted and improved to take advantage of epipolar geometry constraints of stereo images. The last method proposes to replace under/over exposed pixels in the reference image by using valid HDR values from other images in the multi-exposure LDR image sequence.
El limitado rango dinámico de las imágenes digitales puede ampliarse mezclando varias imágenes adquiridas con diferentes valores de exposición. Esta tesis incluye un detallado resumen del estado del arte y tres métodos diferentes para alinear las imágenes y corregir el efecto ’ghosting’ en imágenes HDR. El primer método está centrado en detectar las áreas afectadas por el movimiento y registrar los objetos dinámicos sobre una imagen de referencia de modo que se logre recuperar información a lo largo de toda la imagen. Nuestra segunda propuesta es un método para obtener imágenes HDR multiscópicas a partir de diferentes exposiciones LDR. Está basado en un algoritmo de ’patch match’ que ha sido adaptado para aprovechar las ventajas de las restricciones de la geometría epipolar de imágenes estéreo. Por último proponemos reemplazar los píxeles saturados en la imagen de referencia usando valores correctos de otras imágenes de la secuencia.

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35

Mao, Songqin. "High water content sludge dewatering via freeze-thaw." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq21188.pdf.

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36

Pernagallo, Salvatore. "Biocompatible polymer microarrays for cellular high-content screening." Thesis, University of Edinburgh, 2010. http://hdl.handle.net/1842/7571.

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The global aim of this thesis was to study the use of microarray technology for the screening and identification of biocompatible polymers, to understand physiological phenomena, and the design of biomaterials, implant surfaces and tissue-engineering scaffolds. This work was based upon the polymer microarray platform developed by the Bradley group. Polymer microarrays were successfully applied to find the best polymer supports for: (i) mouse fibroblast cells and used to evaluate cell biocompatibility and cell morphology. Fourteen polyurethanes demonstrated significant cellular adhesion. (ii) Analysis of the adhesion of human erythroleukaemic K562 suspension cells onto biomaterials with particular families of polyurethanes and polyacrylates identified. A DNA microarray study (to access the global gene expression profiles upon cellular binding) demonstrated that interactions between cells and some polyacrylates induced a number of transcriptomic changes. These results suggested that, during these interactions, a chain of cellular changes is triggered, most notably resulting in the downregulation of membrane receptors and ligands. (iii) Identification of polymers with potential applications in the field of stem cell biology. Polymers were identified that showed attachment, promotion and stabilisation of hepatocyte-like cells. A polyurethane support (PU-134) was pinpointed, which significantly improved both hepatocyte-like cell function and “lifespan”. A second project investigated biomaterials that promoted adhesion, growth and function of endothelial progenitor cells. A new polymer matrix was identified which contained the necessary signals to promote endothelial phenotype and function. This has potential application in the creation of blood vessels and the endothelialisation of artificial vessel prostheses and stent coatings for improving angioplasty therapy. (iv) The study of bacterial adhesion, focusing on the adhesion of food-borne pathogenic bacterium Salmonella enterica serovar typhimurium, strain SL1344, and the commensal bacterium Escherichia coli, strain W3110. Several polymers were found to support selective bacterial enrichment, as well as others that minimised bacterial adhesion.

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Conroy, Kristen Monica. "Treating High Salt Content Wastewater with Sand Bioreactors." The Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu149796844505562.

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38

Chaipraparl, Pornpun. "Thai High School Compute Literacy: A Content Analysis." Thesis, University of North Texas, 1989. https://digital.library.unt.edu/ark:/67531/metadc330995/.

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This study examined the extent to which each computer literacy objective domain, each specific mode of instruction, and each type of question were treated in Thai high school computer literacy text materials. Two textbooks and their accompanying teachers' manuals were examined using three analytical schemes as frameworks for the examinations. The Minnesota Educational Computing Consortium (MECC) computer literacy objectives were used to classify the content in the text materials in order to determine the degree of emphasis on each computer literacy objective domain. The Hawaii state Department of Education (HSDE) instructional modes were used to classify the content in the text materials in order to determine the degree of emphasis on each mode of instruction. Bloom's taxonomy of education, cognitive domain, was used to classify the review questions and exercises in the text materials in order to determine the degree of emphasis on each cognitive level. Detailed findings are given as numerals, percentages, and decimal values. Perspectives are offered on the need for textbooks which reflect the values and feelings objectives. Conclusions were that (a) text materials focus most on the programming/algorithms objectives and tend to exclude the values and feelings objectives; (b) text materials use only three modes of instruction, focusing first on the topic mode, second on the tutee mode, and last on the tool mode; (c) text material questions focus more on higher cognitive than on lower cognitive levels.

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39

Pardo, Carlos. "Novel technologies for high-throughput and high-content studies on zebrafish larvae." Thesis, Harvard University, 2013. http://dissertations.umi.com/gsas.harvard:10755.

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The zebrafish larva is an ideal candidate for in vivo high-throughput screening: it is a small vertebrate, it is optically transparent, possesses complex organs, and is easy to culture. In addition, genetic mutants and models of human diseases are widely available. Despite these attractive features there are no tools capable of screening at sufficient throughput and resolution to fully exploit the zebrafish. Here, I present a collection of technologies that enable high-throughput studies on zebrafish larvae at cellular resolution.
Engineering and Applied Sciences

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40

Rohde, Christopher 1979. "Methods and technologies for high-throughput and high-content small animal screening." Thesis, Massachusetts Institute of Technology, 2012. http://hdl.handle.net/1721.1/71486.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 2012.
Cataloged from PDF version of thesis.
Includes bibliographical references (p. 157-170).
High-throughput and high-content screening (HTS and HCS) of whole animals requires their immobilization for high-resolution imaging and manipulation. Here we present methods to enable HTS and HCS of the nematode Caenorhabditis elegans (C. elegans). First we present microfluidic technologies to rapidly isolate, immobilize, image and manipulate individual animals. These technologies include 1. a high-speed microfluidic sorter that can isolate and immobilize C. elegans in a well defined geometry for screening phenotypic features in physiologically active animals, 2. an integrated chip containing individually addressable screening-chamber devices for incubation and exposure of individual animals to biochemical compounds and high-resolution time-lapse imaging of multiple animals and 3. a design for delivery of compound libraries in standard multiwell plates to microfluidic devices and also for rapid dispensing of screened animals into multiwell plates. We then present an improved immobilization method that restrains animals with sufficient stability to perform femtosecond laser microsurgery and multiphoton imaging, without any apparent effects on animal health. We subsequently screen the contents of a small-molecule library for factors affecting neural regeneration following femtosecond laser microsurgery of C. elegans using these technologies. This screen identifies the kinase inhibitor staurosporine as a strong inhibitor of neural regeneration, and does so in a concentration and neuronal cell type-specific manner. Finally, we present a simple device for immobilizing C. elegans inside standard microtiter plates that is compatible with existing HTS systems. The device consists of an array of metal pins connected to individually-controlled thermoelectric coolers. 'We use this to perform femtosecond laser microsurgery on C. elegans in microtiter plates and to analyze the regeneration dynamics over time. This analysis shows that neurons tend regenerate in single short bursts that occur stochastically within the first two days post-surgery.
by Christopher B. Rohde.
Ph.D.

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41

Ramamoorthy, Padmapriya. "HIGH MOLECULAR WEIGHT TEAR PROTEINS AND OCULAR SURFACE MUCINS IN CONTACT LENS-RELATED DRY EYE." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1316523981.

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42

Friedel, Caroline Christina. "Analysis of High-Throughput Data - Protein-Protein Interactions, Protein Complexes and RNA Half-life." Diss., lmu, 2009. http://nbn-resolving.de/urn:nbn:de:bvb:19-96883.

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43

Onoyama, Hiroyuki. "Estimation of Nitrogen Content of Rice Plants and Protein Content of Brown Rice Using Ground-Based Hyperspectral Imagery." Kyoto University, 2016. http://hdl.handle.net/2433/215597.

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Анотація:

Kyoto University (京都大学)
0048
新制・課程博士
博士(農学)
甲第19771号
農博第2167号
新制||農||1040(附属図書館)
学位論文||H28||N4987(農学部図書室)
32807
京都大学大学院農学研究科地域環境科学専攻
(主査)教授飯田訓久, 教授近藤直, 准教授中村公人
学位規則第4条第1項該当

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44

Östensson, Frida. "Grain protein content and its assocoation with the NAC-protein genes HvNAM1 and HvNAM2 in Nordic barley." Thesis, Linköpings universitet, Biologi, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-129390.

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Hunger is a problem faced by many people all over the world, and as the population grows, so does the need for food such as cereals. Because of this, the need for food with higher protein and nutrient content will be increasingly important. NAM-B1, a NAC-protein gene in wheat, has been shown to control the grain protein content and nutrient values, as well as senescence. In barley, two orthologous genes have been found, HvNAM1 and HvNAM2. This study focuses on Nordic barley accessions and how haplotypes of HvNAM1 and HvNAM2 correlate to the grain protein content (GPC) and nutrient content. No correlations between the different haplotypes of the HvNAM genes and the nutrient content and GPC were found. No differences in nutrient content and GPC were found in Nordic accessions originating from Sweden, Norway, Finland, or Denmark, nor were differences found for improvements status groups or for six-row barley and two-row barley. The Nordic accessions were shown to generally have high GPC when compared to control groups Karl and Lewis. However, even if the results of this study indicate that the HvNAM genes do not have major effects on the nutrient contents or GPC, Nordic barley might still be good material for plant improvement. Other factors such as other genes, environmental effects, and gene expression should therefore be investigated.

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45

Pandey, Bikesh. "Analysis of Protein-Protein Interaction Networks Using High Performance Scalable Tools." ScholarWorks@UNO, 2018. https://scholarworks.uno.edu/honors_theses/113.

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Protein-Protein Interaction (PPI) Research currently generates an extraordinary amount of publications and interest in fellow computer scientists and biologists alike because of the underlying potential of the source material that researchers can work with. PPI networks are the networks of protein complexes formed by biochemical events or electrostatic forces serving a biological function [1]. Since the analysis of the protein networks is now growing, we have more information regarding protein, genomes and their influence on life. Today, PPI networks are used to study diseases, improve drugs and understand other processes in medicine and health that will eventually help mankind. Though PPI network research is considered extremely important in the field, there is an issue – we do not have enough people who have enough interdisciplinary knowledge in both the fields of biology and computer science; this limits our rate of progress in the field. Most biologists that are not expert coders need a way of calculating graph values and information that will help them analyze the graphs better without having to manipulate the data themselves. In this research, I test a few ways of achieving results through the use of available frameworks and algorithms, present the results and compare each method’s efficacy. My analysis takes place on very large datasets where I calculate several centralities and other data from the graph using different metrics, and I also visualize them in order to gain further insight. I also managed to note the significance of MPI and multithreading on the results thus obtained that suggest building scalable tools will help improve the analysis immensely.

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46

Mohan, Nitin. "Low-Power High-Performance Ternary Content Addressable Memory Circuits." Thesis, University of Waterloo, 2006. http://hdl.handle.net/10012/2873.

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Ternary content addressable memories (TCAMs) are hardware-based parallel lookup tables with bit-level masking capability. They are attractive for applications such as packet forwarding and classification in network routers. Despite the attractive features of TCAMs, high power consumption is one of the most critical challenges faced by TCAM designers. This work proposes circuit techniques for reducing TCAM power consumption. The main contribution of this work is divided in two parts: (i) reduction in match line (ML) sensing energy, and (ii) static-power reduction techniques. The ML sensing energy is reduced by employing (i) positive-feedback ML sense amplifiers (MLSAs), (ii) low-capacitance comparison logic, and (iii) low-power ML-segmentation techniques. The positive-feedback MLSAs include both resistive and active feedback to reduce the ML sensing energy. A body-bias technique can further improve the feedback action at the expense of additional area and ML capacitance. The measurement results of the active-feedback MLSA show 50-56% reduction in ML sensing energy. The measurement results of the proposed low-capacitance comparison logic show 25% and 42% reductions in ML sensing energy and time, respectively, which can further be improved by careful layout. The low-power ML-segmentation techniques include dual ML TCAM and charge-shared ML. Simulation results of the dual ML TCAM that connects two sides of the comparison logic to two ML segments for sequential sensing show 43% power savings for a small (4%) trade-off in the search speed. The charge-shared ML scheme achieves power savings by partial recycling of the charge stored in the first ML segment. Chip measurement results show that the charge-shared ML scheme results in 11% and 9% reductions in ML sensing time and energy, respectively, which can be improved to 19-25% by using a digitally controlled charge sharing time-window and a slightly modified MLSA. The static power reduction is achieved by a dual-VDD technique and low-leakage TCAM cells. The dual-VDD technique trades-off the excess noise margin of MLSA for smaller cell leakage by applying a smaller VDD to TCAM cells and a larger VDD to the peripheral circuits. The low-leakage TCAM cells trade off the speed of READ and WRITE operations for smaller cell area and leakage. Finally, design and testing of a complete TCAM chip are presented, and compared with other published designs.

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47

Suwannakarn, Kaewta. "Biodiesel production from high free fatty acid content feedstocks." Connect to this title online, 2008. http://etd.lib.clemson.edu/documents/1211389077/.

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48

Munoz, Antonio. "High performance platform independent content analysis for network processing." Thesis, Queen's University Belfast, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.602692.

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The Internet is the global infrastructure for communication, education, entertainment and commerce. As network systems increase in connection speeds and data volume, high performance network intrusion detection and prevention systems must evolve to protect users and businesses from organized and opportunistic crimes motivated by financial and political interests. A detailed study of several well-known network intrusion detection and prevention systems (e.g. Snort) revealed the platform dependency of security rules notation. This thesis describes the design and implementation of Snort2regex, an efficient and accurate tool for compiling Snort rules into regular expression syntax. The regular expression syntax provides a platform independent notation that ensures high levels of security in multiple environments. Several alternative parallel architectures are introduced to attempt to improve the performance of network intrusion detection and prevention systems. I~ order to show the benefits of the Snort2regex compiler, this work also presents SnortEX, a novel software based network intrusion detection and prevention system that benefits from the scalability of the parallel architectures previously introduced. The proposed architecture of SnortEX was evaluated. and several methods of optimization are studied [0 improve the performance and integration between the Snort2regex compiled rule set and SnortEX. Finally, the system is benchmarked and shows a 3 to 17x improvement in performance against a standard Snort implementation.

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49

Marshall, Andrew. "Turbulent flame propagation characteristics of high hydrogen content fuels." Diss., Georgia Institute of Technology, 2015. http://hdl.handle.net/1853/53859.

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Increasingly stringent pollution and emission controls have caused a rise in the use of combustors operating under lean, premixed conditions. Operating lean (excess air) lowers the level of nitrous oxides (NOx) emitted to the environment. In addition, concerns over climate change due to increased carbon dioxide (CO2) emissions and the need for energy independence in the United States have spurred interest in developing combustors capable of operating with a wide range of fuel compositions. One method to decrease the carbon footprint of modern combustors is the use of high hydrogen content (HHC) fuels. The objective of this research is to develop tools to better understand the physics of turbulent flame propagation in highly stretch sensitive premixed flames in order to predict their behavior at conditions realistic to the environment of gas turbine combustors. This thesis presents the results of an experimental study into the flame propagation characteristics of highly stretch-sensitive, turbulent premixed flames generated in a low swirl burner (LSB). This study uses a scaling law, developed in an earlier thesis from leading point concepts for turbulent premixed flames, to collapse turbulent flame speed data over a wide range of conditions. The flow and flame structure are characterized using high speed particle image velocimetry (PIV) over a wide range of fuel compositions, mean flow velocities, and turbulence levels. The first part of this study looks at turbulent flame speeds for these mixtures and applies the previously developed leading points scaling model in order to test its validity in an alternate geometry. The model was found to collapse the turbulent flame speed data over a wide range of fuel compositions and turbulence levels, giving merit to the leading points model as a method that can produce meaningful results with different geometries and turbulent flame speed definitions. The second part of this thesis examines flame front topologies and stretch statistics of these highly stretch sensitive, turbulent premixed flames. Instantaneous flame front locations and local flow velocities are used to calculate flame curvatures and tangential strain rates. Statistics of these two quantities are calculated both over the entire flame surface and also conditioned at the leading points of the flames. Results presented do not support the arguments made in the development of the leading points model. Only minor effects of fuel composition are noted on curvature statistics, which are mostly dominated by the turbulence. There is a stronger sensitivity for tangential strain rate statistics, however, time-averaged values are still well below the values hypothesized from the leading points model. The results of this study emphasize the importance of local flame topology measurements towards the development of predictive models of the turbulent flame speed.

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50

Nilsson, Daniel. "Doping of high-Al-content AlGaN grown by MOCVD." Doctoral thesis, Linköpings universitet, Halvledarmaterial, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-106733.

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The high-Al-content AlxGa1-xN, x > 0.70, is the principal wide-band-gap alloy system to enable the development of light-emitting diodes operating at the short wavelengths in the deep-ultraviolet, λ < 280 nm. The development of the deep-ultraviolet light-emitting diodes (DUV LEDs) is driven by the social and market impact expected from their implementation in portable units for water disinfection and based on the damaging effect of the deep-ultraviolet radiation on the DNA of various microorganisms. Internationally, intense research and technology developments occur in the past few years, yet, the external quantum efficiency of the DUV LEDs is typically below 1%. One of the main material issues in the development of the DUV LEDs is the achievement of n- and ptype doped layers of high-Al-content AlxGa1-xN with low resistivity, which is required for the electrical pumping of the diodes. The doping process, however, becomes significantly more complex with increasing the Al content and the resistivity value can be as high as 101-102 Ω cm for n-type AlN doped by silicon, and 107-108 Ω cm for p-type AlN doped by magnesium. The present study is therefore focused on gaining a better understanding of the constraints in the doping process of the high-Al-content AlxGa1-xN alloys, involving mainly the silicon dopant. For this purpose, the epitaxial growth of the high-Al-content AlxGa1-xN and AlN by the implementation of the distinct hot-wall MOCVD is developed in order to achieve layers of good structural and morphological properties, and with low content of residual impurities, particularly oxygen and carbon. Substitutional point defects such as ON and CN may have a profound impact on the doping by their involvement in effects of n-type carrier compensation. The process temperature can be set from 1000 °C and up to 1400 °C in the present study, which is a principal advantage in order to optimize the material properties of the high-Al-content AlxGa1-xN and AlN. The epitaxial growth of the high-Alcontent AlxGa1-xN and AlN is largely performed on 4H-SiC substrates motivated by (i) the lattice mismatch of ~ 1% along the basal plane (the smallest among other available substrates including Si and sapphire), (ii) the good thermal conductivity of 3.7 W cm-1 K-1, which is essential to minimize the self-heating during the operation of any light-emitting diode, and (iii) the limited access to true-bulk AlN wafers. The Si doping is investigated over a large range of [Si] ~ 1×1017 cm-3 - 1×1020 cm-3. Only the high doping range of [Mg] ~ (1-3)×1019 cm-3 is targeted motivated by the large thermal ionization energy of this common acceptor (from 200 meV in GaN to about 630 meV in AlN). The material characterization involves extensive implementation of atomic force microscopy (AFM), x-ray diffraction (XRD), cathodoluminescence (CL), secondary ion mass spectrometry (SIMS), capacitancevoltage measurements, as well as measurements of the conductivity of the layers by contactless microwave-based technique. The possibility to perform electron paramagnetic resonance (EPR) measurements on the Si-doped high-Al-content AlxGa1-xN is essential in order to establish any effect of self-compensation of the shallow donor state of silicon through the related so-called DX state. The EPR measurements corroborate the study of the incorporation kinetics of silicon and oxygen at various process temperatures and growth rates. The outcome of this study is accordingly summarized and presents our understanding for (i) the complex impact of silicon and oxygen on the n-type conductivity of Al0.77Ga0.23N, which is the alloy composition at which a drastic reduction of the n-type conductivity of high-Al-content AlxGa1-xN is commonly reported (paper 1); (ii) the strain and morphology compliance during the intentional doping by silicon and magnesium, and its correlation with the resistivity in the highly doped layers of Al0.82Ga0.18N alloy composition (paper2); (iii) the n-type conductivity of highly-Si-doped Al0.72Ga0.28N layers as bound by the process temperature (paper 3); and (iv) the shallow donor or DX behavior of the Si dopant in conductive AlxGa1-xN layers, 0.63 ≤ x ≤ 1 (paper 4). It is noted that the measured n-type conductivity in reference layers of Al0.77Ga0.23N, alternatively Al0.72Ga0.28N, alloy composition is on par with the state-of-the-art values, i.e. ≤ 0.05 Ω cm, and 0.012 Ω cm, respectively. A room-temperature resistivity of 7 kΩ cm is measured in Mg-doped layers of Al0.85Ga0.15N alloy composition, which is superior to the state-of-art values (paper 5). The performance of the transport properties of the high-Al-content AlxGa1-xN layers is expected to improve with improvement of their material quality. This can be achieved by improvement of the crystalline quality of the AlN-on-SiC template and by the implementation of true-bulk AlN substrates. The AlN heteroepitaxial growth at the process temperatures of 1100-1200 °C is therefore investigated (paper 6). The lattice constants, structural and optical properties of true-bulk, homoepitaxial and heteroepitaxial AlN material grown at high process temperatures of up to 1400 °C is further reported (paper 7).

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