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DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 黃念祖(Nien-Tsu Huang) | |
dc.contributor.author | Richard Lee Lai | en |
dc.contributor.author | 賴彥廷 | zh_TW |
dc.date.accessioned | 2021-06-17T02:21:26Z | - |
dc.date.available | 2021-08-17 | |
dc.date.copyright | 2020-08-24 | |
dc.date.issued | 2020 | |
dc.date.submitted | 2020-08-18 | |
dc.identifier.citation | 1 Phelan, M. C. J. C. p. i. n. Techniques for mammalian cell tissue culture. 38, A. 3B. 1-A. 3B. 19 (2007). 2 Lee, J., Cuddihy, M. J. Kotov, N. A. J. T. E. P. B. R. Three-dimensional cell culture matrices: state of the art. 14, 61-86 (2008). 3 Zheng, C., Chen, G. E., Pang, Y. Huang, Y. An integrated microfluidic device for long-term culture of isolated single mammalian cells. Science China Chemistry 55, 502-507, doi:10.1007/s11426-012-4493-1 (2012). 4 Wheeler, A. R. et al. Microfluidic device for single-cell analysis. 75, 3581-3586 (2003). 5 Turan, B. et al. A pillar-based microfluidic chip for T-cells and B-cells isolation and detection with machine learning algorithm. ROBOMECH Journal 5, 27, doi:10.1186/s40648-018-0124-8 (2018). 6 Park, J. Y. et al. Single-cell trapping in larger microwells capable of supporting cell spreading and proliferation. Microfluid Nanofluidics 8, 263-268, doi:10.1007/s10404-009-0503-9 (2010). 7 Huang, N.-T. et al. An integrated microfluidic platform for in situ cellular cytokine secretion immunophenotyping. Lab on a Chip 12, 4093-4101, doi:10.1039/C2LC40619E (2012). 8 Tsuchiya, S. et al. Establishment and characterization of a human acute monocytic leukemia cell line (THP‐1). 26, 171-176 (1980). 9 Mehling, M. Tay, S. J. C. o. i. B. Microfluidic cell culture. 25, 95-102 (2014). 10 Halldorsson, S., Lucumi, E., Gómez-Sjöberg, R. Fleming, R. M. T. Advantages and challenges of microfluidic cell culture in polydimethylsiloxane devices. Biosensors and Bioelectronics 63, 218-231, doi:https://doi.org/10.1016/j.bios.2014.07.029 (2015). 11 Huang, N.-T., Hwong, Y.-J. Lai, R. L. A microfluidic microwell device for immunomagnetic single-cell trapping. Microfluidics and Nanofluidics 22, 16, doi:10.1007/s10404-018-2040-x (2018). 12 Yang, H. Gijs, M. A. M. Micro-optics for microfluidic analytical applications. Chemical Society Reviews 47, 1391-1458, doi:10.1039/C5CS00649J (2018). 13 Huang, N.-T., Zhang, H.-l., Chung, M.-T., Seo, J. H. Kurabayashi, K. Recent advancements in optofluidics-based single-cell analysis: optical on-chip cellular manipulation, treatment, and property detection. Lab on a Chip 14, 1230-1245, doi:10.1039/C3LC51211H (2014). 14 Wang, X. et al. Enhanced cell sorting and manipulation with combined optical tweezer and microfluidic chip technologies. 11, 3656-3662 (2011). 15 Khoshmanesh, K. et al. Dielectrophoretic platforms for bio-microfluidic systems. 26, 1800-1814 (2011). 16 Lai, R. L. Huang, N.-T. Dimensional analysis and parametric studies of the microwell for particle trapping. Microfluidics and Nanofluidics 23, 121, doi:10.1007/s10404-019-2289-8 (2019). 17 Narayanamurthy, V., Nagarajan, S., Firus Khan, A. a. Y., Samsuri, F. Sridhar, T. M. Microfluidic hydrodynamic trapping for single-cell analysis: mechanisms, methods and applications. Analytical Methods 9, 3751-3772, doi:10.1039/C7AY00656J (2017). 18 Lee, P. J., Hung, P. J., Rao, V. M. Lee, L. P. Nanoliter scale microbioreactor array for quantitative cell biology. 94, 5-14, doi:10.1002/bit.20745 (2006). 19 Rettig, J. R. Folch, A. Large-Scale Single-Cell Trapping And Imaging Using Microwell Arrays. Analytical Chemistry 77, 5628-5634, doi:10.1021/ac0505977 (2005). 20 Leong, T. G., Randall, C. L., Benson, B. R., Zarafshar, A. M. Gracias, D. H. J. L. o. a. C. Self-loading lithographically structured microcontainers: 3D patterned, mobile microwells. 8, 1621-1624 (2008). 21 Kobel, S., Valero, A., Latt, J., Renaud, P. Lutolf, M. Optimization of microfluidic single-cell trapping for long-term on-chip culture. Lab on a Chip 10, 857-863, doi:10.1039/B918055A (2010). 22 Deng, B. et al. Parameter screening in microfluidics based hydrodynamic single-cell trapping. 2014 (2014). 23 Jin, D. et al. A microfluidic device enabling high-efficiency single-cell trapping. 9, 014101 (2015). 24 Zhou, Y. et al. A microfluidic platform for trapping, releasing and super-resolution imaging of single-cells. 232, 680-691 (2016). 25 Perillo, B. et al. ROS in cancer therapy: the bright side of the moon. Experimental Molecular Medicine 52, 192-203, doi:10.1038/s12276-020-0384-2 (2020). 26 Liou, G.-Y. Storz, P. Reactive oxygen species in cancer. Free Radic Res 44, 479-496, doi:10.3109/10715761003667554 (2010). 27 Simon, H.-U., Haj-Yehia, A. Levi-Schaffer, F. J. A. Role of reactive oxygen species (ROS) in apoptosis induction. 5, 415-418 (2000). 28 Pelicano, H., Carney, D. Huang, P. J. D. R. U. ROS stress in cancer cells and therapeutic implications. 7, 97-110 (2004). 29 Nogueira, V. Hay, N. Molecular pathways: reactive oxygen species homeostasis in cancer cells and implications for cancer therapy. Clin Cancer Res 19, 4309-4314, doi:10.1158/1078-0432.CCR-12-1424 (2013). 30 Lo, K.-Y., Wu, S.-Y. Sun, Y.-S. A microfluidic device for studying the production of reactive oxygen species and the migration in lung cancer cells under single or coexisting chemical/electrical stimulation. Microfluidics and Nanofluidics 20, 15, doi:10.1007/s10404-015-1683-0 (2016). 31 Ayuso, J. M. et al. Development and characterization of a microfluidic model of the tumour microenvironment. Scientific Reports 6, 36086, doi:10.1038/srep36086 (2016). 32 Moussavi-Harami, S. F. et al. Microfluidic device for simultaneous analysis of neutrophil extracellular traps and production of reactive oxygen species. Integr Biol (Camb) 8, 243-252, doi:10.1039/c5ib00225g (2016). 33 Križaj, D. et al. From mechanosensitivity to inflammatory responses: new players in the pathology of glaucoma. Current eye research 39, 105-119, doi:10.3109/02713683.2013.836541 (2014). 34 Chu, S.-H. et al. A microfluidic device for in situ fixation and super-resolved mechanosensation studies of primary cilia. 13, 014105 (2019). 35 Marshall, K. L. Lumpkin, E. A. The molecular basis of mechanosensory transduction. Adv Exp Med Biol 739, 142-155, doi:10.1007/978-1-4614-1704-0_9 (2012). 36 Zablotskii, V., Syrovets, T., Schmidt, Z. W., Dejneka, A. Simmet, T. J. B. Modulation of monocytic leukemia cell function and survival by high gradient magnetic fields and mathematical modeling studies. 35, 3164-3171 (2014). 37 Zablotskii, V., Polyakova, T., Lunov, O. Dejneka, A. How a High-Gradient Magnetic Field Could Affect Cell Life. Scientific Reports 6, 37407, doi:10.1038/srep37407 (2016). 38 Zablotskii, V. et al. Life on magnets: stem cell networking on micro-magnet arrays. 8 (2013). 39 Zablotskii, V. et al. Down-regulation of adipogenesis of mesenchymal stem cells by oscillating high-gradient magnetic fields and mechanical vibration. 105, 103702 (2014). 40 Miyakoshi, J. J. P. i. b. biology, m. Effects of static magnetic fields at the cellular level. 87, 213-223 (2005). 41 Sabo, J. et al. Effects of static magnetic field on human leukemic cell line HL-60. 56, 227-231 (2002). 42 Hernández-Hernández, H., Cruces-Solis, H., Elías-Viñas, D. Verdugo-Díaz, L. J. A. o. m. r. Neurite outgrowth on chromaffin cells applying extremely low frequency magnetic fields by permanent magnets. 40, 545-550 (2009). 43 Santoro, N. et al. Effect of extremely low frequency (ELF) magnetic field exposure on morphological and biophysical properties of human lymphoid cell line (Raji). 1357, 281-290 (1997). 44 Venkatachalam, K. Montell, C. J. A. R. B. TRP channels. 76, 387-417 (2007). 45 Michalick, L. Kuebler, W. M. TRPV4—A Missing Link Between Mechanosensation and Immunity. 11, doi:10.3389/fimmu.2020.00413 (2020). 46 Yin, J. et al. Role of Transient Receptor Potential Vanilloid 4 in Neutrophil Activation and Acute Lung Injury. 54, 370-383, doi:10.1165/rcmb.2014-0225OC (2016). 47 Hirt, C. W., Amsden, A. A. Cook, J. An arbitrary Lagrangian-Eulerian computing method for all flow speeds. Journal of computational physics 14, 227-253 (1974). 48 Mustin, B. Stoeber, B. Low cost integration of 3D-electrode structures into microfluidic devices by replica molding. Lab on a Chip 12, 4702-4708, doi:10.1039/C2LC40728K (2012). 49 Munson, B. R., Okiishi, T. H., Huebsch, W. W. Rothmayer, A. P. Fluid mechanics. (Wiley Singapore, 2013). 50 Freund, E. et al. Plasma-derived reactive species shape a differentiation profile in human monocytes. 9, 2530 (2019). 51 Genin, M., Clement, F., Fattaccioli, A., Raes, M. Michiels, C. M1 and M2 macrophages derived from THP-1 cells differentially modulate the response of cancer cells to etoposide. BMC Cancer 15, 577-577, doi:10.1186/s12885-015-1546-9 (2015). 52 Haschek, W. M., Rousseaux, C. G. Wallig, M. A. in Fundamentals of Toxicologic Pathology (Second Edition) (eds Wanda M. Haschek, Colin G. Rousseaux, Matthew A. Wallig) 9-42 (Academic Press, 2010). | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/68450 | - |
dc.description.abstract | 在本論文中,我們提出一個具有多個微流井的微流道晶片進行懸浮型細胞 (單核細胞株,THP-1) 的捕捉及長時間的培養,並觀察它們在不同磁場刺激條件下受到的影響。首先,我們透過數值模擬來最佳化單細胞捕捉的微流井構造。模擬的參數包含刮長寬比 (W/L) 、寬深寬比 (D/W) 以及細胞尺寸跟微流井長度的比例 (R/L) 。初步結果顯示,當W/L越大的時候,捕捉的效率會越高,而當D/W越大時,細胞比較不會被沖走。R/L則是越小越容易被捕捉。接著我們利用細胞進行實驗,證明當 W/L = 2、D/W = 0.5 跟 R/L = 0.125 時,單細胞捕捉效率(細胞捕捉率跟單細胞捕捉率的乘積)是最高的。因此,我們選擇 W/L = 2 微流井構造來行磁場刺激實驗。找到最佳的細胞導植入濃度以後,我們將進行不同種類磁場的刺激並且判斷它們對THP-1細胞的影響。結果顯示,梯度為 2x10^2 T/m 的高磁場梯度的條件下會增加細胞ROS 的分泌,過了36小時甚至比沒有施加磁場的情況多了兩倍。該刺激同時也會造成細胞生長的抑制 (比控制組低了25%) 以及真圓度的下降 (由控制組的0.9下降為0.8)。這是因為有梯度的磁場會對細胞膜造成剪應力,並影響到負責調控ROS濃度的機械感應通道TRPV4。因此,我們的晶片可以用來進行大量單細胞捕捉以及精準的磁場梯度引發之機械刺激。 | zh_TW |
dc.description.abstract | We aim to develop a microfluidic platform and the accompanying image processing algorithms for the long-term cultivation and observation of the suspension cell line types (monocytes, THP-1) under different physiological conditions. First, we conduct simulations to determine the optimal microwell geometry for single single-cell trapping. Parameters such as width to cross-sectional length (W/L), width to depth (D/W) and radius to cross-sectional length ratio (R/L) ratios are discussed in our study. Next, we conduct actual experiments to verify our simulations. Triangular microwells with W/L = 1 and W/L = 2 have single cellsingle-cell occupancy rates (given as the product of the single cellsingle-cell trapping rate and well occupancy) of 16.7% and 35.8%, respectively. Therefore, we choose W/L = 2 and D/W=0.5 for our purposes. For these two values, the ideal R/L ratio is found to be around 0.125. After finding the optimal cell loading conditions, we conduct experiments under different magnetic field gradients and determine their effect on THP-1 cell culture. Results show that a high magnetic field gradient on the order of 2x10^2 T/m would increases the concentration of Reactive Oxygen Species (ROS) secretion by 200% over 36 hours. It also results in both decreased cell size (reduction of 25% compared with the control group) and circularity (0.8 compared with 0.9 for control). This is because magnetic field gradients induce shear and pressure forces on the cellular membrane, affecting the mechanosensitive ion channel TRPV4, which is responsible for the regulation of ROS concentrations. Therefore, our chip can be used in high-throughput single cellsingle-cell trapping and provide precise magnetic field gradient induced mechanical stimulus. | en |
dc.description.provenance | Made available in DSpace on 2021-06-17T02:21:26Z (GMT). No. of bitstreams: 1 U0001-1708202013562400.pdf: 4885609 bytes, checksum: 97a5db803cc2d63442a363ec1f491669 (MD5) Previous issue date: 2020 | en |
dc.description.tableofcontents | 中文摘要……………………………………………………………...viii Abstract………………………………………………………………...ix Chapter 1 Introduction..……………………………………………1 1.1 Background………………..…………………………………………….1 1.2 Literature Review..……………………………………………………...2 1.2.1 Microfluidics in Long-Term Cell Culture…………..………………...2 1.2.2 Single-Cell Trapping…...………………………………...…………...2 1.2.3 Reactive Oxygen Species …………...…………………………….......4 1.2.4 Magnetic Forces……………………………………………………....5 1.2.5 Relationship Between TRP channels and ROS……………………….7 1.3 Research Motivation …………………..………………………………..9 1.4 Thesis Structure………………………...…………………………….....9 Chapter 2 Microfluidic Design and Simulation…...…….………..10 2.1 Governing Equations………………………………………………….10 2.1.1 Governing Equations for Fluid Flow………………………………...10 2.2 Optimization of the Main Channel……………………………………12 2.3 Optimization of Microwells …………………………………………...13 2.3.1 Streamline Simulation……………………………………………….15 2.3.2 Aspect Ratio and Particle Position…………………………………...17 2.3.3 Effect of R/L on Particle Trapping…………………………………..19 2.3.4 Effect of Well Depth on Trapping……………………………………21 2.4 Magnetic Field Simulation Results……………………………………22 Chapter 3 Device Fabrication and Experimental Setup…………24 3.1 Microfluidic Device Fabrication………………………………………24 3.1.1 Photolithography……………………………………………………..24 3.1.2 Soft lithography…………………………...…………………………25 3.2 Magnet Fabrication……………………………………………………27 3.3 Experimental Design and Setup………………………………………30 3.3.1 Cell Culture Conditions……………………………………………...30 3.3.2 THP-1 Cell Growth in microfluidics………………………………...31 3.3.3 Image Acquisition…………………………………………………...32 3.4 Image Processing Techniques…………………………………………33 3.4.1 ROS Signal Processing………………………………………………33 3.4.2 Effects of Magnetic Fields on Cell Size and Circularity…………….34 Chapter 4 Results and Discussion………………………….36 4.1 Microbead Trapping Experiments……………………………………36 4.1.1 Effect of Sheath Flow on Trapping…………………………………..36 4.1.2 Effect of Microwell Geometry on Trapping efficiency………………37 4.2 Cell Trapping Experiments…………………………………………...39 4.2.1 Single-cell Trapping…………………………………………………39 4.2.2 Cell Loading and Preparation………………………………………..40 4.3 Effect of Magnetic Fields on Cell Size and Circularity………………42 4.4 Effect of Magnetic Fields on ROS…………………………………….44 Chapter 5 Conclusions…………………………………………….46 Chapter 6 Future Work…………………………………………...47 Chapter 7 Appendix……………………………………………….49 7.1 Simulation Protocol…………………..………………………………..49 7.1.1 Introduction………………………………………………………….49 7.1.2 Calculation of the Magnetic Field……………………………………49 7.2 Cell Culture and Experiment Protocol……………………………….50 7.2.1 Cell Culture………………………………………………………….50 7.2.2 Chip Preparation……………………………………………………..50 7.2.3 Cell Loading…………………………………………………………50 7.3 Supplementary for Cell Morphology and ROS………………………51 References……………………………………………………………..52 | |
dc.language.iso | en | |
dc.title | 流體動力學式之細胞捕捉及磁場刺激微流井微流道晶片 | zh_TW |
dc.title | A Microwell Microfluidic Device for Cell Trapping and Magnetic Gradient Stimulation | en |
dc.type | Thesis | |
dc.date.schoolyear | 108-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 林致廷(Chih-Ting Lin),楊東霖(Tony Yang),盧彥文(Yen-Wen Lu) | |
dc.subject.keyword | 微流道,單細胞捕捉, | zh_TW |
dc.subject.keyword | Microfluidics,Single cell trapping, | en |
dc.relation.page | 54 | |
dc.identifier.doi | 10.6342/NTU202003738 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2020-08-19 | |
dc.contributor.author-college | 電機資訊學院 | zh_TW |
dc.contributor.author-dept | 生醫電子與資訊學研究所 | zh_TW |
顯示於系所單位: | 生醫電子與資訊學研究所 |
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