Doctoral Dissertations

Date of Award

8-2007

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Mechanical Engineering

Major Professor

Kenneth D. Kihm

Committee Members

Anthony English, Majid Keyhani, Jayne Wu

Abstract

My doctoral research has focused on the development of microscale optical techniques for examining micro/bio fluidics. Preliminary work measured the velocity field in a microchannel, by optical slicing, using Confocal Laser Scanning Microscopy (CLSM). Next, Optical Serial Sectioning Microscopy (OSSM) was applied to examine thermometry by detecting the free Brownian motion of nano-particles suspended in mediums at different temperatures. An extension of this work used objective-based Total Internal Reflection Fluorescence Microscopy (TIRFM) to examine the hindered Brownian motion of nano-particles that were very close to a solid surface (within 1 mm).

An optically transparent and electrically conductive Indium Tin Oxide (ITO) biosensor and an integrated dynamic live cell imaging system were developed to dynamically examine changes in cell coverage area, cell morphology, cell-substrate adhesion, and cell-cell interaction. To our knowledge this is the first sensor capable of conducting simultaneous optical and electrical measurements. This system consists of an incubator, which keeps cells viable by providing the necessary environmental conditions (37 °C temperature and 5 % CO2), and multiple microscopy techniques, including multispectrum Interference Reflection Microscopy (MS-IRM), TIRFM, Epi-fluorescence Microscopy, Phase Contrast Microscopy (PCM), and Differential Interference Contrast Microscopy (DICM). Along with investigations of cytometric proliferation including cellular barrier functions, in vitro cytotoxicity experiments were also conducted to examine the effect of a drug (cytochalasin D, a toxic agent) on cellular motility and cellular morphology. These cytotoxicity results give us a fundamental understanding of the cellular processes induced by the drug, which will be invaluable in the search for methods of preventing metastases. In this research, MS-IRM is used to examine the focal contacts and the gap morphology between cells and substrates, DICM is used to examine the coverage area of cells, and impedance measurements are used to correlate these two parameters.

Advances in the understanding of vascular bio-transport in endothelial cells will have an impact on many aspects of cell biology, tissue engineering, and pharmacology. Particularly important will be the ability to test the popular hypothesis that the cell barrier function is regulated by specific cytoskeleton elements controlling intercellular and extracellular coupling.

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