CHIKV is the prototype of Alphaviruses and it causes an acute febrile illness with rash, severely painful arthralgias, and sometimes arthritis. While CHIKV has first been identified in the 1950s in Africa, recent outbreaks of CHIKV in the islands of the Indian Ocean and particular in Italia have re-drawn attention to CHIKV. In the past CHIKV disease was considered self-limiting and non-fatal. However, a number of deaths on Reunion (Anonym, 2006) during the outbreak, which was affected directly or indirectly by CHIKV, have changed this view. To defeat CHIKV outbreaks diagnostic tools and anti CHIKV therapies are urgently needed. In this thesis, we generated tools to investigate CHIKV at the molecular level by serological tests. CHIKV was isolated from a German woman who was infected during her holidays on the Mauritius Island. To characterize this viral isolate the complete viral genome was amplified by PCR and molecular cloned. In order to analyse antibody responses of infected individuals some of the structural and non-structural genes were subcloned in bacterial expression vectors. The NSP2, proteinase, capsid, E1 and E2 were subsequently expressed in E.coli using purified successfully. In this thesis, the structural proteins were used to develop a screening test for anti-CHIKV antibodies in patient derived serum samples. These tests were evaluated with pre-characterized anti-CHIKV sera (30 samples) obtained from the BNI Hamburg and 100 serum samples from German blood donors used as negative controls. Immunoblotting analysis revealed that up to 77% of precharacterised positive sera could recognize the recombinant proteins and there were no detectable reactivity of CHIKV-negative German donor sera. The recombinant proteins were also recognized by 71.4% of positive sera in the newly established ELISA. In order to go further in analyses of the results, an in house IFA was performed. Positive sera (21 samples) were used. The results showed that all of them reacted positive, but this assay was less sensitive than the IFA from BNI. In comparison with the IFA result from BNI Hamburg, the results were not congruent in all test performed. This could be due to various drawbacks of the tests. A cross reaction in Alphaviruses and the different strains are mentioned as well as the denatured forms of the structural proteins. Besides the main structural proteins (E1, E2 and C), other proteins such as non-structural proteins, uncleaved precursor proteins could participate in the different outcomes of serological assays. In order to go further in the CHIKV diagnoses, the CHIKV recombinant proteins were applied to screen the anti-CHIKV antibodies in the Vietnamese population, who are considered to live in the high risk regions. In serological tests, 158 sera of Vietnamese donors were incubated with the recombinant proteins or the fixed CHIKV infected cells. The results showed that 24% of Vietnamese donor sera recognized the recombinant proteins in immunoblot assay, while 36% scored positive in the ELISA assay. In IFA, the sera considered positive were 11.4%. While some discrepancies in serological tests were found, these results showed that the ratio of CHIKV-positive sera seem to be equal to the other regions in the world, which are affected by CHIKV. It is suggested that CHIKV infection in Vietnam has been repeatedly misdiagnosed. This study cohort consisted only of samples originating from Hanoi area of Northern Vietnam, thus, future studies should expand to include samples from other Vietnam areas. To do this the various subtypes of the virus in the different regions should be isolated and the sequences of these viruses should be well characterized.

Apesar de ser uma criação recente, o cartão de crédito desponta como um dos mais utilizados meios de pagamento, além de instrumento de viabilização de crédito na sociedade atual. É diferenciação relevante a ser notada entre o cartão de crédito, peça plástica, e o sistema de cartão de crédito. Enquanto o primeiro se refere apenas ao documento de legitimação, ou seja, instrumento físico capaz de demonstrar uma relação existente entre o titular do cartão e o emissor, um dos contratos que integram o sistema, o segundo é o produto dos contratos coligados que possibilitam a concretização da operação. Além do contrato já indicado, o contrato entre fornecedor e emissor e o contrato entre fornecedor e titular formam, em um modelo básico, os contratos envolvidos na operação, coligados funcionalmente, ou seja, apesar de manter sua estrutura formal segmentada, formam um conjunto que viabiliza a operação. Os contratos envolvidos, apesar da tipicidade social, com a reiteração de contratações semelhantes, abrangendo os mesmos direitos e obrigações que identificam o modelo, são legalmente atípicos, já que as prestações envolvidas não permitem identificação com os tipos legais atualmente positivados nem assemelhação que permita aplicação das regras próprias daqueles, restando, no direito brasileiro, a vigência de normas e princípios gerais aos contratos e obrigações envolvidos. Os contratos coligados demandam uma recompreensão dos princípios tradicionais dirigidos e aplicáveis aos contratos atomizados, com diversos efeitos para o estudo dos contratos de cartão de crédito e a busca de soluções, até mesmo para as tradicionais questões controvertidas. / Despite being a recent creation, credit cards rise as one of the most used means of payment, besides being instruments of credit viabilization in the current society. There is a relevant differentiation to be observed between credit cards, plastic pieces, and the credit card system. While the former only refers to the legitimation document, that is, to the physical instrument capable of evidencing a relationship existing between the card holder and the issuer, one of the contracts that integrate the system, the latter is the product of the linked contracts that allow the operation execution. Besides that already indicated contract, the contract between suppliers and issuers and the contract between suppliers and holders form, in a basic model, the contracted involved in the operation, functionally linked, that is, although keeping their formal structure segmented, they form a set that feasibilizes the operation. The involved contracts, despite the social typicity, with the reiteration of similar contractings, involving the same rights and obligations that identify the model, are legally atypical, as the considerations involved do not allow identification with the legal types currently ascertained, nor similarity allowing application of their own rules, there being, in the Brazilian laws, the validity of general norms and principles governing the contracts and obligations involved. The linked contracts require a re-understanding of the traditional principles directed towards and applicable to atomized contracts, with various effects for the study of credit card contracts and for the quest for solutions, even for traditionally controvert issues.

Neste trabalho estudamos a dinâmica de partículas e cordas num campo gravitacional com métrica degenerada. Utilizamos o formalismo Hamiltoniano para determinar os vínculos da teoria e a dinâmica efetiva do sistema. Classificamos a estrutura dos vínculos de acordo com a forma de métrica e dimensão do espaço (-tempo). Obtemos também a representação das simetrias em termos de isometrias geradas por auto-vetores nulos da métrica. / In this work we stud the dynamics of particles and strings in a gravitational Field with degenerate metric. We use the Hamiltonian formalism to find out the constraints and the effective dynamics of the system. We classify the constraint structure according to the formo the me-tric and the space(time) dimension. We also obtain the representation of the symmetries in terms of the isometries generated by the null eigenvectors of the metric.

There is a disproportionate burden of disease (measured in daily-adjusted life years, or DALYs) in low-income countries. Much of this disparity is due to infectious diseases: 53% of DALYs in Africa are due to infectious diseases, compared with only 3% in the American continents. This disparity is largely due to differences in electrical and transport infrastructure as well as access to skilled personnel and monetary resources. Current diagnostic solutions are primarily designed for high-resource settings and therefore these solutions cannot be easily translated to a lower-resource setting. In order to tackle this health disparity, new solutions must be designed specifically for a lower-resource setting. In this dissertation, we take a translational approach to engineering appropriate diagnostics for resource-limited settings. First, we develop a handheld smartphone accessory to perform an assay similar to enzyme-linked immunosorbent assay (ELISA), traditionally a laboratory-based test. In 15 minutes, it provides an objective diagnostic readout important for minimal training, while using an average of 1.6mW of power and costing only $34. We further develop the device to provide a quantitative hemoglobin measurement simultaneously with an HIV immunoassay, for use in antenatal care screening. The multiplexing two assay types that are clinically relevant has the potential to streamline workflow. While specifications can be demonstrated in the laboratory, the true test of the device must be performed in the field. We brought our smartphone accessory to three health centers in Kigali, Rwanda to be used by healthcare workers with no prior experience in ELISA. After a short 30 minute training, the healthcare workers were able to obtain diagnostic results comparable to other immunoassays run under field conditions. With a simple and user-friendly design, we sought to further expand the usage of our device as a self-testing device, having patients test themselves. Lastly, we explore manufacturable thermoplastics as a material for a microfluidic diagnostic for nucleic acid detection. The sum of this work aims to gain insight into methods of design, testing, and implementation of translational design.

Neste trabalho estudamos a dinâmica de partículas e cordas num campo gravitacional com métrica degenerada. Utilizamos o formalismo Hamiltoniano para determinar os vínculos da teoria e a dinâmica efetiva do sistema. Classificamos a estrutura dos vínculos de acordo com a forma de métrica e dimensão do espaço (-tempo). Obtemos também a representação das simetrias em termos de isometrias geradas por auto-vetores nulos da métrica. / In this work we stud the dynamics of particles and strings in a gravitational Field with degenerate metric. We use the Hamiltonian formalism to find out the constraints and the effective dynamics of the system. We classify the constraint structure according to the formo the me-tric and the space(time) dimension. We also obtain the representation of the symmetries in terms of the isometries generated by the null eigenvectors of the metric.

Autism is the fastest growing serious developmental disability in the United States and more children will be diagnosed with autism this year than with AIDS, diabetes, and cancer combined (Autism Speaks, 2013). Supporting and caring for the child with autism can be devastating socially and emotionally for the families (Neely, Amatea, Doan, & Tannen, 2012). The purpose of this study is to investigate the changes in the life of families that have a child diagnosed with autism. Four families were interviewed in this qualitative study and each was from East Tennessee. These different families were led by a mother and father, a grandmother, and two divorced mothers. Through this research, I learned how each family was impacted by autism. Autism dramatically affected each family and each was impacted in many different aspects of life.

v anglickém jazyce Theoretical and Practical Aspect of Unit-linked Insurance Abstract United-linked insurance (ULI) is the most widely sold type of life insurance in the Czech Republic; frequently, however, this product is sold to clients without respecting their investment profile and financial needs (mis-selling); mis-selling, in turn, results in the loss of confidence in the system of financial distribution and its efficiency. Even though the current legislation offers tools to mitigate mis-selling (not only) of ULI, in practice such legal regulation proves to be insufficient. The causes of this relate to both the distributors and their customers, as well as to the financial products themselves. Behavioural research has proven marked shortcomings in the economic and legal understanding of a human being as a homo economicus, i.e. as a rationally acting agent. A number of studies evidence that people oftentimes demonstrate limited rationality, which negatively impacts financial distribution, in that these cognitive errors may be purposefully and skilfully abused by distributors who use their marketing and sales strategies to the clients' disadvantage. In this respect, this thesis analyses the current financial-political customer protection doctrines; the status of the ULI customer protection,...

Abnormal T lymphocyte reactions in both autism and Alzheimer's disease (AD) have been reported. This research investigated the possibility that these abnormalities may involve circulating antithymic antibodies. Plasma samples from autistic patients, AD patients, and normal-matched controls were tested for reactivity against murine thymocytes. In the first of 3 studies results of the enzyme-linked immunosorbent assay (ELISA) were statistically significant for binding (P < 0.001) between antithymic antibodies in plasmas of AD patients and murine thymocytes. Binding (P < 0.05) in low dilutions (1/2.5 and 1/5} of autistic patient plasmas was also observed. In the second study, plasmas of neither autistic nor AD patients significantly inhibited DNA synthesis of thymic cells in the presence of interleukin-1 (IL-l} and phytohemagglutinin (PHA). In the third study, no significant increases (P > 0.05) in cytotoxic activities were detected using AD patient plasmas and both untreated and heat-treated autistic patient plasmas. After further testing, these heat-treated plasmas diluted 1/64 and 1/128 had increased cytotoxicities (P Therefore, circulating antithymic antibodies may be involved in abnormal T lymphocyte reactions in autism and AD. Since they probably do not act alone, future research should study these complex abnormalities using human thymocytes.

Spider-silk is known as one of the stronger natural materials, unfortunately it is impossible to farm spiders due to their territorial and cannibalistic nature. To address this issue, researchers have studied spider-silk to discover how it is produced in nature. From their results, spider-silk is composed of large sized proteins produced in two different cell types. Using this knowledge, researchers created transgenic organisms capable of producing spider-silk proteins in large quantities. Using these proteins, several groups have created fibers, films, hydrogels, and adhesives with robust and versatile properties. Wet-spinning is a technique commonly used to create fibers from spider-silk proteins. These fibers unfortunately do not compare to the mechanical properties of natural silk. To address this researchers have used a method known as electrospinning to create spider-silk fibers with substantially smaller diameters. In doing so, these electrospun fibers have increased surface area and enhanced mechanical properties. Using this method, our group has modified the electrospinner to be able to produce continuous fine diameter yarns composed of hundreds of nanofibers with mechanical properties surpassing that of natural silk. Fibers aside, spider-silk proteins can be used to create a variety of different biocompatible materials. To further enhance these materials, our group has utilized a technique traditionally used for observation. This technique employs a high intensity light source to initiate cross-links within the proteins. With this method, our spider-silk protein materials have increased their mechanical properties by a factor of seven. These materials can further be modified through post-treatments, resulting in tunable materials with diverse and robust mechanical properties.

FAPα ist ein Membranglykoprotein mit Dipeptidyl-Peptidase- und Typ I Kollagenase-Aktivität, das eine wichtige Rolle im Rahmen des Gewebeumbaus und der Angiogenese spielt. Die Expression von FAPα wurde für eine Reihe von Geweben gezeigt. So konnte bereits gezeigt werden, dass FAPα nach Myokardinfarkt auf humanen kardialen Fibroblasten (HCF) verstärkt exprimiert wird. Ausgehend von der Erkenntnis, dass eine im Blut zirkulierende Form von FAPα, APCE, gefunden wurde, sollte in der vorliegenden Arbeit eine mögliche Assoziation des in HCF exprimierten FAPα mit dem zirkulierenden APCE untersucht werden. Hierfür wurden ebenfalls die Signalwege, die zur Induktion von FAPα führen, näher untersucht. Im Rahmen der vorliegenden Arbeit konnte gezeigt werden, dass das von humanen kardialen Fibroblasten exprimierte FAPα ebenfalls im Zellkulturüberstand nachgewiesen werden kann. Wie bereits beschrieben, führte eine Stimulation mit TGFβ1 zur Expression von FAPα. Diese ist abhängig von Smad-3 und konnte durch Zugabe des Inhibitors SB431542 blockiert werden. Auch Smad-2 scheint die FAPα-Expression zu beeinflussen. Ein Effekt von EGR-1, CTGF und TNFα auf die FAPα-Expression konnte hingegen nicht nachgewiesen werden. Darüber hinaus wurde im Rahmen der vorliegenden Arbeit ein ELISA zur Messung von humanem FAPα im Plasma erstmalig etabliert. In der Evaluation des ELISA wurden die Grenzwerte für die Detektion und Quantifizierung bestimmt, die Intra- und Intervariabilität des ELISAs berechnet und die Linearität der Wiederfindung von humanem rekombinantem FAPα bewertet, sowie die Stabilität von FAPα nach mehrfachen Einfrier-Auftau-Zyklen und nach der Lagerung bei verschiedenen Temperaturen evaluiert. Da hier nur kommerziell erwerbliche Materialien verwendet wurden, ist eine Anwendung durch Dritte leicht durchführbar. Es wurde weiterhin eine klinische Studie zur Messung der FAPα-Plasmaspiegel in 101 gesunden Blutspendern und 407 Patienten mit akutem Koronarsyndrom (ACS) sowie von 25 Patienten mit stabiler koronarer Herzerkrankung (KHK) durchgeführt. Hierbei konnten erstmals Referenzwerte für zirkulierendes FAPα im Plasma beschrieben werden. Die FAPα-Plasmaspiegel der gesunden Kontrollgruppe unterschieden sich nicht von denen der Patienten mit stabiler KHK. Bei Patienten mit akutem Koronarsyndrom war die FAPα-Konzentration im Vergleich mit den FAPα-Plasmaspiegeln der Kontrollgruppe hingegen signifikant erniedrigt. Auch zeigte sich bei den Patienten mit FAPα-Spiegeln im kleinsten Quartil eine erhöhte Mortalität. Die biologische Funktion von FAPα bzw. dessen mögliche Pathophysiologie im Rahmen eines ACS sowie die mit niedrigen FAPα-Spiegeln assoziierte Mortalität sind noch unklar und bleiben Gegenstand der weiteren Forschung. / FAPα is a membrane protein with dipeptidyl-peptidase and type-I-collagenase activity involved in angiogenesis and tissue repair. A number of cells and tissues including human cardiac fibroblasts (HCF) after myocardial infarction express FAPα. Due to the recent discovery of a circulating form of FAPα, APCE, a possible association between the cell-bound form of FAPα and APCE was studied in HCF. Furthermore, the signaling pathways that lead to an induction of FAPα were analyzed. It could be demonstrated that HCF shed FAPα, which can be detected in the cell culture supernatant. Moreover, in accordance with earlier results, FAPα expression is induced by TGFβ1. This is dependent on Smad-3 and could be blocked by the inhibitor SB431542. Additionally, Smad-2, but not EGR-1, CTGF or TNFα are involved in FAPα expression. Within this work an ELISA for the quantification of FAPα in human plasma was established and the limits of detection and quantification, the intra- and interassay variability and the linearity of recovery of human FAPα were determined. Furthermore, the stability of FAPα after multiple thaw-freeze-cycles or different storage conditions was evaluated. Because of the exclusive use of commercially available products, others could easily repeat this ELISA. FAPα plasma concentrations in 101 healthy blood donors, 407 patients with acute coronary syndrome (ACS) and in 25 patients with stable coronary artery disease were quantified by ELISA. This is the first time reference values for FAPα plasma levels could be derived. There was no significant difference between healthy donors and patients with stable coronary artery disease. However, compared to the healthy control group, FAPα levels were significantly lower in patients with ACS. Importantly, ACS patients with FAPα levels in the lowest quartile showed an increased mortality. The biological function of FAPα and its possible pathophysiological role in ACS are still unclear and remain the object of future studies.