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New Tools for Trapping and Separation in Gas Chromatography and Dielectrophoresis : Improved Performance by Aid of Computer SimulationAldaeus, Fredrik January 2007 (has links)
<p>Computer simulations can be useful aids for both developing new analytical methods and enhancing the performance of existing techniques. This thesis is based on studies in which computer simulations were key elements in the development of several new tools for use in gas chromatography and dielectrophoresis. In gas chromatography, gaseous analytes are separated by exploiting differences in their partitioning between different phases, and after their partitioning parameters have been determined the separations can be computationally predicted, and optimized, for a wide range of operating conditions. Similarly, in dielectrophoresis, particles with differing polarizability or size can be separated, and since particle trajectories within a separation device can be predicted using computations, the suitability of new designs, applications of forces and combinations of operational parameters can be assessed without necessarily making or empirically testing all of the variants.</p><p>Using two existing numerical methods combined with semi-empirical determinations of retention behavior, temperature-programmed gas chromatograms were predicted with less than one percent deviations from experimental data, and a new method for improving the capacity of a gas-trapping device was predicted and experimentally verified. In addition, two new concepts with potential capacity to enhance dielectrophoretic separations were developed and tested in simulations. The first provides a promising way to improve the trapping of bacteria in media with elevated conductivity by using super-positioned electric fields, and the second a way to increase selectivity in the separation of bio-particles by using multiple dielectrophoretic cycles. The studies also introduced a more accurate method for determining the conductivity of suspensions of bacteria, and a new computational method for determining the dielectrophoretic behavior of particles in concentrated suspensions.</p><p>The scientific studies are summarized and discussed in the main text of this thesis, and presented in detail in seven appended papers.</p>
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Single-Molecule Detection and Optical Scanning in Miniaturized FormatsMelin, Jonas January 2006 (has links)
<p>In later years polymer replication techniques have become a frequently employed fabrication method for microfluidic and micro-optical devices. This thesis describes applications and further developments of microstructures replicated in polymer materials. </p><p>A novel method for homogenous amplified single-molecule detection utilizing a microfluidic readout format is presented. The method enables enumeration of single biomolecules by transforming specific molecular recognition events at nanometer dimensions to micrometer-sized DNA macromolecules. This transformation process is mediated by target specific padlock probe ligation, followed by rolling circle amplification (RCA) resulting in the creation of one rolling circle product (RCP) for each recognized target. Throughout this transformation the discrete nature of the molecular population is preserved. By hybridizing a fluorescence-labeled DNA detection oligonucleotide to each repeated sequence of the RCP, a confined cluster of fluorophores is generated, which makes optical detection and quantification possible. Spectral multiplexing is also possible since the spectral profile of each RCP can be analyzed separately. The microfluidic data acquisition process is characterized in detail and conditions that allow for quantification limited only by Poisson sampling statistics is established. The molecular characteristics of RCPs in solution are also investigated.</p><p>Furthermore a novel thermoplastic microfluidic platform is described. The platform allows for observation of the microchannels using high magnification optics and also offers the possibility of on-chip cell culture and the integration of mechanical actuators.</p><p>A novel fabrication process for the integration of polymer micro-optical elements on silicon is presented. The process is used for fabrication of a micro-optical system consisting of a laser and a movable microlens making beam steering possible. Such a micro-scanning system could potentially be used for miniaturized biochemical analysis.</p>
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Single-Molecule Detection and Optical Scanning in Miniaturized FormatsMelin, Jonas January 2006 (has links)
In later years polymer replication techniques have become a frequently employed fabrication method for microfluidic and micro-optical devices. This thesis describes applications and further developments of microstructures replicated in polymer materials. A novel method for homogenous amplified single-molecule detection utilizing a microfluidic readout format is presented. The method enables enumeration of single biomolecules by transforming specific molecular recognition events at nanometer dimensions to micrometer-sized DNA macromolecules. This transformation process is mediated by target specific padlock probe ligation, followed by rolling circle amplification (RCA) resulting in the creation of one rolling circle product (RCP) for each recognized target. Throughout this transformation the discrete nature of the molecular population is preserved. By hybridizing a fluorescence-labeled DNA detection oligonucleotide to each repeated sequence of the RCP, a confined cluster of fluorophores is generated, which makes optical detection and quantification possible. Spectral multiplexing is also possible since the spectral profile of each RCP can be analyzed separately. The microfluidic data acquisition process is characterized in detail and conditions that allow for quantification limited only by Poisson sampling statistics is established. The molecular characteristics of RCPs in solution are also investigated. Furthermore a novel thermoplastic microfluidic platform is described. The platform allows for observation of the microchannels using high magnification optics and also offers the possibility of on-chip cell culture and the integration of mechanical actuators. A novel fabrication process for the integration of polymer micro-optical elements on silicon is presented. The process is used for fabrication of a micro-optical system consisting of a laser and a movable microlens making beam steering possible. Such a micro-scanning system could potentially be used for miniaturized biochemical analysis.
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New Tools for Trapping and Separation in Gas Chromatography and Dielectrophoresis : Improved Performance by Aid of Computer SimulationAldaeus, Fredrik January 2007 (has links)
Computer simulations can be useful aids for both developing new analytical methods and enhancing the performance of existing techniques. This thesis is based on studies in which computer simulations were key elements in the development of several new tools for use in gas chromatography and dielectrophoresis. In gas chromatography, gaseous analytes are separated by exploiting differences in their partitioning between different phases, and after their partitioning parameters have been determined the separations can be computationally predicted, and optimized, for a wide range of operating conditions. Similarly, in dielectrophoresis, particles with differing polarizability or size can be separated, and since particle trajectories within a separation device can be predicted using computations, the suitability of new designs, applications of forces and combinations of operational parameters can be assessed without necessarily making or empirically testing all of the variants. Using two existing numerical methods combined with semi-empirical determinations of retention behavior, temperature-programmed gas chromatograms were predicted with less than one percent deviations from experimental data, and a new method for improving the capacity of a gas-trapping device was predicted and experimentally verified. In addition, two new concepts with potential capacity to enhance dielectrophoretic separations were developed and tested in simulations. The first provides a promising way to improve the trapping of bacteria in media with elevated conductivity by using super-positioned electric fields, and the second a way to increase selectivity in the separation of bio-particles by using multiple dielectrophoretic cycles. The studies also introduced a more accurate method for determining the conductivity of suspensions of bacteria, and a new computational method for determining the dielectrophoretic behavior of particles in concentrated suspensions. The scientific studies are summarized and discussed in the main text of this thesis, and presented in detail in seven appended papers.
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Integrated Microfluidic Optical Manipulation Technique: Towards High Throughput Single Cell AnalysisCharron, Luc 20 August 2012 (has links)
An all-optical micromanipulation technique is presented in the framework of precise cell selection within a cell culture and multiplexed transport capabilities for microfluidic single cell analysis applications. The technique was developed by combining an optical tweezer setup with a novel integrated waveguide cell propulsion method referred to as end-face waveguide propulsion (EFWP). The EFWP technique delivers optical forces to a particle generating thrust. The thesis is divided into two sections: simulation and experimental validation. In the first section a new simulation technique based on ray optics theory (ROT) and the beam propagation method (BPM) is used to predict particle velocity and trajectory along a microfluidic propagation channel. In this work, the ROT-BPM technique is used to analyse and optimize the waveguide geometry to maximize particle velocity. Analysis of the impact of common microchip manufacturing limitations on velocity is performed to determine acceptable fabrication process tolerances. The second section presents experimental results of polymer microspheres and acute myeloid leukemia (AML) cells as biological targets. The experimental results are compared with simulations performed in the first section. Correction factors are added to the simulations to reflect the experimental device parameters. Thermal e_ects due to photon absorption within the fluidic channels are also investigated and corrected for. The final analysis indicates that the ROT-BPM technique developed in this work can be used to adequately predict particle velocity and trajectory path. EFWP currently delivers the fastest particle velocities compared to other optical micromanipulation techniques currently available in microfluidic applications. While the technique is focused on addressing chemical cytometry precise particle selectivity and high throughput needs, EFWP can also be used in many other single cell applications.
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Development of a Microfluidic Device for Selective Electrical Lysis of Plasma Membranes of Single CellsShah, Duoaud F. 11 January 2011 (has links)
A primary objective of modern biology is to understand the molecular mechanisms which underlie cellular functions and a crucial part of this task is the ability to manipulate and analyze individual cells. As a result of interdisciplinary research, microfluidics may become the forefront of analytical methods used by biologists. This technology can be used to gain unprecedented opportunities for cell handling, lysis and investigation on a single cell basis. This thesis presents the development of a microfluidic device capable of selecting individual cells and performing selective electrical lysis of the plasma membrane, while verifying intactness of the nuclear membrane. The device is fabricated by an improved photolithography method and integrates molten solder as electrodes for lysis by a DC electric field. Quantification of lysis is accomplished by video and image analysis, and measurement of the rate of ion diffusion from the cell.
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Development of a Microfluidic Device for Selective Electrical Lysis of Plasma Membranes of Single CellsShah, Duoaud F. 11 January 2011 (has links)
A primary objective of modern biology is to understand the molecular mechanisms which underlie cellular functions and a crucial part of this task is the ability to manipulate and analyze individual cells. As a result of interdisciplinary research, microfluidics may become the forefront of analytical methods used by biologists. This technology can be used to gain unprecedented opportunities for cell handling, lysis and investigation on a single cell basis. This thesis presents the development of a microfluidic device capable of selecting individual cells and performing selective electrical lysis of the plasma membrane, while verifying intactness of the nuclear membrane. The device is fabricated by an improved photolithography method and integrates molten solder as electrodes for lysis by a DC electric field. Quantification of lysis is accomplished by video and image analysis, and measurement of the rate of ion diffusion from the cell.
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Integrated Microfluidic Optical Manipulation Technique: Towards High Throughput Single Cell AnalysisCharron, Luc 20 August 2012 (has links)
An all-optical micromanipulation technique is presented in the framework of precise cell selection within a cell culture and multiplexed transport capabilities for microfluidic single cell analysis applications. The technique was developed by combining an optical tweezer setup with a novel integrated waveguide cell propulsion method referred to as end-face waveguide propulsion (EFWP). The EFWP technique delivers optical forces to a particle generating thrust. The thesis is divided into two sections: simulation and experimental validation. In the first section a new simulation technique based on ray optics theory (ROT) and the beam propagation method (BPM) is used to predict particle velocity and trajectory along a microfluidic propagation channel. In this work, the ROT-BPM technique is used to analyse and optimize the waveguide geometry to maximize particle velocity. Analysis of the impact of common microchip manufacturing limitations on velocity is performed to determine acceptable fabrication process tolerances. The second section presents experimental results of polymer microspheres and acute myeloid leukemia (AML) cells as biological targets. The experimental results are compared with simulations performed in the first section. Correction factors are added to the simulations to reflect the experimental device parameters. Thermal e_ects due to photon absorption within the fluidic channels are also investigated and corrected for. The final analysis indicates that the ROT-BPM technique developed in this work can be used to adequately predict particle velocity and trajectory path. EFWP currently delivers the fastest particle velocities compared to other optical micromanipulation techniques currently available in microfluidic applications. While the technique is focused on addressing chemical cytometry precise particle selectivity and high throughput needs, EFWP can also be used in many other single cell applications.
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Development of Cell Lysis Techniques in Lab on a chipShahini, Mehdi January 2013 (has links)
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.
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Modeling, Fabrication, and Test of a CMOS Integrated Circuit Platform for Electrophoretic Control of On-Chip Heterogeneous Fluids: toward Particle Separation on a Custom CMOS ChipWake, Heather Anne January 2009 (has links)
<p>Electrophoresis is the migration of charged particles in a heterogeneous fluid under the influence of an electric field. This project is work toward an electrophoretic separation system on a custom CMOS chip. Modeling, fabrication, and testing of an AMI ABN 1.5 um CMOS chip for this application is discussed. The unique approach is to build the entire system using conventional CMOS integrated circuit technology, such that the separation area is fabricated on the chip with integrated control and detection circuitry. To achieve the desired functionality, a novel configuration of an electrophoresis system is implemented. In this system, instead of using only one electrode at each end of the separation area, a multitude of electrodes beneath the entire separation area are utilized, enabling better control of high electric fields using very small voltages over small areas. Electronic circuits control the position and strength of the electric field to drive the separations and to simultaneously detect the location and concentration of samples within the separation area. Ultimately, the project was successful at showing that implementing an electrophoresis system on standard CMOS is possible.</p> / Dissertation
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