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Micro-channel Array Platform: Studies on Vacuum-assisted Cell Trapping and 3D Micro-electroporation

Chang, Lingqian
http://rave.ohiolink.edu/etdc/view?acc_num=osu1376591662

Abstract Details

2013, Master of Science, Ohio State University, Biomedical Engineering.
Trapping of single cells in the functional regions is an important theme of microfluidic systems. Particularly in the 3D micro-channel array based electroporation systems (3D MEP), a good alignment between cells and channels gives rise to a uniform and safe transfection at the single-cell level. In the present work we carried out studies to seed cells to a micro-channel array chip using vacuum-assisted hydrodynamics trap. A 3D MEP was developed to deliver transfect factors into the trapped cells for cell biology studies. We tried glass etching and silicon etching to fabricate a micro-channel array chip with pore size less than 5 micrometers. In the process of hydrofluoric acid based isotropic etch of glass, we developed a bowl-shaped micro-pore array using a Cr/Au/photoresist mask, yet the size of micro-channels was as big as 30 micrometers. A micro-channel array with size less than 5 micrometers was fabricated using the potassium hydroxide based wet etching and the fluorine gas based dry etching. We studied the alkaline resistance of different materials used for both the mask layer and the protection layer. The results showed that the sandwich mask (SiO2-Si3N4-SiO2) by PECVD had good performance in the silicon wet etching. Next, we studied on the vacuum-assisted cells trapping. A vacuum setup was developed to generate a negative pressure to draw cells onto the micro-channel array, eventually to form a cell array on the chip. We tried to understand the relationships between six independent variables in the trapping procedure and the trapping efficiency. The cell density, the negative pressure and the cell buffer volume were directly proportional to the trapping efficiency. The best channel size 3 ~ 4 micrometers was identified with respect to the cell deformation. Poly (ethylene glycol) (PEG) was found to improve the trapping efficiency by facilitating the detachment of the cells from the chip surface. We found that the pipetting method worked better than the chip shaken for detaching cells. We tried to trap 1,000 cells onto 1,000 channels, and achieved 70% trap efficiency under the optimum conditions. 95% trapped cells were health after 24 hours on-chip culture, indicating the micro-channel array environment was compatible to cells. We have developed a 3D MEP system to electroporate the cells trapped on the micro-channel array. We delivered PI into MEFs randomly loaded on the chip. The electroporation took place only in the cells aligned to the micro-channel array. The relative fluorescence intensity implied that the 3D MEP offered a uniform delivery of PI dye. For heterogeneity study of leukemia cells, we delivered GATA2 molecular beacon (MB) into K562 cells and Jurkat cells trapped on the micro-channel array. The results showed the positive fluorescence of GATA2 in K562 cells, indicating an up-regulation of intracellular GATA2 gene. On the contrary, there was no significantly positive signal of GATA2 in Jurkat cells. The difference demonstrated the 3D MEP system was capable of distinguishing the leukemia cell types based on GATA2 MB transfection at the single-cell level, and paved the road for monitoring the leukemia-initiating cells from patient samples.
L.James Lee (Committee Chair)
Derek Hansford (Committee Member)
Wu Lu (Other)
163 p.
Biomedical Engineering

Recommended Citations

Citations

  • Chang, L. (2013). Micro-channel Array Platform: Studies on Vacuum-assisted Cell Trapping and 3D Micro-electroporation [Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1376591662

    APA Style (7th edition)

  • Chang, Lingqian. Micro-channel Array Platform: Studies on Vacuum-assisted Cell Trapping and 3D Micro-electroporation. 2013. Ohio State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1376591662.

    MLA Style (8th edition)

  • Chang, Lingqian. "Micro-channel Array Platform: Studies on Vacuum-assisted Cell Trapping and 3D Micro-electroporation." Master's thesis, Ohio State University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=osu1376591662

    Chicago Manual of Style (17th edition)

osu1376591662
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© 2013, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.