144 research outputs found

    Cell Trapping Utilizing Insulator-based Dielectrophoresis in The Open-Top Microchannels

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    The ability to manipulate or separate a biological small particle, such as a living cell and embryo, is fundamental needed to many biological and medical applications. The insulator-based dielectrophoresis (iDEP) trapping is composed of conductless tetragon structures in micro-chip. In this study, a lower conductive material of photoresist was adopted as a structure in open-top microchannel instead of a metallic wire to squeeze the electric field in a conducting solution, therefore, creating a high field gradient with a local maximum. The microchip with the open-top microchannels was designed and fabricated herein. The insulator-based DEP trapping microchip with the open-top microchannels was designed and fabricated in this work. The cells trapped by DEP force could be further treated or cultured in the open-top microchannel ; however, those trapped in the microchip with enclosed microchannels could not be proceeded easily

    Higher Order Electrokinetic Effects for Applied Biological Analytics

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    abstract: Microfluidic systems have gained popularity in the last two decades for their potential applications in manipulating micro- and nano- particulates of interest. Several different microfluidics devices have been built capable of rapidly probing, sorting, and trapping analytes of interest. Microfluidics can be combined with separation science to address challenges of obtaining a concentrated and pure distinct analyte from mixtures of increasingly similar entities. Many of these techniques have been developed to assess biological analytes of interest; one of which is dielectrophoresis (DEP), a force which acts on polarizable analytes in the presence of a non-uniform electric fields. This method can achieve high resolution separations with the unique attribute of concentrating, rather than diluting, analytes upon separation. Studies utilizing DEP have manipulated a wide range of analytes including various cell types, proteins, DNA, and viruses. These analytes range from approximately 50 nm to 1 µm in size. Many of the currently-utilized techniques for assessing these analytes are time intensive, cost prohibitive, and require specialized equipment and technical skills. The work presented in this dissertation focuses on developing and utilizing insulator-based dielectrophoresis (iDEP) to probe a wide range of analytes; where the intrinsic properties of an analyte will determine its behavior in a microchannel. This is based on the analyte’s interactions with the electrokinetic and dielectrophoretic forces present. Novel applications of this technique to probe the biophysical difference(s) between serovars of the foodborne pathogen, Listeria monocytogenes, and surface modified Escherichia coli, are investigated. Both of these applications demonstrate the capabilities of iDEP to achieve high resolution separations and probe slight changes in the biophysical properties of an analyte of interest. To improve upon existing iDEP strategies a novel insulator design which streamlines analytes in an iDEP device while still achieving the desirable forces for separation is developed, fabricated, and tested. Finally, pioneering work to develop an iDEP device capable of manipulating larger analytes, which range in size 10-250 µm, is presented.Dissertation/ThesisDoctoral Dissertation Chemistry 201

    Separation and concentration of water-borne contaminants utilizing insulator-based dielectrophoresis.

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    Dielectrophoretic characterization of particles and erythrocytes

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    Medical lab work, such as blood testing, will one day be near instantaneous and inexpensive via capabilities enabled by the fast growing world of microtechnology. In this research study, sorting and separation of different ABO blood types have been investigated by applying alternating and direct electric fields using class=SpellE\u3edielectrophoresis in microdevices. Poly(dimethylsiloxane) (PDMS) microdevices, fabricated by standard photolithography techniques have been used. Embedded perpendicular platinum (Pt) electrodes to generate forces in AC dielectrophoresis were used to successfully distinguish positive ABO blood types, with O+ distinguishable from other blood types at \u3e95% confidence. This is an important foundation for exploring DC dielectrophoretic sorting of blood types. The expansion of red blood cell sorting employing direct current insulative class=SpellE\u3edielectrophoresis (DC-iDEP) is novel. Here Pt electrodes were remotely situated in the inlet and outlet ports of the microdevice and an insulating obstacle generates the required dielectrophoretic force. The presence of ABO antigens on the red blood cell were found to affect the class=SpellE\u3edielectrophoretic deflection around the insulating obstacle thus sorting cells by type. To optimize the placement of insulating obstacle in the microchannel, COMSOL Multiphysics® simulations were performed. Microdevice dimensions were optimized by evaluating the behaviors of fluorescent polystyrene particles of three different sizes roughly corresponding to the three main components of blood: platelets (2-4 µm), erythrocytes (6-8 µm) and leukocytes (10-15 µm). This work provided the operating conditions for successfully performing size dependent blood cell insulator based DC dielectrophoresis in PDMS microdevices. In subsequent studies, the optimized microdevice geometry was then used for continuous separation of erythrocytes. The class=SpellE\u3emicrodevice design enabled erythrocyte collection into specific channels based on the cell’s deflection from the high field density region of the obstacle. The channel with the highest concentration of cells is indicative of the ABO blood type of the sample. DC resistance measurement system for quantification of erythrocytes was developed with single PDMS class=SpellE\u3emicrochannel system to be integrated with the DC- class=SpellE\u3eiDEP device developed in this research. This lab-on-a-chip technology application could be applied to emergency situations and naturalcalamities for accurate, fast, and portable blood typing with minimal error

    Dielectrophoretic characterization of particles and erythrocytes

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    Medical lab work, such as blood testing, will one day be near instantaneous and inexpensive via capabilities enabled by the fast growing world of microtechnology. In this research study, sorting and separation of different ABO blood types have been investigated by applying alternating and direct electric fields using class=SpellE\u3edielectrophoresis in microdevices. Poly(dimethylsiloxane) (PDMS) microdevices, fabricated by standard photolithography techniques have been used. Embedded perpendicular platinum (Pt) electrodes to generate forces in AC dielectrophoresis were used to successfully distinguish positive ABO blood types, with O+ distinguishable from other blood types at \u3e95% confidence. This is an important foundation for exploring DC dielectrophoretic sorting of blood types. The expansion of red blood cell sorting employing direct current insulative class=SpellE\u3edielectrophoresis (DC-iDEP) is novel. Here Pt electrodes were remotely situated in the inlet and outlet ports of the microdevice and an insulating obstacle generates the required dielectrophoretic force. The presence of ABO antigens on the red blood cell were found to affect the class=SpellE\u3edielectrophoretic deflection around the insulating obstacle thus sorting cells by type. To optimize the placement of insulating obstacle in the microchannel, COMSOL Multiphysics® simulations were performed. Microdevice dimensions were optimized by evaluating the behaviors of fluorescent polystyrene particles of three different sizes roughly corresponding to the three main components of blood: platelets (2-4 µm), erythrocytes (6-8 µm) and leukocytes (10-15 µm). This work provided the operating conditions for successfully performing size dependent blood cell insulator based DC dielectrophoresis in PDMS microdevices. In subsequent studies, the optimized microdevice geometry was then used for continuous separation of erythrocytes. The class=SpellE\u3emicrodevice design enabled erythrocyte collection into specific channels based on the cell’s deflection from the high field density region of the obstacle. The channel with the highest concentration of cells is indicative of the ABO blood type of the sample. DC resistance measurement system for quantification of erythrocytes was developed with single PDMS class=SpellE\u3emicrochannel system to be integrated with the DC- class=SpellE\u3eiDEP device developed in this research. This lab-on-a-chip technology application could be applied to emergency situations and naturalcalamities for accurate, fast, and portable blood typing with minimal error

    Improving the Design and Application of Insulator-Based Dielectrophoretic Devices for the Assessment of Complex Mixtures

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    Dielectrophoresis (DEP) is an electrokinetic (EK) transport mechanism that exploits polarization effects when particles are exposed to a non-uniform electric field. This dissertation focused on the development of high-performance insulator-based DEP (iDEP) devices. A detailed analysis of the spatial forces that contribute to particle movement in an iDEP device is provided. In particular, this analysis shows how particle size and shape affects the regions where particles are likely to be retained due to dielectrophoretic trapping. The performance of these trapping regions was optimized using a systematic approach that integrates the geometrical parameters of the array of insulating structures. Devices that decrease the required electrical potential by ~80% where found. The optimization strategy enabled the detection of structures that promote and discourage particle trapping. By combining the best and worst structures in a single asymmetric structure, a novel iDEP device was designed. This device selectively enriches the larger particles in a sample and drives the smaller particles away from the enrichment region. A quick enrichment and elution of large cells was achieved. This is important when dealing with samples containing eukaryotic cells, which can be harmed by the electrical treatment. Yeast cells were successfully separated from polystyrene particles in under 40 seconds using this device and a high cell viability of 85% was achieved. Finally, an enhancement of traditional iDEP devices is proposed, where some insulating posts are replaced by conducting structures. That is, insulating and conductive posts are intimately combined within the same array. The performance of this hybrid device is presented to show the advantage of using insulating structures with microelectrodes in the same array to dominate particle movement

    MICROFLUIDIC PARTICLE AND CELL MANIPULATION USING RESERVOIR-BASED DIELECTROPHORESIS

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    Controlled manipulation of synthetic particles and biological cells from a complex mixture is important to a wide range of applications in biology, environmental monitoring, and pharmaceutical industry. In the past two decades microfluidics has evolved to be a very useful tool for particle and cell manipulations in miniaturized devices. A variety of force fields have been demonstrated to control particle and cell motions in microfluidic devices, among which electrokinetic techniques are most often used. However, to date, studies of electrokinetic transport phenomena have been primarily confined within the area of microchannels. Very few works have addressed the electrokinetic particle motion at the reservoir-microchannel junction which acts as the interface between the macro (i.e., reservoir) and the micro (i.e., microchannel) worlds in real microfluidic devices. This Dissertation is dedicated to the study of electrokinetic transport and manipulation of particles and cells at the reservoir-microchannel junction of a microfluidic device using a combined experimental, theoretical, and numerical analysis. First, we performed a fundamental study of particles undergoing electrokinetic motion at the reservoir-microchannel junction. The effects of AC electric field, DC electric field, and particle size on the electrokinetic motion of particles passing through the junction were studied. A two-dimensional numerical model using COMSOL 3.5a was developed to investigate and understand the particle motion through the junction. It was found that particles can be continuously focused and even trapped at the reservoir-microchannel junction due to the effect of reservoir-based dielectrophoresis (rDEP). The electrokinetic particle focusing increases with the increase in AC electric field and particle size but decreases with the increase in DC electric field. It was also found that larger particles can be trapped at lower electric fields compared to smaller counterparts. Next, we utilized rDEP to continuously separate particles with different sizes at the reservoir-microchannel junction. The separation process utilized the inherent electric field gradients formed at the junction due to the size difference between the reservoir and the microchannel. It was observed, that the separation efficiency was reduced by inter-particle interactions when particles with small size differences were separated. The effect of enhanced electrokinetic flow on the separation efficiency was investigated experimentally and was observed to have a favorable effect. We also utilized rDEP approach to separate particles based on surface charge. Same sized particles with difference in surface charge were separated inside the microfluidic reservoir. The streaming particles interacted with the trapped particles and reduced the separation efficiency. The influences from the undesired particle trapping have been found through experiments to decrease with a reduced AC field frequency. Then, we demonstrated a continuous microfluidic separation of live yeast cells from dead cells using rDEP. Because the membrane of a cell gets distorted when it loses its viability, a higher exchange of ions results from such viability loss. The increased membrane conductivity of dead cells leads to a different Claussius-Mossoti factor from that of live cells, which enables their selective trapping and continuous separation based on cell viability. A two-shell numerical model was developed to account for the varying conductivities of different cell layers, the results of which agree reasonably with the experimental observations. We also used rDEP to implement a continuous concentration and separation of particles/cells in a stacked microfluidics device. This device has multiple layers and multiple microchannels on each layer so that the throughput can be significantly increased as compared to a single channel/single layer device. Finally, we compared the two-dimensional and three-dimensional particle focusing and trapping at the reservoir-microchannel junction using rDEP. We observed that the inherent electric field gradients in both the horizontal and vertical planes of the junction can be utilized if the reservoir is created right at the reservoir-microchannel junction. Three-dimensional rDEP utilizes the additional electric field gradient in the depth wise direction and thus can produce three-dimensional focusing. The electric field required to trap particles is also considerably lower in three-dimensional rDEP as compared to the two-dimensional rDEP, which thus considerably reduces the non-desired effects of Joule heating. A three-dimensional numerical model which accounted for the entire microfluidic device was also developed to predict particle trajectories

    Micro/Nano-Chip Electrokinetics

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    Micro/nanofluidic chips have found increasing applications in the analysis of chemical and biological samples over the past two decades. Electrokinetics has become the method of choice in these micro/nano-chips for transporting, manipulating and sensing ions, (bio)molecules, fluids and (bio)particles, etc., due to the high maneuverability, scalability, sensitivity, and integrability. The involved phenomena, which cover electroosmosis, electrophoresis, dielectrophoresis, electrohydrodynamics, electrothermal flow, diffusioosmosis, diffusiophoresis, streaming potential, current, etc., arise from either the inherent or the induced surface charge on the solid-liquid interface under DC and/or AC electric fields. To review the state-of-the-art of micro/nanochip electrokinetics, we welcome, in this Special Issue of Micromachines, all original research or review articles on the fundamentals and applications of the variety of electrokinetic phenomena in both microfluidic and nanofluidic devices

    Isomotive dielectrophoresis for enhanced analyses of cell subpopulations.

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    As the relentless dream of creating a true lab-on-a-chip device is closer to realization than ever before, which will be enabled through efficient and reliable sample characterization systems. Dielectrophoresis (DEP) is a term used to describe the motion of dielectric particles/ cells, by means of a non-uniform electric field (AC or DC). Cells of different dielectric properties (i.e., size, interior properties, and membrane properties) will act differently under the influence of dielectrophoretic force. Therefore, DEP can be used as a powerful, robust, and flexible tool for cellular manipulation, separation, characterization, and patterning. However, most recent DEP applications focus on trapping, separation, or sorting particles. The true value of DEP lies in its analytical capabilities which can be achieved by utilizing isomotive dielectrophoresis (isoDEP). In isoDEP, the gradient of the electric field-squared is constant, hence, upon the application of electric field, all particles/cells that share the same dielectric properties will feel the same constant dielectrophoretic force i.e., translate through the micro-channel at the same velocity. However, DEP is not the only acting force upon particles inside an isoDEP device, other electrokinetics, including but not limited to electrothermal hydrodynamics, might act on particles simultaneously. Within this dissertation, electrothermal-based experiments have been conducted to assess the effect of such undesired forces. Also, to maximize the relative DEP force over other forces for a given cell/particle size, design parameters such as microchannel width, height, fabrication materials, lid thickness, and applied electric field must be properly tuned. In this work, scaling law analyses were developed to derive design rules that relate those tunable parameters to achieve the desired dielectrophoretic force for cell analysis. Initial results indicated that for a particle suspended in 10 mS/m media, if the channel width and height are below 10 particle diameters, the electrothermal-driven flow is reduced by ∼ 500 times compared to the 500 µm thick conventional isoDEP device. Also, Replacing glass with silicon as the device’s base for an insulative-based isoDEP, reduces the electrothermal induced flow by ∼ 20 times. Within this dissertation, different device designs and fabrication methods were attempted in order to achieve an isoDEP platform that can characterize and differentiate between live and dead phytoplankton cells suspended in the same solution. Unfortunately, unwanted electrokinetics (predicted by the previously mentioned scaling law analysis) prevented comprehensive isoDEP analysis of phytoplankton cells. Due to isoDEP device limitations and other complications, other techniques were pursued to electrically characterize phytoplankton cells in suspension. An electrochemical-based platform utilizing impedance spectroscopy measurements was used to extract the electrical properties of phytoplankton cells in suspension. Impedance spectroscopy spectra were acquired, and the single-shell model was applied to extract the specific membrane capacitance, cytoplasm permittivity, and conductivity of assumingly spherical cells in suspension utilizing Maxwell’s mixture theory of a controlled volume fraction of cells. The impedance of suspensions of algae were measured at different frequencies ranging from 3 kHz to 10 MHz and impedance values were compared to investigate differences between two types of cells by characterizing their change in cytoplasm permittivity and membrane capacitance. Differentiation between healthy control and nitrogen-depleted cultured algae was attempted. The extracted specific membrane capacitances of Chlamydomonas and Selenastrum were 15:57 ± 3:62 and 40:64 ± 12:6 mF/m2 respectively. Successful differentiation based on the specific membrane capacitance of different algae species was achieved. However, no significant difference was noticed between nitrogen abundant and nitrogen depleted cultures. To investigate the potential of isoDEP for cell analysis, a comparison to existing dielectrophoresis-based electrokinetic techniques was encouraged, including electrorotation (ROT) microfluidic platforms. The ROT microfluidic chip was used to characterize M17, HEK293, T-lymphocytes, and Hela single cells. Through hands-on experience with ROT, the advantages and disadvantages of this approach and isoDEP are apparent. IsoDEP proves to be a good characterization tool for subpopulation cell analysis with potential higher throughput compared to ROT while maintaining simple fabrication and operation processes. To emphasize the role of dielectrophoresis in biology, further studies utilizing the 3DEP analytical system were used to determine the electrical properties of Drosophila melanogaster (Kc167) cells ectopically expressing Late embryogenesis abundant (LEA) proteins from the anhydrobiotic brine shrimp, Artemia franciscana. Dielectrophoretic-based characterization data demonstrates that single expression of two different LEA proteins, AfrLEA3m and AfrLEA6, both increase cytoplasmic conductivity of Kc167 cells to a similar extend above control values. The extracted DEP data supported previously reported data suggesting that AfrLEA3m can interact directly with membranes during water stress. This hypothesis was strengthened using scanning electron microscopy, where cells expressing AfrLEA3m were found to retain their spherical morphology during desiccation, while control cells exhibited a larger variety of shapes in the desiccated state
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