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PhD Dissertation_Chang_07192.pdf (6.11 MB)

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3D Nanochannel Array Platform for High-throughput Cell Manipulation and Nano-electroporation

Chang, Lingqian
http://rave.ohiolink.edu/etdc/view?acc_num=osu1468876520

Abstract Details

2016, Doctor of Philosophy, Ohio State University, Biomedical Engineering.
Electroporation is one of the most common non-viral methods for gene delivery. Recent progress in gene therapy has offered special opportunities to electroporation for in vitro and in vivo applications. However, conventional bulk electroporation (BEP) inevitably causes serious cell damage and stochastic transfection between cells. Microfluidic electroporation (MEP) has been claimed to provide benign single cell transfection for the last decade. Nevertheless, the intracellular transport in both MEP and BEP systems is highly diffusion-dominant, which prevents precise dose control and high uniformity. In this Ph.D. research, we developed a 3D nanochannel-electroporation (3D NEP) platform for mass cell transfection. A silicon-based nanochannel array (3D NEP) chip was designed and fabricated for cell manipulation and electroporation. The chip, designed as Z-directional microchannel – nanochannel array, was fabricated by clean room techniques including projection photolithography and deep reactive-ion etching (DRIE). The fabricated 3D NEP chip is capable of handling 40,000 cells per 1 cm2, up to 1 million per wafer (100 mm diameter). High-throughput cell manipulation technologies were investigated for precise alignment of individual cells to the nanochannel array, a key step for NEP to achieve dose control. We developed three techniques for cell trapping in this work. (1) Magnetic tweezers (MTs) were integrated on the chip to remotely control cells under a programmed magnetic field. (2) A positive dielectrophoresis (pDEP) power system was built as an alternative to trap cells onto the nanochannel array using DEP force. (3) A novel yet simple `dipping-trap’ method was used to rapidly trap cells onto a nanochannel array, aligned by a micro-cap array pattern on the 3D NEP chip, which eventually offered 70 – 90 % trapping efficiency and 90 % specificity. 3D NEP platforms were assembled for cell transfection based on the Si-based nanochannel array chip and cell manipulation techniques. Cells were patterned on the nanochannel array and collectively were electroporated in parallel, injected with cargo in Z-direction. Controlling the dose was demonstrated with the external pulse durations at high-throughput. The `electrophoretic’- expedited delivery of large molecular weight plasmids were demonstrated with large numbers of primary cells simultaneously, which cannot be achieved in BEP and MEP. Two clinically valuable case studies were performed with our 3D NEP for living cell sensing / interrogation. (1) In the case of in vitro transfection of primary cardiomyocytes, we studied the dose-effects of miR-29 on mitochondrial changes and the suppression of the Mcl-1 gene in adult mouse cardiomyocytes by precisely controlling the miR-29 dose injected. (2) Glioma stem cells (GSCs), a type of cell hypothesized to be highly aggressive and to lead to the relapses of gliobastoma in human brain, was studied at single cell resolution on 3D NEP platform. The developed 3D NEP system moves towards clinically oriented and user-friendly tools for life science applications. The batch-treated cells with controlled dosage delivery provide a useful tool for single cell analysis. The pioneering experiments in this work have demonstrated the 3D NEP for the applications of cell reprogramming, adoptive immunotherapy, in vitro cardiomyocytes transfection and glioma stem cells study.
Ly James Lee (Advisor)
Wu Lu (Advisor)
Derek Hansford (Committee Member)
179 p.
Biomedical Engineering; Electrical Engineering; Mechanical Engineering; Nanotechnology

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Citations

  • Chang, L. (2016). 3D Nanochannel Array Platform for High-throughput Cell Manipulation and Nano-electroporation [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1468876520

    APA Style (7th edition)

  • Chang, Lingqian. 3D Nanochannel Array Platform for High-throughput Cell Manipulation and Nano-electroporation. 2016. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1468876520.

    MLA Style (8th edition)

  • Chang, Lingqian. "3D Nanochannel Array Platform for High-throughput Cell Manipulation and Nano-electroporation." Doctoral dissertation, Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1468876520

    Chicago Manual of Style (17th edition)

osu1468876520
1,109
© 2016, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.