Identifier

etd-11092009-135336

Degree

Doctor of Philosophy (PhD)

Department

Engineering Science (Interdepartmental Program)

Document Type

Dissertation

Abstract

The goal of this study is to combine molecular and microdevice methods to characterize and quantify viability of single mammalian cells. Fluorescent-based assays were optimized for adherent HeLa and suspension Jurkat cells and were used as a tool for validation of a microfabricated diagnostic device. Cell and substrate/surface interactions were considered for designing a microfluidic device that can be used to characterize cell viability for quantitative biomedical and cell biology applications, which require label-free, real-time monitoring of cells. Several interdisciplinary methods are employed to evaluate electrical impedance differences between live and dead Jurkat cells in a microfluidic device. Biological Micro-Electro-Mechanical Systems (BioMEMS) offer many advantages over the conventional macroscale approaches to biomedical diagnostics, such as reduced reagents, costs, and power consumption; shorter reaction time; portability; versatility; and potential for parallel, integrated operations, thus having the potential to revolutionize how many current cell-based biomolecular assays are performed. A microchip device to detect cell viability at the single-cell level in real-time has much potential for pharmacological drug screening or point-of-care diagnostics. Optimal cell media conditions such as pH and osmolarity are evaluated to ensure cell viability and adequate sensitivity for detecting cell events via electrical impedance measurements. A fluorescent cell assay using Calcein was optimized for optical validation of Jurkat cell viability studies for cells flowing through a microchannel. Fluorescence microscopy was combined with acquired electrical impedance (at 2 MHz) to validate the presence and viability of each cell at the detection electrodes. The microchip design parameters such as substrate material and geometry of microchannel and electrodes were based of the average 12 um-diameter of Jurkat cells tested. Here, we demonstrate the design of a polymer-based chip device that is able to differentiate between live and dead Jurkat cells on the basis of electrical impedance magnitude and phase signals, which could be related to inherent dielectric differences of live and dead cells. The overall outcome of this study provides groundwork for quantifying cell viability of single cells on-chip, in real-time, in a flow-through system, without the use of expensive fluorescent labels.

Date

2009

Document Availability at the Time of Submission

Secure the entire work for patent and/or proprietary purposes for a period of one year. Student has submitted appropriate documentation which states: During this period the copyright owner also agrees not to exercise her/his ownership rights, including public use in works, without prior authorization from LSU. At the end of the one year period, either we or LSU may request an automatic extension for one additional year. At the end of the one year secure period (or its extension, if such is requested), the work will be released for access worldwide.

Committee Chair

Monroe, W. Todd

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