Date on Master's Thesis/Doctoral Dissertation

8-2017

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Mechanical Engineering

Degree Program

Mechanical Engineering, PhD

Committee Chair

Keynton, Robert

Committee Co-Chair (if applicable)

Williams, Stuart

Committee Member

Brehob, Ellen

Committee Member

Cohn, Robert

Committee Member

Sumanasekera, Gamini

Committee Member

El-Baz, Ayman

Author's Keywords

electroporation; nanochannel; microfluidic; cell

Abstract

For traditional electroporation devices, there are a number of problems associated with these devices such as insufficient understanding of its theoretical mechanism, low cell viability, inadequate electroporation efficiency, excess voltage applied to generate required electric field due to the large size of these devices and sample contamination. Although newly developed microfluidic electroporation devices have solved most of the above existing problems in traditional bulk electroporation devices, they appear to lack the ability to control the precise dose of biomolecules or genes transfecting into cells and, from a manufacturing perspective, the fabrication methods do not enable repeatable production of such devices on the large scale. Here, we introduce a new, repeatable method for fabricating 3-D Micro/Nanofluidic electroporation platforms and characterize these platforms to demonstrate their ability to electroporate live cells. Some of the new methods developed in this work include a direct-write fiber technique via three-axis robotic dispensing system, dry film resist photolithography, film-to-film bonding and replica molding to create the desired electroporation platform. A robotic dispensing system was utilized to control the fiber diameter, which was determined vii by the: 1) prescribed dispense time; 2) pressure of the dispensing system valve; 3) rate at which the stage traversed; 4) diameter of the dispensing tip; 5) polymer solution viscosity and surface tension; and, 6) programmed drawing length. Thin dry film photoresist was utilized to replace liquid photoresist in order to achieve high-quality film-to-film bonding after drawing nanofibers onto one substrate containing the thin-film structure. Polydimethylsiloxane (PDMS) was employed as the bulk material to fabricate the target micro/nano electroporation substrate using replica molding and micro/nanofibers etching. Characterization of the direct-write fiber technique via robotic dispensing system to acquire suspended and complex fibers of the required dimension repeatedly under prescribed conditions were completed. Combining this fiber direct-write method and traditional clean room techniques, a total of 18 micro- to nano-scale electroporation devices (6 for each group of 1 ìm, 500 nm, and 300 nm diameter) were successfully developed and mass produced in two weeks with relatively high repeatability (within 20% of the design). Finally, metrology and characterization studies were performed on the electroporation platforms to validate the micro/nanochannel’s existence and its connectivity to two micro-chambers. Furthermore, biomolecules and other fluorescent particles were successfully transported through the micro/nanochannel and transferred (via electroporation) into the cells. Preliminary results of electroporation experiment performed on this micro/nano-electroporation platform illustrated that the duration of the entire electroporation process was significantly shorter than times reported previously by other investigator’s nano-electroporation platforms.

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