Rapid creation of devices with microscale features is a vital step in the commercialization of a wide variety of technologies, such as microfluidics, fuel cells and self-healing materials. The current standard for creating many of these microstructured devices utilizes the inexpensive, flexible material poly-dimethylsiloxane (PDMS) to replicate microstructured molds. This process is inexpensive and fast for small batches of devices, but lacks scalability and the ability to produce large surface-area materials. The novel fabrication process presented in this paper uses a cylindrical mold with microscale surface patterns to cure liquid PDMS prepolymer into continuous microstructured films. Results show that this process can create continuous sheets of micropatterned devices at a rate of 1.9 in2/sec (~1200 mm2/sec), almost an order of magnitude faster than soft lithography, while still retaining submicron patterning accuracy.

<p>The analysis of low abundant biological molecules is often challenging due to their chemical properties, low concentration and limited sample volumes. Neuropeptides are one group of molecules that fits these criteria. Neuropeptides also play an important role in biological functions, which makes them extra interesting to analyze. A classic chemical analysis involves sampling, sample preparation, separation and detection. In this thesis, an enhanced solid supported microdialysis method was developed and used as a combined sampling- and preparation technique. In general, significantly increased extraction efficiency was obtained for all studied peptides. To be able to control the small sample volumes and to minimize the loss of neuropeptides because of unwanted adsorption onto surfaces, the subsequent analysis steps were miniaturized to a micro total analysis system (µ-TAS), which allowed sample pre-treatment, injection, separation, manipulation and detection. </p><p>In order to incorporate these analysis functions to a microchip, a novel microfabrication protocol was developed. This method facilitated three-dimensional structures to be fabricated without the need of clean room facilities. </p><p>The sample pre-treatment step was carried out by solid phase extraction from beads packed in the microchip. Femtomole levels of neuropeptides were detected from samples possessing the same properties as microdialysates. The developed injection system made it possible to conduct injections from a liquid chromatographic separation into a capillary electrophoresis channel, which facilitated for advanced multidimensional separations. An electrochemical sample manipulation system was also developed. In the last part, different electrospray emitter tip designs made directly from the edge of the microchip substrate were developed and evaluated. The emitters were proven to be comparable with conventional, capillary based emitters in stability, durability and dynamic flow range. Although additional developments remain, the analysis steps described in this thesis open a door to an integrated, on-line µ-TAS for neuropeptides analysis in complex biological samples.</p>

The analysis of low abundant biological molecules is often challenging due to their chemical properties, low concentration and limited sample volumes. Neuropeptides are one group of molecules that fits these criteria. Neuropeptides also play an important role in biological functions, which makes them extra interesting to analyze. A classic chemical analysis involves sampling, sample preparation, separation and detection. In this thesis, an enhanced solid supported microdialysis method was developed and used as a combined sampling- and preparation technique. In general, significantly increased extraction efficiency was obtained for all studied peptides. To be able to control the small sample volumes and to minimize the loss of neuropeptides because of unwanted adsorption onto surfaces, the subsequent analysis steps were miniaturized to a micro total analysis system (µ-TAS), which allowed sample pre-treatment, injection, separation, manipulation and detection. In order to incorporate these analysis functions to a microchip, a novel microfabrication protocol was developed. This method facilitated three-dimensional structures to be fabricated without the need of clean room facilities. The sample pre-treatment step was carried out by solid phase extraction from beads packed in the microchip. Femtomole levels of neuropeptides were detected from samples possessing the same properties as microdialysates. The developed injection system made it possible to conduct injections from a liquid chromatographic separation into a capillary electrophoresis channel, which facilitated for advanced multidimensional separations. An electrochemical sample manipulation system was also developed. In the last part, different electrospray emitter tip designs made directly from the edge of the microchip substrate were developed and evaluated. The emitters were proven to be comparable with conventional, capillary based emitters in stability, durability and dynamic flow range. Although additional developments remain, the analysis steps described in this thesis open a door to an integrated, on-line µ-TAS for neuropeptides analysis in complex biological samples.

Submitted by Jaqueline Silva (jtas29@gmail.com) on 2016-10-03T14:17:36Z No. of bitstreams: 2 Dissertação - Eunício de Oliveira Lobo Junior - 2016.pdf: 5628245 bytes, checksum: cb5cfdfdb21a0a9bed0292decf6c094f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Jaqueline Silva (jtas29@gmail.com) on 2016-10-03T14:18:22Z (GMT) No. of bitstreams: 2 Dissertação - Eunício de Oliveira Lobo Junior - 2016.pdf: 5628245 bytes, checksum: cb5cfdfdb21a0a9bed0292decf6c094f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2016-10-03T14:18:22Z (GMT). No. of bitstreams: 2 Dissertação - Eunício de Oliveira Lobo Junior - 2016.pdf: 5628245 bytes, checksum: cb5cfdfdb21a0a9bed0292decf6c094f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2016-03-10 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / This report describes the development of two alternative platforms for electrophoretic runs in microsystems. Firstly, the development of a hybrid capillary system that dispenses microfabrication steps is presented using fused silica capillaries interconnected by a commercial crossed shape interface. This hybrid system was coupled with contactless conductivity detector (C4D) to allow the determination of inorganic cations in biological samples. Electrokinetic sample injection was performed through gated mode approach for the first time in this arrangement. Operational parameters such as: (i) wave frequency and amplitude applied in C4D system, (ii) electrical potential applied in injection, (iii) injection time, (iv) detection point, (v) effect of capillary conditioning as well as and (vi) recovery percentage were extensively investigated and optimized. Better separations of cationic mixture containing NH4+, K+, Na+, Ca2+ and Mg2+ were achieved using a buffer system composed of 50 mM Lactic Acid, 20 mM Histidine and 3 mM 18-crown-6 on a capillary with effective length of 14.5 cm. . Addition of internal standard was used to ensure analytical reproducibility and allow the recording of merit figures. Linear behaviors were observed in concentration ranges between 10 and 100 M for NH4+, K+, Ca2+ e Mg2+, and 20-200 M for Na+. The limit of detection values calculated were 3.75 μM (NH4+), 3.70 μM (K+), 7.50 μM (Na+), 5.00 μM (Ca2+) and 5.35 μM (Mg2+). The concentration levels achieved for cations in biological samples ranged from 4,1 μM to 200 μM. Besides the hybrid system, this report also describes the development of an alternative methodology for the fabrication of high-relief masters for soft-lithography in poly(dimethylsiloxane) (PDMS) substrate. One of the innovative features makes reference to the use of low cost commercial photoresist from textile industry - poly(vinyl acetate) (PVAc) - which exhibits low toxicity. PVAc films were deposited on printed cirtuitry boards through the use of a homemade spincoater developed by desktop cooler, with rotation time control. This methodology allowed the production of high relief masters and PDMS channels with width and depth of 50 μm and 40 μm, respectively. Channels and masters profiles They were characterized with the following techniques: scan electron microscopy, perfilometry, optical and electrical. PDMS electrophoresis devices were successfully used for the separation of major inorganic cations. / Esta dissertação descreve o desenvolvimento de duas plataformas alternativas para a realização de eletroforese em microssistemas. Inicialmente é descrita um sistema eletroforético híbrido que dispensa etapas de microfabricação utilizando capilares de sílica fundida, conectados por uma interface comercial com formato em cruz. Este sistema capilar híbrido foi acoplado com detecção condutométrica sem contato (C4D) e aplicado na determinação de cátions inorgânicos (NH4+, K+, Na+, Ca2+, Mg2+) em amostras biológicas. A injeção de amostras foi realizada eletrocineticamente no modo gated, sendo o primeiro estudo em capilares utilizando esta modalidade de injeção. Foram avaliados os parâmetros operacionais de funcionamento do sistema incluindo (i) frequência e amplitude da onda senoidal aplicada ao sistema de detecção, (ii) potencial elétrico aplicado na injeção, (iii) tempo de injeção, (iv) composição do tampão, (v) ponto de detecção, (vi) efeito do condicionamento do capilar e (vii) recuperação. As melhores separações para uma mistura contendo os cátions inorgânicos foram obtidas usando-se um sistema tamponante composto de ácido lático 50 mM, histidina 20 mM e éter coroa (18-crown-6) 3 mM em um capilar com comprimento efetivo de 14,5 cm. As figuras de mérito analítico foram obtidas a partir da adição do íon Li+ como padrão interno, o qual assegurou confiabilidade nas análises quantitativas. A partir da otimização dos parâmetros analíticos, as curvas analíticas para os íons NH4+, K+, Ca2+ e Mg2+ exibiram comportamento linear (R2>0,99) na faixa de 10-100 M enquanto a curva analítica para o íon Na+ proporcionou resposta linear na faixa de 20-200 M. Os limites de detecção encontrados para os cinco cátions foram entre 3,75 μM (NH4+), 3,75 μM (K+), 7,50 μM (Na+), 5,00 μM (Ca2+) e 5,35 μM (Mg2+). O sistema desenvolvido foi explorado para a determinação dos cátions inorgânicos em amostras de urina, saliva e lágrimas. As concentrações encontradas nas amostras biológicas variaram de 4,1 μM a 200 μM. Além do sistema híbrido, a dissertação também apresenta uma metodologia de baixo custo para produção de moldes em alto relevo para litografia suave em poli(dimetilsiloxano) (PDMS). A principal inovação é o uso de fotoresiste de baixo custo, que se trata de uma emulsão fotossensível de poli(acetato de vinila) (PVAc) utilizada na indústria têxtil e que apresenta baixa toxicidade. Outra inovação é o controle da altura dos moldes utilizando um spincoater de produção própria, com controle de tempo de rotação. Com esta metodologia foram produzidos moldes em alto relevo, e microchips em PDMS com 50 μm de largura e 40 μm de altura. Foram realizadas separações eletroforéticas dos cátions NH4+,K+,Na+,Ca2+,Mg2+e Li+. As eficiências de separação variaram entre 73.000 e 120.000 pratos/m. O que comprova que a metodologia alternativa apresenta aplicabilidade microfluídica