Date on Master's Thesis/Doctoral Dissertation

1-2021

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Electrical and Computer Engineering

Degree Program

Electrical Engineering, PhD

Committee Chair

Williams, Stuart

Committee Co-Chair (if applicable)

Naber, John

Committee Member

Naber, John

Committee Member

Harnett

Committee Member

Harnett, Cindy

Committee Member

McNamara, Shamus

Author's Keywords

microfluidics; dielectrophoresis cell; analysis; electrorotation; isomotive dielectrophoresis; isoDEP

Abstract

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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