Electrowetting on dielectric (EWOD) is a technique for reducing the apparent contact angle of a fluid droplet, which has many promising applications in the fields of optics, digital displays, and lab-on-a-chip research. In this thesis, a design is presented for a novel single circuit device for achieving continuous droplet motion, by using the current-rectifying properties of valve metals to create diode-like behavior. This contrasts with existing designs, which require an array of individual electrodes to achieve motion in discrete steps. We are able to demonstrate continuous droplet motion across a 28mm-long test strip with an applied voltage of 303 V and a velocity of 5.59 mm/s (at 370 V) using an ionic-fluid electrolyte (BMIM-PF6), and have achieved actuation at as low as 185 V, with a maximum observed velocity (at 300 V) of 13.8 mm/s using a 1M sodium sulfate solution.

Surgery of axons in C. elegans using ultrafast laser pulses, and observing their subsequent regrowth opens a new frontier in neuroscience, since such research holds a great potential for the development of novel therapies and cures to neurodegenerative diseases. In order to make the required large-scale genetic screenings in C. elegans possible and thus obtain statistically significant biological data, an automated laser axotomy system needs to be developed. Microfluidic devices hold the promise of improved throughput by integrating different functional modules into a single chip. The first step to developing a microfluidic device for laser axotomy is to devise an on-chip worm trapping method, which maintains a high degree of immobilization to sever axons without using anesthetics. In this thesis, we present a novel method that uses a thin, deflectable PDMS membrane that individually traps worms in a microfluidic device. Axons can successfully be severed with the same accuracy as those using conventional paralyzing techniques. This device also incorporates recovery chambers for housing worms after surgery and for time-lapse imaging of axonal regrowth without the repeated use of anesthetics. Towards accomplishing an automated, high-throughput laser axotomy system, we developed an improved microfluidic design based on the same mechanical immobilization technique. This second generation device allows for serially processing of a large quantity of worms rapidly using a semi-automated system. Integrated to the opto-mechanical platform, a software program utilizing image processing techniques is developed. This semi-automated program can automatically identify the location of worms, their neuronal cell bodies, focus on the axons of interest, and align the laser beam with the axon via a PID based viso-servo feedback algorithm. Statistic data demonstrate that there is no significant difference in axonal reconnection rates between surgeries performed on-chip and using anesthetics. To improve flow control, a three-dimensional novel microfluidic valve structure is designed and fabricated. This novel valve structure allows for a complete sealing of the flow channel, without degrading optical conditions for imaging and laser ablation in the trapping area. Finally, we developed a prototypical microfluidic assembly that will eventually be able to interface a well-plate to automatically deliver population of worms from individual wells to the automated chip for axotomy. This interface consists of a microfluidic multiplexer to significantly reduce the number of solenoid valves needed to individually address each well. / text

This dissertation develops biosensors for rapid detection of pathogens for environmental monitoring and food safety applications and utilizes the multidisciplinary and multi-application characteristics of biosensors to develop a lesson plan that can be implemented in secondary education classrooms. The detection methods evolve from particle immunoagglutination assay, PDMS optofluidic lab-on-a-chip, and spectrum analysis to smartphone and image analysis without any reagent; the potential application in secondary education also underlines the extended value of biosensors. In the first paper presented here, an optofluidic lab-on-a-chip system and subsequent sampling procedure were developed for detecting bacteria from soil samples utilizing Mie scattering detection of particle immunoagglutination assay. This system and protocol detected the presence of Escherichia coli K12 from soil particles in near real-time (10 min) with a detection limit down to 1 CFU mL⁻¹ and has the potential to be implemented in the field. We also compared the interaction between E. coli and soil particles to the two-step protein-surface interaction. In the second paper, a smartphone-utilized biosensor consisting of a near-infrared (NIR) LED (wavelength of 880 nm) and a digital camera of a smartphone was developed for detecting microbial spoilage on ground beef, without using any reagents. The method was further improved by programming a smartphone application that allows the user to position the smartphone at an optimum distance and a range of angles utilizing its internal gyro sensor to measure a series of scatter intensities against the detection angle. This handheld device can be used as a preliminary screening tool to monitor microbial contamination on meat products. In the third paper, we designed a lesson plan for secondary education classrooms using biosensors as a core and branching out to different applications and fields of study with the goal of heightening students' interest and motivation toward attaining degrees and careers in STEM fields. Results revealed that the lesson was more effective in affecting younger students than older students, and more effective in teaching about the applications of biosensors than about the techniques of biosensor development.

This dissertation develops biosensors for rapid detection of pathogens for environmental monitoring and food safety applications and utilizes the multidisciplinary and multi-application characteristics of biosensors to develop a lesson plan that can be implemented in secondary education classrooms. The detection methods evolve from particle immunoagglutination assay, PDMS optofluidic lab-on-a-chip, and spectrum analysis to smartphone and image analysis without any reagent; the potential application in secondary education also underlines the extended value of biosensors. In the first paper presented here, an optofluidic lab-on-a-chip system and subsequent sampling procedure were developed for detecting bacteria from soil samples utilizing Mie scattering detection of particle immunoagglutination assay. This system and protocol detected the presence of Escherichia coli K12 from soil particles in near real-time (10 min) with a detection limit down to 1 CFU mL⁻¹ and has the potential to be implemented in the field. We also compared the interaction between E. coli and soil particles to the two-step protein-surface interaction. In the second paper, a smartphone-utilized biosensor consisting of a near-infrared (NIR) LED (wavelength of 880 nm) and a digital camera of a smartphone was developed for detecting microbial spoilage on ground beef, without using any reagents. The method was further improved by programming a smartphone application that allows the user to position the smartphone at an optimum distance and a range of angles utilizing its internal gyro sensor to measure a series of scatter intensities against the detection angle. This handheld device can be used as a preliminary screening tool to monitor microbial contamination on meat products. In the third paper, we designed a lesson plan for secondary education classrooms using biosensors as a core and branching out to different applications and fields of study with the goal of heightening students' interest and motivation toward attaining degrees and careers in STEM fields. Results revealed that the lesson was more effective in affecting younger students than older students, and more effective in teaching about the applications of biosensors than about the techniques of biosensor development.

Digital microfluidics (DMF) has recently emerged as a popular technology for a wide range of applications. In DMF, nL-mL droplets containing samples and reagents are controlled(i.e., moved, merged, mixed, and dispensed from reservoirs) by applying a series of electrical potentials to an array of electrodes coated with a hydrophobic insulator. DMF is distinct from microchannel-based fluidics as it allows for precise control over multiple reagent phases (liquid and solid) in heterogeneous systems with no need for complex networks of microvalves. In this thesis, digital microfluidics has been applied to address key challenges in the fields of chemistry, biology and medicine. For applications in chemistry, the first two-plate digital microfluidic platform for synchronized chemical synthesis is reported. The new method, which was applied to synthesizing peptide macrocycles, is fast and amenable to automation, and is convenient for parallel scale fluid handling in a straightforward manner. For applications in biology, I present the first DMF-based method for extraction of proteins (via precipitation) in serum and cell lysate. The performance of the new method was comparable to that of conventional techniques, with the advantages of automation and reduced analysis time. The results suggest great potential for digital microfluidics for proteomic biomarker discovery. Furthermore, I integrated DMF with microchannels for in-line biological sample processing and separations. Finally, for applications in medicine, I developed the first microfluidic method for sample clean-up and extraction of estrogen from one-microliter droplets of breast tissue homogenates, blood, and serum. The new method is fast and automated, and features >1000x reduction in sample use relative to conventional techniques. This method has significant potential for applications in endocrinology and breast cancer risk reduction. In addition, I describe a new microfluidic system incorporating a digital microfluidic platform for on-chip blood spotting and processing, and a microchannel emitter for direct analysis by mass spectrometry. The new method is fast, robust, precise, and is capable of quantifying analytes associated with common congenital disorders such as homocystinuria, phenylketonuria, and tyrosinemia.

Digital microfluidics (DMF) has recently emerged as a popular technology for a wide range of applications. In DMF, nL-mL droplets containing samples and reagents are controlled(i.e., moved, merged, mixed, and dispensed from reservoirs) by applying a series of electrical potentials to an array of electrodes coated with a hydrophobic insulator. DMF is distinct from microchannel-based fluidics as it allows for precise control over multiple reagent phases (liquid and solid) in heterogeneous systems with no need for complex networks of microvalves. In this thesis, digital microfluidics has been applied to address key challenges in the fields of chemistry, biology and medicine. For applications in chemistry, the first two-plate digital microfluidic platform for synchronized chemical synthesis is reported. The new method, which was applied to synthesizing peptide macrocycles, is fast and amenable to automation, and is convenient for parallel scale fluid handling in a straightforward manner. For applications in biology, I present the first DMF-based method for extraction of proteins (via precipitation) in serum and cell lysate. The performance of the new method was comparable to that of conventional techniques, with the advantages of automation and reduced analysis time. The results suggest great potential for digital microfluidics for proteomic biomarker discovery. Furthermore, I integrated DMF with microchannels for in-line biological sample processing and separations. Finally, for applications in medicine, I developed the first microfluidic method for sample clean-up and extraction of estrogen from one-microliter droplets of breast tissue homogenates, blood, and serum. The new method is fast and automated, and features >1000x reduction in sample use relative to conventional techniques. This method has significant potential for applications in endocrinology and breast cancer risk reduction. In addition, I describe a new microfluidic system incorporating a digital microfluidic platform for on-chip blood spotting and processing, and a microchannel emitter for direct analysis by mass spectrometry. The new method is fast, robust, precise, and is capable of quantifying analytes associated with common congenital disorders such as homocystinuria, phenylketonuria, and tyrosinemia.

The recent breakthroughs in genomics and molecular diagnostics will not be reflected in health-care systems unless the biogenetic or other nucleic acid-based tests are transferred from the laboratory to clinical market. Developments in microfabrication techniques brought lab-on-a-chip (LOC) into being the best candidate for conducting sample preparation for such clinical devices, or point-of-care testing set-ups. Sample preparation procedure consists of several stages including cell transportation, separation, cell lysis and nucleic acid purification and detection. LOC, as a subset of Microelectromechanical systems (MEMS), refers to a tiny, compact, portable, automated and easy-to-use microchip capable of performing the sample-preparation stages together. Complexity in micro-fabrications and inconsistency of the stages oppose integration of them into one chip. Among the variety of mechanisms utilized in LOC for cell lysis, electrical methods have the highest potential to be integrated with other microchip-based mechanisms. There are, however, major limitations in electrical cell lysis methods: the difficulty and high-cost fabrication of microfluidic chips and the high voltage requirements for cell lysis. Addressing these limitations, the focus of this thesis is on realization of cell lysis microchips suitable for LOC applications. We have developed a new methodology of fabricating microfluidic chips with electrical functionality. Traditional lithography of microchannel with electrode, needed for making electro-microfluidic chips, is considerably complicated. We have combined several easy-to-implement techniques to realize electro-microchannel with laser-ablated polyimide. The current techniques for etching polyimide are by excimer lasers in bulky set-ups and with involvement of toxic gas. We present a method of ablating microfluidic channels in polyimide using a 30W CO2 laser. Although this technique has poorer resolution, this approach is more cost effective, safer and easier to handle. We have verified the performance of the fabricated electro-microfluidic chips on electroporation of mammalian cells. Electrical cell lysis mechanisms need an operational voltage that is relatively high compared to other cell manipulation techniques, especially for lysing bacteria. Microelectro-devices have dealt with this limitation mostly by reducing the inter-distance of electrodes. The technique has been realized in tiny flow-through microchips with built-in electrodes in a distance of a few micrometers which is in the scale of cell size. In addition to the low throughput of such devices, high probability of blocking cells in such tiny channels is a serious challenge. We have developed a cell lysis device featured with aligned carbon nanotube (CNT) to reduce the high voltage requirement and to improve the throughput. The vertically aligned CNT on an electrode inside a MEMS device provides highly strengthened electric field near the tip. The concept of strengthened electric field by means of CNT has been applied in field electron emission but not in cell lysis. The results show that the incorporation of CNT in lysing bacteria reduces the required operational voltage and improves throughput. This achievement is a significant progress toward integration of cell lysis in a low-voltage, high-throughput LOC. We further developed the proposed fabrication methodology of micro-electro-fluidic chips, described earlier, to perform electroporation of single mammalian cell. We have advanced the method of embedding CNT in microchannel so that on-chip fluorescent microscopy is also feasible. The results verify the enhancement of electroporation by incorporating CNT into electrical cell lysis. In addition, a novel methodology of making CNT-embedded microfluidic devices has been presented. The embedding methodology is an opening toward fabrication of a CNT-featured LOC for other applications.

No description available.

Biological/biochemistry analyses traditionally require bulky instruments and a great amount of volume of biological/chemical agents, and many procedures have to be performed in certain locations such as medical centers or research institutions. These limitations usually include time delay in testing. The delays may be critical for some aspects such as disease prevention or patient treatment. One solution to this issue is the realization of point-of-care (POC) testings for patients, a domain in public health, meaning that health cares are provided near the sites of patients using well-designed and portable medical devices. Transportation of samples between local and central institutions can therefore be reduced, facilitating early and fast diagnosis. A closely related topic in engineering, lab-on-a-chip (LOC), has been discussed and practiced in recent years. LOC emphasizes integrating several functions of laboratory processes in a small portable device and performing analysis using only a very small amount of sample volume, to achieve low-cost and rapid analysis. From an engineer's point of view, LOC is the strategy to practice the idea of POC testing. This dissertation aimed at exploring the POC potentials of porous membrane-base LOC devices, which can be used to simplify traditional and standard laboratory procedures. In this study, three LOC prototypes are shown and discussed. First the protein sensor incorporating with silica nanofiber membrane, which has shown 32 times more improvement of sensitivity than a conventional technique and a much shorter detection time; secondly the bacteria filter chip that uses a sandwiched aluminum oxide membrane to stabilize the bacteria and monitor the efficacy of antibiotics, which has reduced the test time from 1 day of the traditional methods to 1 hour; the third is the sensor combining microfluidics and silica nanofiber membrane to realize Surface Enhanced Raman Spectroscopy on bio-molecules, which has enhancement factor 10^9 and detection limit down to nanomolar, but simple manufacturing procedures and reduced fabrication cost. These results show the porous-base membrane LOC devices may have potentials in improving and replacing traditional detection methods and eventually be used in POC applications.

abstract: In this thesis, I present a lab-on-a-chip (LOC) that can separate and detect Escherichia Coli (E. coli) in simulated urine samples for Urinary Tract Infection (UTI) diagnosis. The LOC consists of two (concentration and sensing) chambers connected in series and an integrated impedance detector. The two-chamber approach is designed to reduce the non-specific absorption of proteins, e.g. albumin, that potentially co-exist with E. coli in urine. I directly separate E. coli K-12 from a urine cocktail in a concentration chamber containing micro-sized magnetic beads (5 µm in diameter) conjugated with anti-E. coli antibodies. The immobilized E. coli are transferred to a sensing chamber for the impedance measurement. The measurement at the concentration chamber suffers from non-specific absorption of albumin on the gold electrode, which may lead to a false positive response. By contrast, the measured impedance at the sensing chamber shows ~60 kÙ impedance change between 6.4x104 and 6.4x105 CFU/mL, covering the threshold of UTI (105 CFU/mL). The sensitivity of the LOC for detecting E. coli is characterized to be at least 3.4x104 CFU/mL. I also characterized the LOC for different age groups and white blood cell spiked samples. These preliminary data show promising potential for application in portable LOC devices for UTI detection. / Dissertation/Thesis / M.S. Electrical Engineering 2011