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La coopétition dans les petites entreprises : le cas des agences immobilières / Coopetition in small businesses : the case of real estate agenciesMira, Benjamin 09 December 2016 (has links)
L’objectif de cette recherche est d’étudier la coopétition dans les petites entreprises. La coopétition est un sujet de recherche depuis vingt ans maintenant. Cette stratégie fait particulièrement l’objet d’un nombre croissant de publications depuis la fin des années 2000. La coopétition apparaît dans la littérature comme une stratégie paradoxale porteuse de performances mais également de risques pour les entreprises. Ces risques peuvent réduire la performance des entreprises ou bien conduire à l’échec de cette stratégie. Un management de la coopétition apparaît donc comme primordial pour bénéficier pleinement des avantanges de cette stratégie. Cependant, la plupart des études ont été conduites dans des grandes entreprises. Nous ne savons que très peu de choses sur la coopétition dans les petites entreprises. De par la spécificité des petites entreprises d’une part, et la multiplication des stratégies de coopétition dans les petites entreprises d’autre part, la recherche sur la coopétition dans ces entreprises présente de forts enjeux théoriques et empiriques. Cette étude cherche donc à expliquer comment fonctionne la coopétition dans les petites entreprises, c’est-à-dire à décrire ses modalités, à en définir un management efficace pour gérer les risques coopétitifs et à en évaluer l’impact sur la performance. Nous mobilisons une approche qualitative et une approche quantitative pour répondre à ces questions en étudiant le cas des agences immobilières en France. Les résultats de cette étude montrent que : (1) la coopétition dans les petites entreprises obéit drastiquement à la Loi proxémique ; (2) les principes d’intégration individuelle du paradoxe coopétitif, de séparation organisationnelle et de co-management des activités communes entre coopétiteurs sont nécessaires pour manager les risques coopétitifs ; (3) la coopétition impacte positivement la performance dans un contexte de petites entreprises. Les résultats montrent tout l’intérêt de la coopétition pour les petites entreprises et contribue à la connaissance de la théorie générale de la coopétition. / The objective of this research is to study coopetition in small businesses. Coopetition is a subject of research for twenty years now. This strategy is particularly the subject of a growing number of publications since the late 2000s. Coopetition appears in the literature as a paradoxical strategy bringing performance but also risks for companies. These risks may reduce the performance of companies or lead to the failure of this strategy. A particular management of coopetition appears as crucial to fully benefit from the avantanges of this strategy. However, most studies have been conducted in large companies. We know very little about coopetition in small businesses. On the one hand, because of the specificity of small businesses, and on the other hand, the proliferation of coopetition strategies in small businesses, research on coopetition in these companies has strong theoretical and empirical issues. This study seeks to explain how coopetition works in small businesses, that is to say, to describe its modalities, to define an effective management of coopetitive risks and to assess its impact on performance. We mobilize a qualitative approach and a quantitative approach to answer to these questions by studying the case of real estate agencies in France. The results of this study show that: (1) coopetition in small businesses to drastically obeys to the Proximity Law; (2) the management principles of individual integration of coopetitive paradox, of organizational separation, and of co-management of joint activities between coopetitors are necessary to manage coopetitive risks; (3) coopetition positively impacts the performance in a context of small business. The results show the advantage for small businesses to adopt coopetition strategies and contributes to the knowledge on the general theory of coopetition.
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Influência da estrutura espacial e da vegetação sobre a assembléia de aves em remanescentes campestres no sul do Brasil / Influence of spatial and vegetation structure on the bird assemblage in grassland patches in Southern BrazilCamilotti, Vagner Luis January 2009 (has links)
Esse estudo teve como objetivo investigar os fatores estruturantes das taxocenoses de aves que utilizam remanescentes de vegetação campestre na região da Fronteira Oeste do Rio Grande do Sul. Através desse estudo busquei compreender os efeitos da estrutura espacial dos remanescentes (área, formato e proximidade entre remanescentes) e também os efeitos exercidos pela estrutura do habitat no uso desses pelas aves campestres. Amostrei quantitativamente por pontos de contagem a avifauna em 21 manchas de vegetação campestre com diferentes áreas, formas e estrutura da vegetação. Utilizei ferramentas de sensoriamento remoto para avaliar a estrutura espacial dos remanescentes e caracterizei o habitat dessas manchas através de medidas da estrutura da vegetação. Utilizei o método analítico de partição da variância através de análises de regressão múltipla e análise de correlação canônica parciais para quantificar a porção da variância explicada por cada grupo de variáveis (estrutura da vegetação e estrutura espacial dos remanescentes) na variação total da abundância, composição e riqueza da avifauna nas manchas. Registrei um total de 33 espécies de aves campestres, sendo três dessas categorizadas em algum grau de ameaça de extinção: Circus cinereus, Cisthotorus platensis e Sporophila cinnamomea. Meus resultados indicaram que a estrutura espacial das manchas não exerce efeito significativo sobre a avifauna campestre e a essa responde principalmente à estrutura e características da vegetação. Dessa forma, meus resultados sugerem que as assembléias de aves que utilizam os remanescentes de vegetação campestre da região estudada são estruturadas a partir das características do habitat e são fracamente influenciadas pela estruturação espacial desses. Assim, a permanência de manchas próximas de vegetação campestre em meio às áreas de cultivo, mesmo que pequenas, poderá favorecer a permanência de uma alta diversidade de espécies de aves na região estudada. / The goal of this study was to investigate the factors that structure bird assemblages in grassland patches in Campos Biome of Fronteira Oeste region, Rio Grande do Sul, Brazil. Thus, I searched for the effects of the spatial structure of native vegetation patches and the effects caused by their vegetation structure on the avifauna that makes use of them. I utilized remote sensing techniques to evaluate the spatial structure of the patches (area, shape, and proximity among patches) and I characterized the patches’ habitat by measuring the vegetation structure (vegetation height, dead standing vegetation, soil covered by vegetation, percentage coverage by grasses, herbs, and shrubs). I used the variation partitioning method to quantify the portion of variation explained by both spatial and vegetation group of variables in the total avifauna’s abundance, composition, and richness variation. I registered 33 grassland bird species and among them, three are threatened with extinction: Circus cinereus, Cisthotorus platensis, and Sporophila cinnamomea. My results denote an insignificant effect of spatial structure of patches on the grassland birds and a high response to the vegetation structure. In this way, my results suggest that the grassland bird’s assemblages that use native vegetation patches are mainly structured by the habitat characteristics (vegetation structure), and are weakly influenced by the patches spatial arrangement. Thus, the maintenance of neighbour patches, even though small ones, into the arable fields may favour the persistence of a high diversity of grassland birds in the region studied.
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Innovation policy transfer in developing countries : a comparative analysis of organisational schemes in the national innovation systems of Egypt and MoroccoSelmanovic, Sandra January 2015 (has links)
The practice of forcing onto developing countries policies developed in other contexts assumes standardised paths to social and economic growth. These policies should enable rapid economic development by “correcting” local market inefficiencies, without considering the role these may have in preserving stability and preventing violence. Moreover, this approach does not address the compatibility of such policies with the culture, values and norms of the receiving country. The thesis aims at extending our understanding of success factors for policy transfer, focusing on organisational schemes for innovation support. It follows the pragmatic Research paradigm and adopts a deductive approach using mixed methods to study the link between administrative efficiency and innovation performance in developing countries, an under-researched area. This thesis argues that policies successfully implemented in culturally proximal contexts are more likely to lead to similar results in the receiving context. The results of the quantitative analysis indicate a strong relationship between administrative efficiency and innovation performance in low-middle income countries in the Middle East and North Africa. The performance of organisational schemes supporting innovation in the national innovation systems of Egypt and Morocco is qualitatively assessed on the basis of 72 face-to-face interviews with entrepreneurs, policy makers and academics. The research finds that similar organisational schemes have produced similar results in the two case studies, confirming the working hypothesis and supporting an approach to policy transfer based on “relevant” good practice.
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Agroindústrias rurais familiares e a rede de relações sociais nos mercados de proximidade na região do Corede Jacuí Centro/RSAgne, Chaiane Leal January 2010 (has links)
A diversificação das atividades produtivas e sociais é uma realidade da agricultura familiar brasileira. O processamento da produção agropecuária e a inserção do produto agroindustrial nos mercados são formas desta diversificação. Este trabalho teve como principal objetivo descrever as relações que formam a rede social dos mercados de proximidade, ou seja, demonstrar as relações que as ARFs mantêm com outros agentes para realizar as ações de troca e comercialização de produtos agroindustriais. A região estudada compreende os municípios que compõe o Corede Jacuí Centro: Cachoeira do Sul, Cerro Branco, Novo Cabrais, Paraíso do Sul, Restinga Seca, São Sepé e Vila Nova do Sul. O estudo caracterizouse como exploratório-descritivo, com a coleta de informações secundárias e primárias. As informações do IBGE sobre as indústrias rurais serviram de suporte para definir o número de entrevistas que seriam efetuadas em cada município. Foram realizadas 41 entrevistas, com o auxílio de um formulário semiestruturado e diários de campo. Utilizou-se de metodologia qualitativa (análise de conteúdo) e quantitativa (medidas de dispersão: média aritmética simples e desvio padrão; medidas de localização: máximo e mínimo) para a análise dos dados. A formação social da rede dos mercados de proximidade envolve interações dos produtores com atores de instituições e da comunidade, com intermediários, pontes de inserção, consumidores e demais agentes da sociedade civil. Destaca-se a venda direta para consumidores urbanos, representando uma média de 51% da produção vendida, cujas formas de comercialização são: de porta em porta, por encomenda, em feiras, em eventos e na propriedade. Nas relações de comercialização e trocas com parentes e vizinhos, as famílias comercializam uma média de 14% e 1%, respectivamente. Estas interações são evidenciadas pela troca de produtos, matéria-prima e serviços na atividade agroindustrial. Quanto às relações comerciais com as pontes de inserção e intermediários, correspondem a 19% e 15% da produção vendida, respectivamente. Para os agricultores, as características de diferenciação dos seus produtos são: o natural, o jeito de fazer, o sabor, o trabalho do agricultor para produzir, o preço, a certificação SIM e a produção ecológica. Dentre as inovações destacamse: a embalagem, a consistência, o tempero, os sabores, a produção light, o tamanho, a concentração do produto e a modificação da receita. / The diversification of the social and productive activities is what usually happens in the Brazilian household agriculture. The agricultural production processing and the agro industrial product insertion in the markets are manners of this diversification. This paper had as main objective to describe the relations that constitute the social networking of the proximity markets to the change and merchandising of the agroindustrial products. The region studied comprehends the cities that belong to Corede Jacuí Centro: Cachoeira do Sul, Cerro Branco, Novo Cabrais, Paraíso do Sul, Restinga Seca, São Sepé and Vila Nova do Sul. This study was characterized as an exploratory-descriptive analysis, with the gathering of secondary and primary information. The data about agrobusiness provided by IBGE gave a support to define the number of interviews that would be done in each city. 41 interviews have been accomplished, with the assistance of a semi-structured form and field diaries. Qualitative methodology has been used (content analysis) and quantitative (disperse measurements: simple arithmetic average and standard deviation; localization measurements: maximum and minimum) for the data analysis. The social formation of the proximate market networking involves interactions of the producers with actors of institutions and the community, with middlemen, insertion bridges, consumers and other agents of the civil society. The direct sale to the urban consumer has been highlighted, representing an average of 51% of the sold production, whose merchandising forms are: door to door, by order, at fairs, at events and at the property. In the relationships of marketing and exchanges with relatives and neighbors, the families commercialized an average of 14% and 1% respectively. These interactions are spotlighted by the exchange of products, raw material, and services at the agroindustrial activity. Concerned with the commercial relationship with the insertion bridges and middlemen, it corresponds to 19% and 15% of the commercialized production, respectively. According to the peasants, the characteristics that differ their products are: the natural, the way to make, the flavor, the farmer’s work to produce, the price, the SIM certification and the ecological production. Among the innovations some are pointed out: the package, the consistency, the seasoning, the flavors, the light production, the size, the product concentration and the recipe modification.
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Les effets de l'Internet sur les règles de conflit de compétence internationale : comparaison entre les droits irakien, français et américain / The effects of the Internet on international competence conflict rulesObbed, Khair Al Deen Kadhim 28 June 2016 (has links)
La législation irakienne définit le contrat comme étant l’union d'une offre faite par la partie contractante avec l'acceptation d'une autre partie et ce de manière à établir les effets dans l'objet du contrat. Ainsi, la place occupée par le contrat de vente en droit irakien est importante. Lorsque les parties relèvent d’ordres juridiques différents, leurs rapports sont régis par le droit international privé, qui détermine le tribunal apte à trancher le litige. Cette thèse de doctorat vise alors à vérifier la capacité à appliquer les règles de conflit de compétence internationale en droit irakien sur des contrats « virtuels » ou dématérialisés. Comme nous le verrons, dans ce domaine, « virtuel » ne veut pas pour autant dire que ce contrat n’est pas réel, comme le spécifie très clairement la loi irakienne. Il reste rattaché au territoire. Le problème est que le droit irakien, en ignorant les notions de frontière et de territorialité, ne reconnaît pas sa propre « immatérialité ». Cette réalité dans les textes et la pratique implique que les opérations qui se produisent sur Internet ne sont pas prises en compte par les règles de conflits de compétence internationale. C’est la raison pour laquelle nous avons souhaité vérifier et comprendre la capacité et l’effectivité des règles de conflit de compétence internationale dans le cadre de litiges sur Internet. Ce faisant, nous espérons mettre en lumière les règles les plus appropriées, qui correspondent le mieux à la nature du contrat virtuel, à savoir son immatérialité. Pour ce faire, nous entreverrons quelques développements sur les litiges de l’Internet. Ainsi nous disposons de deux domaines de recherche : un premier au niveau de la législation nationale, comme le droit français et le droit américain ; un deuxième au niveau des conventions internationales, comme les conventions des Nations-Unies en 2005, la convention de la Haye en 2005, la convention de Bruxelles en 1968 ou encore les règlements de Bruxelles 2000 et 2012. / Iraqi law defines the contract as the union of an offer made by the contracting party with the acceptance of another party and that in order to establish the effects in the contract. The place of the sales contract under Iraqi law is important. When the parties come from different legal orders, their relations are governed by the private international law which determines the court will decide. This thesis research aims to test the ability to apply the international rules of conflict of jurisdiction under Iraqi law on the virtual contract, which is paperless. We shall see, this is not to mean however that the contract is not real, as clearly specifies Iraqi law. It remains attached to the territory. In contrast, the Iraqi law does not recognize its immaterial that meanwhile ignores borders and notions of territoriality. This reality in the texts and practices therefore makes transactions that occur on the Internet are not taken into account by the rules of international jurisdiction conflicts. That is why, we wanted to check and understand the capacity and effectiveness of international jurisdiction conflict rules in the context of Internet disputes. So we will try to find the most appropriate rules, consistent with the nature of the virtual contract, namely its immateriality. This search will reveal developments in litigation of the Internet. Thus, it takes two directions: first at the national legislation, such as French and US law. Second, at the stage of international conventions such as the United Nations Conventions 2005, Hague Convention, the Brussels Convention 1968 and the 2000 and 2012 regulations.
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A framework for modelling spatial proximityBrennan, Jane, Computer Science & Engineering, Faculty of Engineering, UNSW January 2009 (has links)
The concept of proximity is an important aspect of human reasoning. Despite the diversity of applications that require proximity measures, the most intuitive notion is that of spatial nearness. The aim of this thesis is to investigate the underpinnings of the notion of nearness, propose suitable formalisations and apply them to the processing of GIS data. More particularly, this work offers a framework for spatial proximity that supports the development of more intuitive tools for users of geographic data processing applications. Many of the existing spatial reasoning formalisms do not account for proximity at all while others stipulate it by using natural language expressions as symbolic values. Some approaches suggest the association of spatial relations with fuzzy membership grades to be calculated for locations in a map using Euclidean distance. However, distance is not the only factor that influences nearness perception. Hence, previous work suggests that nearness should be defined from a more basic notion of influence area. I argue that this approach is flawed, and that nearness should rather be defined from a new, richer notion of impact area that takes both the nature of an object and the surrounding environment into account. A suitable notion of nearness considers the impact areas of both objects whose degree of nearness is assessed. This is opposed to the common approach of only taking one of both objects, seen as a reference to assess the nearness of the other to it, into consideration. Cognitive findings are incorporated to make the framework more relevant to the users of Geographic Information Systems (GIS) with respect to their own spatial cognition. GIS users bring a wealth of knowledge about physical space, particularly geographic space, into the processing of GIS data. This is taken into account by introducing the notion of context. Context represents either an expert in the context field or information from the context field as collated by an expert. In order to evaluate and to show the practical implications of the framework, experiments are conducted on a GIS dataset incorporating expert knowledge from the Touristic Road Travel domain.
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Establishing very low speed, disturbance-free flow for anemometry in turbulent boundary layersLanspeary, Peter V. January 1998 (has links)
This document addresses problems encountered when establishing the very low air-flow speeds required for experimental investigations of the mechanisms of low-Reynolds-number boundary-layer turbulence. Small-scale motions in the near-wall region are important features of turbulent boundary-layer dynamics, and, if these features are to be resolved by measurements in air with conventionally-sized hot-wire probes, a well-behaved canonical turbulent boundary layer must be developed at free stream flow speeds no higher than 4 m/s. However, at such low speeds, the turbulent boundary layers developed on the walls of a wind tunnel are very susceptible to perturbation by non-turbulent time-dependent flow structures which originate upstream from the test section in the laminar flow at the inlet and in the contraction. Four different non-turbulent flow structures have been identified. The first is a result of quasi-two-dimensional separation of the laminar boundary-layer from the surfaces of the wind-tunnel contraction. Potential flow simulations show that susceptibility to this form of separation is reduced by increasing the degree of axisymmetry in the cross-section geometry and by decreasing the streamwise curvature of the concave surfaces. The second source of time-dependence in the laminar boundary-layer flow is an array of weak streamwise vortices produced by Goertler instability. The Goertler vortices can be removed by boundary-layer suction at the contraction exit. The third form of flow perturbation, revealed by visualisation experiments with streamers, is a weak large-scale forced-vortex swirl produced by random spatial fluctuations of temperature at the wind-tunnel inlet. This can be prevented by thorough mixing of the inlet flow; for example, a centrifugal blower installed at the inlet reduces the amplitude of temperature nonuniformity by a factor of about forty and so prevents buoyancy-driven swirl. When subjected to weak pressure gradients near the start of a wind-tunnel contraction, Goertler vortices in laminar wall layers can develop into three-dimensional separations with strong counter-rotating trailing vortices. These trailing vortices are the fourth source of unsteady flow in the test-section. They can be suppressed by a series of appropriately located screens which remove the low-speed-streak precursors of the three-dimensional separations. Elimination of the above four contaminating secondary flows permits the development of a steady uniform downstream flow and well-behaved turbulent wall layers. Measurements of velocity in the turbulent boundary layer of the test-section have been obtained by hot-wire anemometry. When a hot-wire probe is located within the viscous sublayer, heat transfer from the hot-wire filament to the wall produces significant errors in the measurements of both the mean and the fluctuating velocity components. This error is known as wall-proximity effect and two successful methods are developed for removing it from the hot-wire signal. The first method is based on the observation that, if all experimental parameters except flow speed and distance from the wall are fixed, the velocity error may be expressed nondimensionally as a function of only one parameter, in the form DeltaU^+=f(y^+). The second method, which also accommodates the effect of changing the hot-wire overheat ratio, is based on a dimensional analyis of heat transfer to the wall. Velocity measurements in the turbulent boundary layer at the mid-plane of a nearly square test-section duct have established that, when the boundary-layer thickness is less than one quarter of the duct height, mean-velocity characteristics are indistinguishable from those of a two-dimensional flat-plate boundary layer. In thicker mid-plane boundary layers, the mean-velocity characteristics are affected by stress-induced secondary flow and by lateral constriction of the boundary-layer wake region. A significant difference between flat-plate and duct boundary layers is also observed in momentum-balance calculations. The momentum-integral equation for a duct requires definitions of momentumd and displacement thickness which are different from those given for flat-plate boundary layers. Momentum-thickness growth rates predicted by the momentum-integral equation for a duct agree closely with measurements of the newly defined duct momentum thickness. Such agreement cannot be obtained in terms of standard flat-plate momentum thickness. In duct boundary layers with Reynolds numbers Re_theta between 400 and 2600, similarity in the wake-region distributions of streamwise turbulence statistics has been obtained by normalising distance from the wall with the flat-plate momentum thickness, theta_2. This result indicates that, in contrast with the mean velocity characteristics, the structure of mid-plane turbulence does not depend on the proportion of duct cross-section occupied by boundary layers and is essentially the same as in a flat-plate boundary layer. For Reynolds numbers less than 400, both wall-region and wake-region similarity fail because near-wall turbulence events interact strongly with the free stream flow and because large scale turbulence motions are directly influenced by the wall. In these conditions, which exist in both duct and flat-plate turbulent boundary layers, there is no distinct near-wall or wake region, and the behaviour of turbulence throughout the boundary layer depends on both wall variables and on outer region variables simultaneously. / Thesis (Ph.D.)--School of Mechanical Engineering, 1998.
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Proximity Ligation : Transforming protein analysis into nucleic acid detection through proximity-dependent ligation of DNA sequence tagged protein-bindersFredriksson, Simon January 2002 (has links)
<p>A novel technology for protein detection, proximity ligation, has been developed along with improved methods for <i>in situ</i> synthesis of DNA microarrays. Proximity ligation enables a specific and quantitative transformation of proteins present in a sample into nucleic acid sequences. As pairs of so-called proximity probes bind the individual target protein molecules at distinct sites, these reagents are brought in close proximity. The probes consist of a protein specific binding part coupled to an oligonucleotide with either a free 3’- or 5’-end capable of hybridizing to a common connector oligonucleotide. When the probes are in proximity, promoted by target binding, then the DNA strands can be joined by enzymatic ligation. The nucleic acid sequence that is formed can then be amplified and quantitatively detected in a real-time monitored polymerase chain reaction. This convenient assay is simple to perform and allows highly sensitive protein detection. Parallel analysis of multiple proteins by DNA microarray technology is anticipated for proximity ligation and enabled by the information carrying ability of nucleic acids to define the individual proteins. Assays detecting cytokines using SELEX aptamers or antibodies, monoclonal and polyclonal, are presented in the thesis.</p><p>Microarrays synthesized <i>in situ</i> using photolithographic methods generate impure products due to damaged molecules and interrupted synthesis. Through a molecular inversion mechanism presented here, these impurities may be removed. At the end of synthesis, full-length oligonucleotides receive a functional group that can then be made to react with the solid support forming an arched structure. The 3’-ends of the oligonucleotides are then cleaved, removing the impurities from the support and allowing the liberated 3’-hydroxyl to prime polymerase extension reactions from the inverted oligonucleotides. The effect of having pure oligonucleotides probes compared to ones contaminated with shorter variants was investigated in allele specific hybridization reactions. Pure probes were shown to have greater ability to discriminate between matched and singly mismatched targets at optimal hybridization temperatures.</p>
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Critical Phenomena and Exchange Coupling in Magnetic HeterostructuresAhlberg, Martina January 2012 (has links)
The continuous phase transition in thin magnetic films and superlattices has been studied using the magneto-optical Kerr effect (MOKE) and polarized neutron scattering (PNR). It has been shown that the critical behavior of amorphous thin films belonging to the 2D XY universality class can be described within the same theory as crystalline sample. This means that quenched disorder only serves as a marginal perturbation in systems with this symmetry. The connection between interlayer exchange coupling and the observed critical behavior in Fe/V superlattices was explored. The results prove that the origin of unusually high values of the exponent β can be traced to a position dependence of the magnetization at elevated temperatures. The magnetization of the outermost layers within the superlattice shows a more pronounced decrease at lower temperatures, compared to the inner layers, which in turn have a more abrupt decrease in the vicinity of the critical temperature. This translates to a high exponent, especially when the layers are probed by a technique where more weight is given to the layers close to the surface, e.g.MOKE. The interlayer exchange coupling as a function of spacer thickness and temperature was also studied in its own right. The data was compared to the literature, and a dependence on the thickness of the magnetic layers was concluded. The phase transition in amorphous FeZr/CoZr multilayers, where the magnetization emanates from ferromagnetic proximity effects, was investigated. Even though the determined exponents of the zero-field magnetization, the susceptibility and the critical isotherm did not correspond to any universality class, scaling plots displayed an excellent data collapse. Samples consisting of Fe δ-layers (0.3-1.4 monolayers) embedded in Pd were studied using element-specific resonant x-ray magnetic scattering. The magnetization of the two constituents showed distinctly different temperature dependences.
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Proximity Ligation : Transforming protein analysis into nucleic acid detection through proximity-dependent ligation of DNA sequence tagged protein-bindersFredriksson, Simon January 2002 (has links)
A novel technology for protein detection, proximity ligation, has been developed along with improved methods for in situ synthesis of DNA microarrays. Proximity ligation enables a specific and quantitative transformation of proteins present in a sample into nucleic acid sequences. As pairs of so-called proximity probes bind the individual target protein molecules at distinct sites, these reagents are brought in close proximity. The probes consist of a protein specific binding part coupled to an oligonucleotide with either a free 3’- or 5’-end capable of hybridizing to a common connector oligonucleotide. When the probes are in proximity, promoted by target binding, then the DNA strands can be joined by enzymatic ligation. The nucleic acid sequence that is formed can then be amplified and quantitatively detected in a real-time monitored polymerase chain reaction. This convenient assay is simple to perform and allows highly sensitive protein detection. Parallel analysis of multiple proteins by DNA microarray technology is anticipated for proximity ligation and enabled by the information carrying ability of nucleic acids to define the individual proteins. Assays detecting cytokines using SELEX aptamers or antibodies, monoclonal and polyclonal, are presented in the thesis. Microarrays synthesized in situ using photolithographic methods generate impure products due to damaged molecules and interrupted synthesis. Through a molecular inversion mechanism presented here, these impurities may be removed. At the end of synthesis, full-length oligonucleotides receive a functional group that can then be made to react with the solid support forming an arched structure. The 3’-ends of the oligonucleotides are then cleaved, removing the impurities from the support and allowing the liberated 3’-hydroxyl to prime polymerase extension reactions from the inverted oligonucleotides. The effect of having pure oligonucleotides probes compared to ones contaminated with shorter variants was investigated in allele specific hybridization reactions. Pure probes were shown to have greater ability to discriminate between matched and singly mismatched targets at optimal hybridization temperatures.
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