Near-membrane and trans-membrane proteins and their interactions with the extracellular matrix (ECM) can yield valuable information about cell dynamics. However, advances in the field of nanoscale cellular processes have been hindered, in part, due to limits imposed by current technology. In this work, a novel evanescent field (EF) imaging technique is designed, modeled, created and tested for near-field imaging in the apical surface of cells. This technique and Förster resonance energy transfer (FRET) were used to investigate interactions between integrins on the cell surface and the ECM protein, fibronectin. The goal was to monitor changes in the integrin density at the cell surface as a function of clustering after binding to fibronectin on the microsphere surface. For the EF technique, quantum dot (QD)-embedded polystyrene microspheres were used to couple light into whispering gallery modes (WGMs) inside the microspheres; the resulting EF at the surface of the microsphere was used as a near-field excitation source with ~50 nm axial resolution for exciting fluorescently-labeled integrins. For FRET measurements (~10 nm axial resolution), QDs (donors) were coated on the surface of microspheres and energy transfer to red fluorescent protein (RFP)-integrin constructs (acceptors) studied. In both techniques, the QD-modified microspheres were mounted on atomic force microscope (AFM) cantilevers, functionalized with fibronectin, and brought into contact with fluorescently-labeled HeLa or vascular smooth muscle (VSM) cells. The results obtained from both methods show the clustering and activity of the integrins and are in good agreement with each other. Amsterdam discrete dipole approximation (ADDA) was used to study the effects of inhomogeneous surrounding refractive index on the quality factor and position of the WGMs due to the attachment of a microsphere to an AFM cantilever. WGMs of various QD-embedded microspheres mounted on AFM cantilevers were experimentally measured and shown to be consistent with the model.

Texte remanié de: Diss.--Bremen--Universität, 1993. / Bibliogr. p. 204-210.

Submitted by Divisão de Documentação/BC Biblioteca Central (ddbc@ufam.edu.br) on 2015-10-21T15:04:23Z No. of bitstreams: 1 Dissertação - Alexandre M. Aguiar.pdf: 8594585 bytes, checksum: c3945e3c68b06eb68321ddb20ca69d85 (MD5) / Approved for entry into archive by Divisão de Documentação/BC Biblioteca Central (ddbc@ufam.edu.br) on 2015-10-21T15:08:27Z (GMT) No. of bitstreams: 1 Dissertação - Alexandre M. Aguiar.pdf: 8594585 bytes, checksum: c3945e3c68b06eb68321ddb20ca69d85 (MD5) / Approved for entry into archive by Divisão de Documentação/BC Biblioteca Central (ddbc@ufam.edu.br) on 2015-10-21T15:14:08Z (GMT) No. of bitstreams: 1 Dissertação - Alexandre M. Aguiar.pdf: 8594585 bytes, checksum: c3945e3c68b06eb68321ddb20ca69d85 (MD5) / Made available in DSpace on 2015-10-21T15:14:08Z (GMT). No. of bitstreams: 1 Dissertação - Alexandre M. Aguiar.pdf: 8594585 bytes, checksum: c3945e3c68b06eb68321ddb20ca69d85 (MD5) Previous issue date: 2015-04-24 / FAPEAM - Fundação de Amparo à Pesquisa do Estado do Amazonas / This master´s dissertation completes a historical-literary analysis of the novel Howards End, written by the English novelist E. M. Forster. The first chapter presents the author´s life, as well as his travels abroad and novels published. Secondly, we analyze the narrative; emphasize the facts, characters and symbolic elements described by the author. Finally, the third chapter, we compare Howards End with On Beauty by Zadie Smith, approaching the facts, characters and symbolic elements written in these works. Howards End was published in 1910 during the Eduardian time, when the Victorian era is over. There was a conflict in England during this time of intense transition between the “new” and “old”, since England has no longer control over its colonies and political and economic questions in Europe seem to increase with totalitarian regimes in German and Italy. In the narrative, there is a resistance to the new changes represented by the sisters Schlegels who are intellectual and emancipated, daughters of an English mother and a German father. On the other hand, the impositions of traditions represented by the clan Wilcox, as well as the integration of another nucleus, the Basts, originally proletarians in that society. The initial conflict begins when the matriarch of the Wilcox, for a symbolic gesture of friendship, decides to leave her old cottage house, Howards End, to the old sister (Margaret), without letting her know about this decision. The families have their paths crossed with that conflict which is the core of most parts of the novel of E. M. Forster, as he seems to suggest that personal relationships represent the only possibility to comprehension of a chaotic world. / Esta dissertação de mestrado faz uma análise histórico-literária do romance Howards End, do escritor inglês E. M. Forster. O primeiro capítulo apresenta a vida do autor, assim como suas viagens ao exterior e os romances publicados. No segundo, faremos uma análise da narrativa; enfatizando os fatos, as personagens e os elementos descritos pelo autor. Por fim, no terceiro, compararemos Howards End com On Beauty, de Zadie, e faremos uma abordagem dos fatos, das personagens, e elementos descritos nessas obras. Howards End foi publicado em 1910 durante a época Eduardiana, quando o período conhecido como Era Vitoriana se encerra. Nesse momento de intensa transição na Inglaterra havia o conflito entre o “novo” e o “velho”, já que a Inglaterra não possuía mais controle sobre suas colônias, e questões políticas e econômicas na Europa parecem surgir com a ascensão de regimes totalitários na Alemanha e Itália. Na narrativa, há a resistência ao novo representado pelas irmãs Schlegel que são intelectuais e emancipadas, filhas de mãe inglesa e pai alemão e, do outro lado, a imposição das tradições representadas pelo clã Wilcox, assim como a integração de outro núcleo, os Bast, de origem proletária e marginalizada naquela sociedade. O conflito inicial se dá quando a matriarca dos Wilcox, por um gesto simbólico de amizade, decide deixar sua velha casa de campo, Howards End para a irmã mais velha Schlegel (Margaret), sem que essa tome conhecimento da decisão. As famílias têm seus destinos entrecruzados a partir desse conflito que é o cerne de boa parte da obra de E. M. Forster, quando ele parece sugerir que as relações interpessoais representam a única possibilidade de compreensão de um mundo caótico.

Selectins are key adhesion molecules responsible for initiating a multistep process that leads a cell out of the blood circulation and into a tissue or organ. The adhesion of cells (expressing ligands) to the endothelium (expressing the selectin i.e.,E-selectin) occurs through spatio-temporally regulated interactions that are mediated by multiple intra- and inter-cellular components. The mechanism of cell adhesion is investigated primarily using ensemble-based experiments, which provides indirect information about how individual molecules work in such a complex system. Recent developments in single-molecule (SM) fluorescence detection allow for the visualization of individual molecules with a good spatio-temporal resolution nanometer spatial resolution and millisecond time resolution). Furthermore, advanced SM fluorescence techniques such as Förster Resonance Energy Transfer (FRET) and super-resolution microscopy provide unique opportunities to obtain information about nanometer-scale conformational dynamics of proteins as well as nano-scale architectures of biological samples. Therefore, the state-of-the-art SM techniques are powerful tools for investigating complex biological system such as the mechanism of cell adhesion. In this project, several constructs of fluorescently labeled E-selectin will be used to study the conformational dynamics of E-selectin binding to its ligand(s) using SM-FRET and combination of SM-FRET and force microscopy. These studies will be beneficial to fully understand the mechanistic details of cell adhesion and migration of cells using the established model system of hematopoietic stem cells (HSCs) adhesion to the selectin expressing endothelial cells (such as the E-selectin expressing endothelial cells in the bone marrow).

No description available.

No description available.

No description available.

The plasma protein complement factor H (fH, 155 kDa) regulates the activity of the alternative pathway of complement activation. Factor H is monomeric, and its 20 CCP modules are arranged in a predominantly elongated conformation, joined by linking sequences that vary in length, with the longest linkers occurring in the central portion of the molecule. CCP modules 1 through 4 of fH host its capacity to act as a cofactor for fI-mediated proteolytic degradation of C3b and its ability to accelerate the decay of the C3 convertase, C3bBb, thereby regulating the so-called tick-over activation of the alternative pathway. Mutations in this part of fH might compromise its function and lead to underregulation of the alternative pathway. It is hypothesized that this can cause predisposition to diseases such as atypical haemolytic uraemic syndrome (aHUS) and age-related macular degeneration (AMD). In the current work, the known disease-associated mutations R53H and R78G were compared to wild-type in terms of fluid-phase cofactor assays, C3b-binding affinity and the ability to accelerate the decay of the convertase. In addition, the protective variant, I62, was also inspected because its protective role might be explained by an increased regulatory activity. The second, linked, aim of this project was to employ Forster resonance energy transfer (FRET) to study the link between conformation and function in fH. FRET is valuable for obtaining long-distance restraints up to a maximum of 100 °A and is therefore particularly useful for inferring domain orientations within multidomain proteins. This approach to measure long-range inter- and intramolecular distances is a convenient way to complement NMR-based structural investigations, which rely on short-range restraints. It is also a valuable complement to X-ray crystallography since it is a solution technique that can be conducted under physiological conditions. By using site-directed mutagenesis in the current work, free cysteines were introduced into CCP modules 1-4 at strategic points, which were then used for attachment of fluorescent tags. C3 possesses an internal thioester which can be labelled with a fluorophore upon activation to C3b. Intermolecular FRET measurements were thus undertaken to gain information about the interaction between the two proteins that is crucial for understanding functional activity. The CCP modules in the centre of fH may be responsible for introducing a bend into fH that brings the N-teminus close to the C-terminus (the latter is important for host versus non-host discrimination) joined by the longest linkers occurring in the whole molecule. This coincidence of two relatively small CCP modules, 12 and 13, with the highest number of eight amino acids between them, is hypothesised to reflect some unique architectural features. To explore the structural details of this portion of fH by FRET, single-labelled cysteine mutants were further modifed to provide a recognition site for transglutaminase (TGase), which can be enzymatically labeled with a second fluorophore. This stoichiometrically-labelled protein was used for intramolecular FRET studies.

GaN and its alloys with Al and In belong to the group III nitride semiconductors and are today the materials of choice for efficient white light emitting diodes (LEDs) enabling energy saving solid state lighting. Currently, there is a great interest in the development of novel inexpensive techniques to fabricate hybrid LEDs combining high quality III-N quantum well (QW) structures with inexpensive colloidal nanoparticles or conjugated polymers. Such hybrid devices are promising for future micro-light sources in full-color displays, sensors and imaging systems. Organics can be engineered to emit at different wavelengths or even white light based on functional groups or by blend of several polymers. This is especially important for the green region, where there is still a lack of efficient LEDs. Besides optoelectronics, other applications such as biochemical sensors or systems for water splitting can be realized using GaN-based nanostructures. Despite a significant progress in the field, there is still a need in fundamental understanding of many problems and phenomena in III-nitride based nanostructures and hybrids to fully utilize material properties on demand of specific applications. In this thesis, hybrid structures based on AlGaN/GaN QWs and colloidal ZnO nano-crystals have been fabricated for down conversion of the QW emission utilizing non-radiative (Förster) resonant energy transfer. Time-resolved photoluminescence (TRPL) was used to investigate the QW exciton dynamics depending on the cap layer thickness in the bare QW and in the hybrid samples. Although the surface potential influences the exciton dynamics, the maximum pumping efficiency assuming a non-radiative energy transfer mechanism was estimated to be ~40 % at 60 K in the structure with thin cap layer of 3 nm. Since bulk GaN of large area is difficult to synthesize, there is a lack of native substrates. Thus, GaN-based structures are usually grown on SiC or sapphire, which results in high threading dislocation density in the active layer of the device and can be the reason of efficiency droop in GaN based LED structures. Fabricating GaN nanorods (NR) can be a way to produce GaN with lower defect density since threading dislocations can be annihilated toward the NR wall during growth. Here, GaN(0001) NRs grown on Si(111) substrates by magnetron sputtering using a liquid Ga target have been investigated. A high quality of NRs have been confirmed by transmission electron microscopy (TEM) and TRPL. Two strong near band gap emission lines at ~3.42 eV and ~3.47 eV related to basal plane stacking faults (SF) and donor-bound exciton (DBE), respectively, have been observed at low temperatures. TRPL properties of the SF PL line suggest that SFs form a regular structure similar to a multiple QWs, which was confirmed by TEM. The SF related PL measured at 5 K for a single NR has a significantly different polarization response compared to the GaN exciton line and is much stronger polarized (> 40 %) in the direction perpendicular to the NR growth axis. Hybrids fabricated using GaN NRs and the green emitting polyfluorene (F8BT) have been studied using micro-TRPL. In contrast to the DBE emission, the recombination time of the SF-related emission was observed to decrease, which might be due to the Förster resonance energy transfer mechanism. Compared to chemical vapor deposition, sputtering allows synthesis at much lower temperatures. Here, sputtering was employed to grow InAlN/GaN heterostructures with an indium content targeted to ~18 %, which is lattice matched to GaN. This means that near strain-free GaN films can be synthesized. It was found that using a lower temperature (~25 C) while depositing the top InAlN results in an improved interface quality compared to deposition at 700 C. In latter case, regions of quaternary alloy of InAlGaN forming structural micro-defects have been observed at the top InAlN/GaN interface in addition to optically active flower-like defect formations.

University of Macau / Faculty of Arts and Humanities. / Department of English