Aflatoxin is a potent biological toxin produced by fungi Aspergillus flavus and A. parasiticus. Current quantification methods for aflatoxins are mostly established on immunoaffinity columns which are both costly and labor intensive. Inspired by smectites’ high aflatoxin adsorption capacity and affinity, a novel aflatoxin quantification sensor based on smectite-polyacrylamide (PAM) nanocomposite was fabricated. First, a smectite-PAM nanocomposite film was synthesized on flat silicon substrates which assembled smectite particles from the clay suspension. A layer-by-layer assembly process was developed to achieve uniform morphology and thickness of the nanocomposite films. During the aflatoxin quantification process, positive correlations between the fluorescence intensity from the aflatoxin B1 (AFB1) adsorbed smectite-PAM nanocomposite films and the AFB1 concentration in the test solutions were obtained. The smectite-PAM nanocomposite film has shown similar AFB1 adsorption capabilities as the smectite. Second, the smectite-PAM nanocomposite film was optimized in order to achieve the aflatoxin quantification at ppb level (below 20ppb) in corn extraction solutions. The smectite was modified by Ba2+, which had demonstrated to be able to improve its aflatoxin adsorption capacity. PAM aqueous solutions with the mass concentration ranging from 0.8% to 0.001% were tested. The results showed that the nanocomposite synthesized from 0.005% concentration of PAM solution generated the best properties. After the optimization, the smectite-PAM nanocomposite films achieved the detection of aflatoxin B1, B2, G1 and G2 (AFB2, AFG1 and AFG2) in 10 ppb corn extraction solution. Aflatoxin quantifications in AFB1 and AFB2 mixture solution, AFB1 and AFB2 mixture solution and AFB1 and AFG1 mixture solution were conducted, and the recoveries of last test ranged from 90.52% to 110.11% at low aflatoxin concentration (below 20 ppb). Third, in order to shorten the quantification duration and simplify the detection process, a novel aflatoxin detection array based on smectite-PAM nanocomposite and an improved fluorometric quantification method were developed. Through a microfluidic chip, the reaction time was reduced to 10~20min. Two concentration levels (20~80ppb/5~15ppb) of aflatoxin B1 spiked corn extraction solutions were tested. In the fluorometric quantification step, a common lab-use 365 nm ultraviolet lamp replaced the spectrofluorometer which simplified and accelerated the process.

The lymphatic system plays an important role in fluid and protein balance within the interstitial spaces. Its dysfunction could result in a number of debilitating diseases, namely lymphedema. Lymphatic vessels utilize both intrinsic and extrinsic mechanisms to pump lymph. Intrinsic pumping involves the active contraction of vessels, a phenomenon that is regulated in part by nitric oxide (NO) produced by lymphatic endothelial cells (LECs). NO production by arterial endothelial cells has been shown to be sensitive to both shear stress and stretch. Therefore, because of the unique mechanical environment of the LECs, we hypothesize that mechanical forces play an important role in regulation of the lymphatic pumping. Parallel-plate flow chambers and indenter-based cyclic stretch devices were constructed and used to apply mechanical loads to LECs. In addition, high-throughput micro-scale channels were developed and tested for shear experiments to address the need to increase the productivity and high- resolution imaging. Twenty-four hours treatment of LECs with different shear stress conditions showed a shear-dependent elevation in NO production. Moreover, 2.5 folds increase in cumulative NO was observed for stretched cells compared to the unstretched cells over six hours period. In conclusion, the upregulation observed in NO production under mechanical stimuli suggest new regulatory mechanisms that can be pharmaceutically targeted. These results provide an unprecedented insight into lymphatic pumping mechanism.

A hybrid molecular dynamics/multiparticle collision dynamics algorithm was constructed to model the pressure-driven flow of a compressible fluid through a nanoscopic channel of square cross-sectional area, as well as the effect of this flow on the configuration of a polymer chain that was tethered to the surface of this nanochannel. In the process of simulating channel flow, a new adiabatic partial slip boundary condition was created as well as a modified source/sink inlet and outlet boundary condition that could maintain a specified pressure gradient across the channel without the large entrance effects typically associated with these algorithms. The results of the flow simulations were contrasted with the results from a series solution to the Navier-Stokes equation for isothermal compressible flow, and showed excellent agreement with the results from the series solution when slip-boundary conditions were applied. A finitely extendible non-linear elastic spring and bead polymer chain was used to simulate the effect of flow on the polymer chain configuration under poor solvent and θ solvent conditions. Under θ solvent conditions, the cyclical dynamics that have been previousy observed for tethered polymer chains in pure shear flows were noted, however they were restricted to the end of the polymer chain. Under poor solvent conditions, the polymer adopted a metastable helix configuration as it collapsed to a globule state. The study also examined interchain and intrachain entanglements in polymers using the granny knot and overhand knot. The mechanisms by which these tangles untied themselves were determined. At low flow rates, the tangles unravelled by the end of the chain migrating through the loops of the tangle. At high flow rates, the tangles behaved like an entrained object as they reptated towards the end of the chain.

Isolation of selected populations from heterogeneous cell mixtures and retrieval of the captured population of interest for regenerative medicine and diagnostics applications is one of the challenges that may be addressed by microfluidics. An affinity adhesion strategy was tested using the tetrapeptides RGDS (arg-gly-asp-ser), REDV (arg-glu-asp-val) and VAPG (val-ala-pro-gly) to modify an alginate hydrogel surface layer to selectively adhere fibroblast (FB), endothelial (EC) and smooth muscle cell (SMC) populations, respectively, of the non-myocyte cardiac cell fraction. Incorporation of peptides into sodium alginate gel surface coatings demonstrated a preferential, seeding density-dependent adhesion relationship on alginate-RGDS when tested with a cardiomyocyte-depleted cell suspension in both static culture and in microfluidic devices. Seeding density-dependent attachment was seen with close to 100% release of viable cells from coated surfaces upon application of ethylenediaminetetraacetic acid (EDTA). Further work will optimize the system with REDV and VAPG to capture ECs and SMCs.

Digital microfluidics is a new technology that permits manipulation of liquid droplets on an array of electrodes. Using this technology, nanoliter to microliter size droplets of different samples and reagents can be dispensed from reservoirs, moved, split, and merged together. Digital microfluidics is poised to become an important and useful tool for biomedical applications because of its capacity to precisely and automatically carry out sequential chemical reactions. In this thesis, a set of tools is presented to accelerate the integration of digital microfluidics into Lab-on-a-Chip platforms for a wide range of applications. An important contribution in this thesis is the development of three rapid prototyping techniques, including the use of laser printing to pattern flexible printed circuit board (PCB) substrates, to make the technology accessible and less expensive. Using these techniques, both digital and channel microfluidic devices can be produced in less than 30 minutes at a minimal cost. These rapid prototyping techniques led to a new method for manipulating liquid droplets on non-planar surfaces. The method, called All Terrain Droplet Actuation (ATDA), was used for several applications, including DNA enrichment by liquid-liquid extraction. ATDA has great potential for the integration of different physico-chemical environments on Lab-on-a-Chip devices. A second important contribution described herein is the development of a new microfluidic format, hybrid microfluidics, which combines digital and channel microfluidics on the same platform. The new hybrid device architecture was used to perform biological sample processing (e.g. enzymatic digestion and fluorescent labeling) followed by electrophoretic separation of the analytes. This new format will facilitate complete automation of Lab-on-a-Chip devices and will eliminate the need for extensive manual sample processing (e.g. pipetting) or expensive robotic stations. Finally, numerical modeling of droplet actuation on single-plate digital microfluidic devices, using electrodynamics, was used to evaluate the droplet actuation forces. Modeling results were verified experimentally using an innovative technique that estimates actuation forces based on resistive forces against droplet motion. The results suggested a list of design tips to produce better devices. It is hoped that the work presented in this thesis will help introduce digital microfluidics to many of the existing Lab-on-a-Chip applications and inspire the development of new ones.

A hybrid molecular dynamics/multiparticle collision dynamics algorithm was constructed to model the pressure-driven flow of a compressible fluid through a nanoscopic channel of square cross-sectional area, as well as the effect of this flow on the configuration of a polymer chain that was tethered to the surface of this nanochannel. In the process of simulating channel flow, a new adiabatic partial slip boundary condition was created as well as a modified source/sink inlet and outlet boundary condition that could maintain a specified pressure gradient across the channel without the large entrance effects typically associated with these algorithms. The results of the flow simulations were contrasted with the results from a series solution to the Navier-Stokes equation for isothermal compressible flow, and showed excellent agreement with the results from the series solution when slip-boundary conditions were applied. A finitely extendible non-linear elastic spring and bead polymer chain was used to simulate the effect of flow on the polymer chain configuration under poor solvent and θ solvent conditions. Under θ solvent conditions, the cyclical dynamics that have been previousy observed for tethered polymer chains in pure shear flows were noted, however they were restricted to the end of the polymer chain. Under poor solvent conditions, the polymer adopted a metastable helix configuration as it collapsed to a globule state. The study also examined interchain and intrachain entanglements in polymers using the granny knot and overhand knot. The mechanisms by which these tangles untied themselves were determined. At low flow rates, the tangles unravelled by the end of the chain migrating through the loops of the tangle. At high flow rates, the tangles behaved like an entrained object as they reptated towards the end of the chain.

Los sistemas de microfluídica conocidos con el nombre de lab-on-a-chip han contribuido al avance en áreas como la química, la biología, la biomedicina, la biodefensa y la microelectrónica, entre otras. Sin embargo, a pesar de sus beneficios, su comercialización está principalmente limitada por los altos costes de producción derivados de las técnicas utilizadas para su fabricación. Es por ello que actualmente se están centrando esfuerzos en utilizar técnicas que disminuyan los costes pero que conserven o incrementen los beneficios de estos dispositivos. Este trabajo de investigación presenta una nueva modalidad de escritura directa con láser (LDW) como una prominente técnica de fabricación, que puede beneficiar el desarrollo de nuevos dispositivos de microfluídica en 3 dimensiones: la escritura directa con láser en 3 dimensiones (LDW-3D). En ella se combinan los beneficios de la técnica de la LDW y una clase de vidrios conocidos con el nombre de vidrios cerámicos fotoestructurables (PSGCs). Con esta técnica se pueden obtener motivos tridimensionales focalizando un haz láser en el interior de un material, el cual debe ser transparente a la radiación que emite dicho láser. La técnica se basa en el efecto de la modificación estructural de un vidrio PSGC conocido con el nombre comercial de Foturan® tras la irradiación con un láser. Luego éste se somete a un tratamiento térmico y finalmente a un ataque químico con HF. Se realizaron ensayos irradiando muestras de Foturan con un láser UV con una duración de pulso de 10 ns y una longitud de onda de 355 nm (nsUV) y un láser de femtosegundos IR con una duración de pulso de 450 fs y una longitud de onda de 1027 nm (fsIR). Los estudios realizados revelan que el láser de nsUV produce un efecto de fotoionización en el Foturan a partir de la absorción de 2 fotones que sensibilizan al agente fotoionizador (Ce+3) contenido en su matriz. En el caso de utilizar el láser de fsIR, la fotoionización de este tipo de vidrio se produce mediante la absorción multifotónica de 8 fotones, actuando sobre niveles de energías que están asociados a defectos e impurezas localizados en la banda prohibida del Foturan. También se han evaluado los diferentes parámetros tecnológicos que permiten la aplicación de la técnica LDW-3D para la fabricación de microestructuras en 3D tales como microcanales y microdepósitos con buen aspecto y correspondencia entre sus dimensiones. En base a estos estudios se fabricó un sistema tridimensional en Foturan utilizando el láser fsIR. Este sistema presenta características que hacen que pueda ser aprovechado en el campo de la microfluídica como micromezclador de soluciones o analizador de microorganismos. / Microfluidic systems known as lab-on-a-chip have contributed to progress in areas such as chemistry, biology, biomedical, biodefense and microelectronics, among others. However, despite their benefits, their marketing is mainly limited by the high production costs resulting from the techniques involved in their manufacture. This is why nowadays researchers are focusing efforts in new techniques to reduce costs but maintaining or increasing the benefits of these devices.This research presents a new type of laser direct writing (LDW) as a prominent fabrication technique, which can benefit the development of new microfluidic devices in three dimensions: laser direct write in 3 dimensions (LDW-3D). It combines the benefits of the technique LDW and a type of glasses known as photo-structural glass ceramics (PSGCs). 3D structures can be obtained through this technique by focusing a laser beam inside a material, which has to be transparent to the radiation emitted by the laser. The technique is based on the effect of structural modification of a PSGC glass called Foturan® after laser irradiation. Afterwards, the glass is submitted to a standard thermal treatment and finally, to an etching step with HF. Tests were performed by irradiating Foturan samples with an UV nanosecond laser with 10 ns pulse duration and 355 nm wavelength (nsUV), and an IR femtosecond laser with 450 fs pulse duration and 1027 nm wavelength (fsIR).The results show that nsUV laser produces the photoionization effect on the Foturan through a 2-photon absorption process which sensitizes the photoactive agent (Ce+3). In the case of using the fsIR laser, photoionization is produced by a 8-photon multiphoton absorption process, involving energy levels that are associated with localized defects and impurities in the Foturan gap. Different technological parameters have been evaluated in order to study the viability of the LDW-3D technique for producing 3D microstructures, such as microchannels and microreservoirs showing good aspect-ratio. Based on these studies, a 3D-system in Foturan was produced by using the fsIR laser. This system presents special features that make it suitable in the field of microfluidics as micromixer or microorganisms analyzer.

Electrophoresis separation corresponds to the motion and separation of dispersed particles under the influence of a constant electric field. In molecular biology, electrophoresis separation plays a major role in identifying, quantifying and studying different biological samples such as proteins, peptides, RNA acids, and DNA. In electrophoresis separation, different characteristics of particles, such as charge to mass ratio, size, and pI, can be used to separate and isolate those particles. For very complex samples, two or more characteristics can be combined to form a multi-dimensional electrophoresis separation system, significantly improving separation efficiency. Much effort has been devoted in recent years to performing electrophoresis separations in microfluidic format. Employing microfluidic technology for this purpose provides several benefits, such as improved transport control, reduced sample volumes, simplicity of operation, portability, greater accessibility, and reduced cost. The aim of this study is to develop microfluidic systems for high-performance separation of biochemical samples using electrophoresis methods. The first part of the thesis concerns the development of a fully integrated microfluidic chip for isoelectric focusing separation of proteins with whole-channel imaging detection. All the challenges posed in fabricating and integrating the chip were addressed. The chip was tested by performing protein and pI marker separations, and the separation results obtained from the chip were compared with those obtained from commercial cartridges. Side-by-side comparison of the results validated the developed chip and fabrication techniques. The research also focuses on improving the peak capacity and separation resolution of two counter-flow gradient electrofocusing methods: Temperature Gradient Focusing (TGF) and Micellar Affinity Gradient Focusing (MAGF). In these techniques, a temperature gradient across a microchannel or capillary is used to separate analytes. With an appropriate buffer, the temperature gradient creates a gradient in the electrophoretic velocity (TGF) or affinity (MAGF) of analytes and, if combined with a bulk counter-flow, ionic species concentrate at unique points where their total velocity is zero, and separate from each other. A bilinear temperature gradient is used along the separation channel to improve both peak capacity and separation resolution simultaneously. The temperature profile along the channel consists of a very sharp gradient used to pre-concentrate the sample, followed by a shallow gradient that increases separation resolution. A simple numerical model was applied to predict the improvement in resolution when using a bilinear gradient. A hybrid PDMS/glass chip integrated with planar micro-heaters for generating bilinear temperature gradients was fabricated using conventional sputtering and soft lithography techniques. A specialized design was developed for the heaters to achieve the desired bilinear profiles using both analytical and numerical modeling. To confirm the temperature profile along the channel, a two-color thermometry technique was also developed for measuring the temperature inside the chip. Separation performance was characterized by separating several different dyes, amino acids and peptides. Experiments showed a dramatic improvement in peak capacity and resolution of both techniques over the standard linear temperature gradients. Next, an analytical model was developed to investigate the effect of bilinear gradients in counter-flow gradient electrofocusing methods. The model provides a general equation for calculating the resolution for different gradients, diffusion coefficients and bulk flow scan rates. The results indicate that a bilinear gradient provides up to 100% improvement in separation resolution over the linear case. Additionally, for some scanning rates, an optimum bilinear gradient can be found that maximizes separation resolution. Numerical modeling was also developed to validate some of the results. The final part of the thesis describes the development of a two-dimensional separation system for protein separation, combining temperature gradient focusing (TGF) and sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis (PAGE) in a PDMS/glass microfluidic chip. An experimental study was performed on separating a mixture of proteins using two characteristics: charge to mass ratio, and size. Experimental results showed a dramatic improvement in peak capacity over each of the one-dimensional separation techniques.

Micro-channels are believed to open up the prospect of precise control of fluid flow and chemical reactions. The capillary effect can be used to pump fluids in micro-channels and the flow generated can dissolve chemicals previously deposited on the walls of the channel. In this work, numerical and experimental approaches have been developed to investigate the wall mass transfer rate generated by capillary driven flows (CD-Flow). The purpose of this work is to analyze the wall mass transfer rates generated by a CD-Flow in a micro-channel. The results have implications in the optimization and design of devices for biological assays. The correlation for Sherwood number, Reynolds number, contact angle and time is reported. This correlation can be a useful tool for design purposes of microfluidic devices that work with fast heterogeneous reaction and have capillary driven flow as passive pumping system. The numerical results have been confirmed by the experimental results. / La perspectiva del uso de micro-canales para el control preciso del flujo y de las reacciones químicas está ampliamente aceptada. Considerando que el efecto de las tensiones superficiales en la micro-escala es significativo, el bombeo pasivo basado en el uso de la tensión superficial para los Lab-on-a-chip resulta ser el método más eficaz.El propósito de este trabajo es analizar la transferencia de masa en la pared en un campo dinámico de un flujo impulsado por capilaridad. Los resultados permitirán mejorar el diseño y optimizar los dispositivos para ensayos biológicos. Se presenta una correlación entre el número de Sherwood, el número de Reynolds, el ángulo de contacto y el tiempo. La correlación puede ser una herramienta útil en el diseño de dispositivos microfluídicos que trabajen con una reacción rápida y heterogénea y usen el bombeo pasivo impulsado por el flujo capilar. Los resultados numéricos han sido confirmados por los resultados experimentales.

<p>A digital microfluidic biochip (DMFB) is a device that digitizes fluidic samples into tiny droplets and operates chemical processes on a single chip. Movement control of droplets can be realized by using electrowetting-on-dielectric (EWOD) technology. DMFBs have high configurability, high sensitivity, low cost and reduced human error as well as a promising future in the applications of point-of-care medical diagnostic, and DNA sequencing. As the demands of scalability, configurability and portability increase, a new DMFB architecture called Microelectrode Dot Array (MEDA) has been introduced recently to allow configurable electrodes shape and more precise control of droplets.</p> <p>The objective of this work is to investigate a routing algorithm which can not only handle the routing problem for traditional DMFBs, but also be able to route different sizes of droplets and incorporate diagonal movements for MEDA. The proposed droplet routing algorithm is based on 3D-A* search algorithm. The simulation results show that the proposed algorithm can reduce the maximum latest arrival time, average latest arrival time and total number of used cells. By enabling channel-based routing in MEDA, the equivalent total number of used cells can be significantly reduced. Compared to all existing algorithms, the proposed algorithm can achieve so far the least average latest arrival time.</p>