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Design and fabrication of microfluidic chip for studying cell motility
|關鍵字:||microfluidics;微流道;metastasis;cell motility;cancer cell migration;cell sorting;轉移;細胞運動性;細胞遷移;細胞篩選||出版社:||機械工程學系所||引用:||1. Sporn, M.B., The war on cancer. Lancet, 1996. 347(9012): p. 1377-81. 2. Clarke, M.F., et al., Cancer stem cells--perspectives on current status and future directions: AACR Workshop on cancer stem cells. Cancer Res, 2006. 66(19): p. 9339-44. 3. Brabletz, T., et al., Opinion: migrating cancer stem cells - an integrated concept of malignant tumour progression. Nat Rev Cancer, 2005. 5(9): p. 744-9. 4. Reya, T., et al., Stem cells, cancer, and cancer stem cells. Nature, 2001. 414(6859): p. 105-11. 5. Decaestecker, C., et al., Can anti-migratory drugs be screened in vitro? A review of 2D and 3D assays for the quantitative analysis of cell migration. Medicinal Research Reviews, 2007. 27(2): p. 149-176. 6. Kam, Y., et al., A novel circular invasion assay mimics in vivo invasive behavior of cancer cell lines and distinguishes single-cell motility in vitro. BMC Cancer, 2008. 8: p. 198. 7. Tavana, H., et al., Rehydration of Polymeric, Aqueous, Biphasic System Facilitates High Throughput Cell Exclusion Patterning for Cell Migration Studies. Advanced Functional Materials, 2011. 21(15): p. 2920-2926. 8. Goetsch, K.P. and C.U. Niesler, Optimization of the scratch assay for in vitro skeletal muscle wound healing analysis. Anal Biochem, 2011. 411(1): p. 158-60. 9. Irimia, D. and M. Toner, Spontaneous migration of cancer cells under conditions of mechanical confinement. Integr Biol (Camb), 2009. 1(8-9): p. 506-12. 10. Mak, M., C.A. Reinhart-King, and D. Erickson, Microfabricated physical spatial gradients for investigating cell migration and invasion dynamics. PLoS One, 2011. 6(6): p. e20825. 11. Murrell, M., R. Kamm, and P. Matsudaira, Tension, free space, and cell damage in a microfluidic wound healing assay. PLoS One, 2011. 6(9): p. e24283. 12. Whitesides, G.M., The origins and the future of microfluidics. Nature, 2006. 442(7101): p. 368-73. 13. Burns, M.A., et al., An integrated nanoliter DNA analysis device. Science, 1998. 282(5388): p. 484. 14. Manz, A., N. Graber, and H.M. Widmer, Miniaturized total chemical analysis systems: a novel concept for chemical sensing. Sensors and actuators B: Chemical, 1990. 1(1-6): p. 244-248. 15. Nie, F.Q., et al., On-chip cell migration assay using microfluidic channels. Biomaterials, 2007. 28(27): p. 4017-4022. 16. Kamholz, A.E., et al., Quantitative analysis of molecular interaction in a microfluidic channel: the T-sensor. Analytical Chemistry, 1999. 71(23): p. 5340-5347. 17. Keenan, T.M. and A. Folch, Biomolecular gradients in cell culture systems. Lab Chip, 2008. 8(1): p. 34-57. 18. Saadi, W., et al., A parallel-gradient microfluidic chamber for quantitative analysis of breast cancer cell chemotaxis. Biomed Microdevices, 2006. 8(2): p. 109-18. 19. Keenan, T.M., C.H. Hsu, and A. Folch, Microfluidic “jets” for generating steady-state gradients of soluble molecules on open surfaces. Applied physics letters, 2006. 89: p. 114103. 20. Shamloo, A., et al., Endothelial cell polarization and chemotaxis in a microfluidic device. Lab Chip, 2008. 8(8): p. 1292-1299. 21. Cailleau, R., M. Olive, and Q.V. Cruciger, Long-term human breast carcinoma cell lines of metastatic origin: preliminary characterization. In Vitro, 1978. 14(11): p. 911-5. 22. Gozgit, J.M., et al., Use of an aggressive MCF-7 cell line variant, TMX2-28, to study cell invasion in breast cancer. Mol Cancer Res, 2006. 4(12): p. 905-13. 23. Wolfe, D.B., D. Qin, and G.M. Whitesides, Rapid prototyping of microstructures by soft lithography for biotechnology. Methods Mol Biol, 2010. 583: p. 81-107. 24. 黃明宏, 聚二甲基矽氧烷應用於可撓液晶顯示器的研究, 2009, 撰者. 25. Tong, Z., et al., Chemotaxis of cell populations through confined spaces at single-cell resolution. PLoS One, 2012. 7(1): p. e29211. 26. Morimura, S. and K. Takahashi, Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I. Int J Cell Biol, 2011. 2011: p. 615912. 27. http://www.atcc.org/Attachments/25935.jpg 28. http://www.atcc.org/Attachments/1980.jpg||摘要:||
本論文將呈現一種研究細胞運動性的微流晶片。目前細胞運動性的研究包括wound healing assay和微流裝置都有些限制。例如wound healing assay可藉由觀察細胞兩個不同時間點得知細胞運動性，但卻無法精確的操作細胞，也無法避免群體細胞之間的相互影響。近年來文獻中提到的一些微流道的方式可以精確的操作細胞並觀察單一細胞的運動性，但仍然無法給予細胞相同的起始條件，以達到更精確、客觀的實驗分析；也無法將不同運動性的細胞篩選並且取出微流道，做不同運動性細胞的後續分析和研究。
本研究為細胞運動性研究設計了一種新的微流晶片。首先，利用migration area與draining channel之間的液壓差將細胞排列成一直線，為細胞創造出相同的起始條件。第二，微流晶片中的每一顆細胞都被觀察和分析。第三，利用collection channels創造出的層流將微流晶片中的細胞按照其位置不同分別被收集出微流晶片。第四，創造濃度梯度場達成細胞趨化研究的功能。本研究提供了一個單一細胞層級的細胞運動性分析平台。
In this thesis, we present a microfluidic chip for studying cell motility. There are some limitations for current cell motility research including wound healing assay and microfluidic device. For example, wound healing assay could observe two different time point of cultured cells. However, it is not able to manipulate cells precisely and avoid interaction between groups of cells in would healing assay. Recently, some reports in the literature could handle many single cells precisely in their microfludic devices. However, those devices still were not able to provide same initial conditions for cells to study motility. Also, those devices could not sort and collect cells based on their motility for the follow-up research.
In this research, a novel microfluidic device was developed to study cell motility. Firstly, cells in the microfluidic chip were able to align in a single line to create same initial condition for cell motility by different hydraulic pressure between migration area and draining channel. Secondly, motility of each cell was observed and analyzed. Thirdly, cells were collected based on their different position in this device by laminar flow created by collection channels. Fourthly, concentration gradient was formed for chemotaxis research. A new platform for cell motility analysis in single cell level was developed.
In this research, human breast cancer cells MDA-MB-231 and MCF-7 are used as samples to test motion of cells. Same initial condition for cells by align cells in one single line was achieved. Single-cell-level spontaneous migration was analyzed and quantified precisely. Velocities and rates of cell motility data were measured and analyzed. Cells were collected to different culture dishes based on their different position in microfluidic chip. Cell recovery rate was quantified. Concentration gradient was formed for chemotaxis research. A platform for cell motility research in single cell level was performed. The functions of microfluidic chip for cell alignment, cell motility analysis, cell collection and gradient formation were demonstrated.
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