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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 沈弘俊(Horn-Jiunn Sheen) | |
dc.contributor.author | Che-Yi Li | en |
dc.contributor.author | 李哲儀 | zh_TW |
dc.date.accessioned | 2023-03-19T23:44:23Z | - |
dc.date.copyright | 2022-09-02 | |
dc.date.issued | 2022 | |
dc.date.submitted | 2022-08-29 | |
dc.identifier.citation | [1] Zborowski, M., Chalmers, J. J., & Lowrie, W. G. (2017). Magnetic cell manipulation and sorting. In Microtechnology for Cell manipulation and Sorting (pp. 15-55). Springer, Cham. [2] Lee, W., Tseng, P., & Di Carlo, D. (2017). Microfluidic cell sorting and separation technology. In Microtechnology for Cell Manipulation and Sorting (pp. 1-14). Springer, Cham. [3] Cheng, J., Rahman, M. A., & Ohta, A. T. (2017). Optical manipulation of cells. In Microtechnology for Cell Manipulation and Sorting, 93-128. [4] Qian, C., Huang, H., Chen, L., Li, X., Ge, Z., Chen, T., ... & Sun, L. (2014). Dielectrophoresis for bioparticle manipulation. International journal of molecular sciences, 15(10), 18281-18309. [5] Khoshmanesh, K., Nahavandi, S., Baratchi, S., Mitchell, A., & Kalantar-zadeh, K. (2011). Dielectrophoretic platforms for bio-microfluidic systems. Biosensors and Bioelectronics, 26(5), 1800-1814. [6] Pohl, H. A., Pollock, K., & Crane, J. S. (1978). Dielectrophoretic force: A comparison of theory and experiment. Journal of Biological Physics, 6(3), 133-160. [7] Jones, T. B. (1979). Dielectrophoretic force calculation. Journal of Electrostatics, 6(1), 69-82. [8] Punjiya, M., Nejad, H. R., Mathews, J., Levin, M., & Sonkusale, S. (2019). A flow through device for simultaneous dielectrophoretic cell trapping and AC electroporation. Scientific reports, 9(1), 1-11. [9] Jubery, T. Z., Srivastava, S. K., & Dutta, P. (2014). Dielectrophoretic separation of bioparticles in microdevices: A review. Electrophoresis, 35(5), 691-713. [10] Docoslis, A., Kalogerakis, N., Behie, L. A., & Kaler, K. V. (1997). A novel dielectrophoresis‐based device for the selective retention of viable cells in cell culture media. Biotechnology and bioengineering, 54(3), 239-250. [11] Yoshimura, Y., Tomita, M., Mizutani, F., & Yasukawa, T. (2014). Cell pairing using microwell array electrodes based on dielectrophoresis. Analytical chemistry, 86(14), 6818-6822. [12] Kim, T. K., & Eberwine, J. H. (2010). Mammalian cell transfection: the present and the future. Analytical and bioanalytical chemistry, 397(8), 3173-3178. [13] Shi, J., Ma, Y., Zhu, J., Chen, Y., Sun, Y., Yao, Y., ... & Xie, J. (2018). A review on electroporation-based intracellular delivery. Molecules, 23(11), 3044. [14] Woods, N. B., Muessig, A., Schmidt, M., Flygare, J., Olsson, K., Salmon, P., ... & Karlsson, S. (2003). Lentiviral vector transduction of NOD/SCID repopulating cells results in multiple vector integrations per transduced cell: risk of insertional mutagenesis. Blood, The Journal of the American Society of Hematology, 101(4), 1284-1289. [15] Schenborn, E. T., & Goiffon, V. (2000). DEAE-dextran transfection of mammalian cultured cells. Transcription Factor Protocols, 147-153. [16] Gordon, J. W., Scangos, G. A., Plotkin, D. J., Barbosa, J. A., & Ruddle, F. H. (1980). Genetic transformation of mouse embryos by microinjection of purified DNA. Proceedings of the National Academy of Sciences, 77(12), 7380-7384. [17] Klein, T. M., & Jones, T. J. (1999). Methods of genetic transformation: the gene gun. In Molecular improvement of cereal crops (pp. 21-42). Springer, Dordrecht. [18] Potter, H., & Heller, R. (2018). Transfection by electroporation. Current protocols in molecular biology, 121(1), 9-3. [19] Divecha, N., & Irvine, R. F. (1995). Phospholipid signaling. Cell, 80(2), 269-278. [20] Herrmann, T., Leavitt, L., & Sharma, S. (2021). Physiology, Membrane. StatPearls [Internet]. [21] Lombard, J. (2014). Once upon a time the cell membranes: 175 years of cell boundary research. Biology direct, 9(1), 1-35. [22] Marszalek, P., Liu, D. S., & Tsong, T. Y. (1990). Schwan equation and transmembrane potential induced by alternating electric field. Biophysical journal, 58(4), 1053-1058. [23] Rubinsky, B. (2007). Irreversible electroporation in medicine. Technology in cancer research & treatment, 6(4), 255-259. [24] Garcia, P. A., Rossmeisl, J. H., Neal, R. E., Ellis, T. L., Olson, J. D., Henao-Guerrero, N., ... & Davalos, R. V. (2010). Intracranial nonthermal irreversible electroporation: in vivo analysis. The Journal of membrane biology, 236(1), 127-136. [25] Chang, D. (Ed.). (1991). Guide to electroporation and electrofusion. Academic Press. [26] Neumann, E., Schaefer‐Ridder, M., Wang, Y., & Hofschneider, P. (1982). Gene transfer into mouse lyoma cells by electroporation in high electric fields. The EMBO journal, 1(7), 841-845. [27] Zimmermann, U. (1986). Electrical breakdown, electropermeabilization and electrofusion. Reviews of Physiology, Biochemistry and Pharmacology, Volume 105, 175-256. [28] Geboers, B., Scheffer, H. J., Graybill, P. M., Ruarus, A. H., Nieuwenhuizen, S., Puijk, R. S., ... & Meijerink, M. R. (2020). High-voltage electrical pulses in oncology: irreversible electroporation, electrochemotherapy, gene electrotransfer, electrofusion, and electroimmunotherapy. Radiology, 295(2), 254-272. [29] Tsong, T. Y. (1989). Electroporation of cell membranes. Electroporation and electrofusion in cell biology, 149-163. [30] Schwan, H. P. (1983). Biophysics of the interaction of electromagnetic energy with cells and membranes. In Biological effects and dosimetry of nonionizing radiation (pp. 213-231). Springer, Boston, MA. [31] Teissie, J., & Rols, M. P. (1993). An experimental evaluation of the critical potential difference inducing cell membrane electropermeabilization. Biophysical journal, 65(1), 409-413. [32] Lin, Y. C., Jen, C. M., Huang, M. Y., Wu, C. Y., & Lin, X. Z. (2001). Electroporation microchips for continuous gene transfection. Sensors and Actuators B: Chemical, 79(2-3), 137-143. [33] Huang, H., Wei, Z., Huang, Y., Zhao, D., Zheng, L., Cai, T., ... & Liang, Z. (2011). An efficient and high-throughput electroporation microchip applicable for siRNA delivery. Lab on a Chip, 11(1), 163-172. [34] Lungu, M. (2004). Electrical separation of plastic materials using the triboelectric effect. Minerals Engineering, 17(1), 69-75. [35] Fan, F. R., Tian, Z. Q., & Wang, Z. L. (2012). Flexible triboelectric generator. Nano energy, 1(2), 328-334. [36] Somkuwar, V. U., Pragya, A., & Kumar, B. (2020). Structurally engineered textile-based triboelectric nanogenerator for energy harvesting application. Journal of Materials Science, 55(12), 5177-5189. [37] Wu, C., Wang, A. C., Ding, W., Guo, H., & Wang, Z. L. (2019). Triboelectric nanogenerator: a foundation of the energy for the new era. Advanced Energy Materials, 9(1), 1802906. [38] Zhu, G., Lin, Z. H., Jing, Q., Bai, P., Pan, C., Yang, Y., ... & Wang, Z. L. (2013). Toward large-scale energy harvesting by a nanoparticle-enhanced triboelectric nanogenerator. Nano letters, 13(2), 847-853. [39] Lin, L., Wang, S., Xie, Y., Jing, Q., Niu, S., Hu, Y., & Wang, Z. L. (2013). Segmentally structured disk triboelectric nanogenerator for harvesting rotational mechanical energy. Nano letters, 13(6), 2916-2923. [40] Liu, Z., Nie, J., Miao, B., Li, J., Cui, Y., Wang, S., ... & Wang, Z. L. (2019). Self‐powered intracellular drug delivery by a biomechanical energy‐driven triboelectric nanogenerator. Advanced Materials, 31(12), 1807795. [41] Tang, W., Han, Y., Han, C. B., Gao, C. Z., Cao, X., & Wang, Z. L. (2015). Self‐powered water splitting using flowing kinetic energy. Advanced Materials, 27(2), 272-276. [42] Sano, M. B., Henslee, E. A., Schmelz, E., & Davalos, R. V. (2011). Contactless dielectrophoretic spectroscopy: examination of the dielectric properties of cells found in blood. Electrophoresis, 32(22), 3164-3171. [43] Garner, A. L., Chen, N., Yang, J., Kolb, J., Swanson, R. J., Loftin, K. C., ... & Schoenbach, K. H. (2004). Time domain dielectric spectroscopy measurements of HL-60 cell suspensions after microsecond and nanosecond electrical pulses. IEEE transactions on plasma science, 32(5), 2073-2084. [44] Wang, X., Becker, F. F., & Gascoyne, P. R. (2002). Membrane dielectric changes indicate induced apoptosis in HL-60 cells more sensitively than surface phosphatidylserine expression or DNA fragmentation. Biochimica et Biophysica Acta (BBA)-Biomembranes, 1564(2), 412-420. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/86243 | - |
dc.description.abstract | 本研究利用微機電技術製作出具有不同幾何結構之電穿刺裝置,以結構之電場放大效應提高細胞電穿刺效率,最終達到同時操縱與電穿刺細胞之目的。此裝置亦能夠以摩擦奈米發電機 (triboelectric nanogenerator, TENG) 作為電訊號來源,以達成電穿刺系統之自供電,並用於shRNA 轉染。 為了能有效達成細胞操縱與電穿刺,本研究以高透光性的氧化銦錫玻璃 (Indium tin oxide glass, ITO glass) 作為裝置之正負電極,方便置於螢光顯微鏡上觀察,另於ITO 層設計數種幾何結構,並以微機電製程製造出一體積小、易操作,且方便觀測之電穿刺晶片。透過表面蝕刻,可在用於抓取細胞之孔洞邊緣產生尖端狀之幾何結構,並產生電場放大效應,降低細胞電穿刺之電訊號源所需電壓值,使細胞不易因承受過大電壓而死亡,電穿刺後之細胞存活率可達近80 %。 本研究主要可分為兩部分,分別為細胞操縱與細胞電穿刺。首先,本研究開發之裝置將外接一電訊號源,以負介電泳力 (negative dielectrophoresis, nDEP) 使細胞進入蝕刻出的孔洞內,達成細胞操縱,再藉由調變電訊號強度達成細胞電穿刺之目的。在成功操縱並固定80 % 之細胞後,細胞將染以細胞增殖示蹤螢光試劑 (carboxyfluorescein diacetate succinimidyl ester, CFDA SE) 與鈣黃綠素 (calcein),驗證於電穿刺後,物質可由細胞內而外或由細胞外而內運輸,並找出電穿刺之最適結構設計。接著,將最適裝置接以摩擦奈米發電機,以達成自供電之細胞電穿刺,並以不同電源供應器進行shRNA 轉染,再將轉染後之樣本進行即時聚合酶鏈式反應 (real-time polymerase chain reaction, real-time PCR),以驗證裝置用於細胞轉染之效率。以本研究之電穿刺裝置進行之shRNA 轉染,能夠成功抑制60 % 之細胞基因表達。 | zh_TW |
dc.description.abstract | In this research, we use microelectromechanical system (MEMS) technology to fabricate devices with different geometric structures. The electric field amplification effect of the device structures is able to improve the efficiency of cell electroporation, and achieve simultaneous cell manipulation and electroporation. Triboelectric nanogenerator (TENG) can be used as an electrical signal source of the devices to achieve self-powered electroporation systems, and be used for shRNA transfection. To effectively achieve cell manipulation and electroporation, we use indium tin oxide glass (ITO glass) with highly transmittance as the positive and negative electrodes of the devices, which is convenient for observation with fluorescence microscope. In addition, several geometric structures are designed on the ITO layer, and electroporation chips which are small in volume, easy to operate and convenient to observe are manufactured by MEMS process. After surface etching, tip-shaped geometric structures can be generated at the edge of the holes used for capturing cells. The structures lead to an electric field amplification effect, which reduces the voltage value required for cell electropuncture, so that the cell won’t be injured due to excessive voltage, to improve cell viability. The cell survival rate after electroporation can reach nearly 80%. This research is divided into two parts, cell manipulation and cell electroporation. In the first part, the devices are connected to an external electrical signal source to make the cells captured into the etched holes with negative dielectrophoresis (nDEP) to achieve cell manipulation, and then achieve cell electroporation by modulating the strength of the electrical signal. After successfully manipulating and fixing 80% of the cells, we stain the cells with carboxyfluorescein diacetate succinimidyl ester (CFDA SE) and calcein to verify that after electroporation, substances can be delivered from inside the cell or from outside the cell, and to find the optimal structural design for electroporation. In the second part, we connect the optimal device to TENG to achieve self-powered cell electroporation, and perform shRNA transfection with different power supplies. After shRNA transfection, we subject transfected samples to real-time polymerase chain reaction (real-time PCR), to verify the efficiency of the device for cell transfection. 60% of the cells' gene expression is successfully inhibited after shRNA transfection. | en |
dc.description.provenance | Made available in DSpace on 2023-03-19T23:44:23Z (GMT). No. of bitstreams: 1 U0001-2608202216094200.pdf: 5938308 bytes, checksum: bf538f2070b1447c69bc20fcfb7b5f3d (MD5) Previous issue date: 2022 | en |
dc.description.tableofcontents | 致謝 I 摘要 II Abstract III 目錄 V 圖目錄 VIII 表目錄 XII 符號目錄 XIII 第一章 導論 1 1.1 前言 1 1.2 研究動機 1 1.3 論文架構 2 第二章 文獻回顧 3 2.1 細胞操縱 (Cell Manipulation) 3 2.1.1 細胞操縱方法 3 2.1.2 介電泳 5 2.2 細胞轉染 (Cell Transfection) 10 2.3 細胞電穿刺 (Cell Electroporation) 12 2.3.1 細胞膜特性 12 2.3.2 跨膜電位 13 2.3.3 電穿刺 13 2.4 摩擦奈米發電機 (Triboelectric Nanogenerator, TENG) 19 2.4.1 摩擦起電效應 19 2.4.2 摩擦奈米發電機 19 2.4.3 應用於電穿刺之摩擦奈米發電機 22 第三章 研究方法與系統設計 26 3.1 研究材料 26 3.2 實驗裝置設計與製程 28 3.2.1 裝置設計 28 3.2.2 裝置製程 34 3.3 細胞培養與染色 41 3.3.1 細胞培養 41 3.3.2 細胞染色 46 3.3.3 細胞轉染 50 3.4 實驗設備 52 3.4.1 光學系統 53 3.4.2 示波器 55 3.4.3 波形產生器 56 3.5 實驗步驟 57 3.5.1 細胞置入實驗裝置 57 3.5.2 細胞操縱實驗 57 3.5.3 細胞電穿刺實驗 58 3.5.4 細胞存活率檢測 59 3.5.5 即時定量聚合酶連鎖反應檢測 60 第四章 實驗結果與討論 63 4.1 細胞介電性質 63 4.2 細胞操縱 63 4.2.1 頻率與介電泳力之關係 63 4.2.2 介電泳力與細胞抓取率之關係 65 4.3 以波形產生器進行之細胞電穿刺 69 4.3.1 CFDA SE Cell Tracer Kit 70 4.3.2 陣列結構對細胞電穿刺之影響 72 4.3.3 Calcein 76 4.4 以摩擦奈米發電機進行之細胞電穿刺 77 4.5 細胞轉染shRNA 79 4.6 細胞存活率 83 4.6.1 以波形產生器進行之電穿刺 83 4.6.2 以摩擦奈米發電機進行之電穿刺 86 4.6.3 細胞轉染shRNA 87 第五章 結論與未來展望 90 5.1 結論 90 5.2 未來展望 91 參考文獻 92 附錄一 97 | |
dc.language.iso | zh-TW | |
dc.title | 用於細胞抓取並以摩擦奈米發電機遞送shRNA之微孔陣列開發 | zh_TW |
dc.title | Development of a Microwell Array for Cell Trapping and shRNA Delivery using Triboelectric Nanogenerator | en |
dc.type | Thesis | |
dc.date.schoolyear | 110-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 范育睿(Yu-Jui Fan),胡哲銘(Che-Ming Hu),沈湯龍(Tang-Long Shen),林宗宏(Zong-Hong Lin) | |
dc.subject.keyword | 微孔陣列,細胞抓取,電穿刺,細胞轉染,摩擦奈米發電機, | zh_TW |
dc.subject.keyword | Microwell array,Cell trapping,Electroporation,Transfection,TENG, | en |
dc.relation.page | 100 | |
dc.identifier.doi | 10.6342/NTU202202862 | |
dc.rights.note | 同意授權(全球公開) | |
dc.date.accepted | 2022-08-30 | |
dc.contributor.author-college | 工學院 | zh_TW |
dc.contributor.author-dept | 應用力學研究所 | zh_TW |
dc.date.embargo-lift | 2027-08-29 | - |
顯示於系所單位: | 應用力學研究所 |
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