臺灣博碩士論文加值系統

本研究對三維介電泳微流體晶片於血漿中生物標誌分子(前列腺特異抗原)電
動行為作系統化的數值模擬。粒子使用前列腺特異抗原，直徑約 10 nm 溶液使用
核酸電泳緩衝液 (Tris-Borate-EDTA)以及血漿，血漿以市售之動物血液放入離心機
分離製作。微流道晶片則利用微機電製程製作，基板為載玻片利用濕式蝕刻玻璃製
作，上蓋使用雕刻機，雕刻壓克力板並使用聚二甲基矽氧烷(PDMS)翻模製作，最
後將玻璃基板與 PDMS 使用氧電漿清洗機進行表面改質接合完成晶片。首先利用
多重物理量耦合軟體 COMSOL Multiphysics 對微流道執行電動力學模擬，模擬電
場強度、電滲流、高頻/低頻介電泳力分析。模擬結果顯示在電場梯度方面施予交
流電 75 Vpp，頻率 15 kHz 時流道隘口處的局部電場達到 10^6 V/m，電滲流方面速度
達到 24.7 µm/s，並得知隘口間距越小電場越大電滲流速度也越高。介電泳力方面
單純施加交流電時，在高頻域時產生負向介電泳力，在低頻域時產生正向介電泳力，
另外再多施加直流電 0.5 V 時，粒子則濃縮至流道隘口右側形成分子壩。實驗結果
顯示在前列腺特異抗原在血漿當中時，給予交流電 75 Vpp 與 15 kHz 時則在 55 秒
內濃縮至 10^6 倍在隘口處，此三維微流道比起傳統二維微流道有更大的反應體積及
高達 30000 倍的截面比具有高電場濃縮倍率。

This study systematically simulates the electrokinetic behavior of a 3D microfluidic dielectrophoresis chip for the concentration of biomarker molecules (prostate specific antigen, PSA) in plasma. PSA molecules with a diameter of 10 nm are diluted in a nucleic acid electrophoresis buffer solution (Tris-Borate-EDTA) and a plasma solution. The plasma is prepared by placing commercially available animal blood into a centrifuge for separation. The microchannel chip is fabricated using micro-electro-mechanical systems (MEMS) technology.The substrate is a glass microscope slide that undergoes wet etching. The upper cover is carved from an acrylic board using CNC micromachining and then coated with polydimethylsiloxane (PDMS). Finally, the glass substrate and PDMS are
bonded together using surface modification with an oxygen plasma cleaner to create the chip. Firstly, the simulation of electrokinetic microchannels is conducted using the COMSOL Multiphysics software. These simulations include the examination of electric field intensity, electroosmotic flow, and high/low-frequency dielectrophoresis forces. The simulation results show that the local electric field at the contraction microchannel reaches 106 V/m, and the electroosmotic velocity reaches 24.7 μm/s when the AC field intensity is 75 Vpp with a frequency of 15 kHz. The smaller the contraction, the greater the electric field intensity, and the higher the electroosmotic velocity. For dielectrophoresis force, when the AC field is applied, a negative dielectrophoresis force is generated in the high-frequency domain, while a positive dielectrophoresis force is generated in the low-frequency domain. When an additional AC field of 0.5 V/cm is applied, the particles will concentrate on the right side of the contraction microchannel, forming a molecular dam. The test results demonstrate a 10^6-fold increase in PSA concentration within 55 seconds after injecting it into the plasma. This phenomenon occurs when the system is exposed to an alternating current (AC) field with an intensity of 75 Vpp and a frequency of 15 kHz. Compared to traditional 2D microchannels, 3D
microchannels offer larger reaction volumes, a cross-section ratio up to 30,000 times higher, and a higher concentration ratio in the electric field.

摘要............................................................... i
Abstract...........................................................ii
誌謝.............................................................. iv
目錄............................................................... v
表目錄............................................................ vii
圖目錄.............................................................viii
符號說明........................................................... xii
緒論................................................................ 1
1.1 前言............................................................ 1
1.2 研究動機與目的................................................... 1
1.3 文獻回顧........................................................ 2
1.4 論文架構........................................................ 9
實驗設備與材料介紹.................................................. 10
2.1 製程設備....................................................... 10
2.2 氧電漿接合實驗設備.............................................. 13
2.3 正立式螢光系統顯微鏡介電泳實驗之設備 ............................ 13
2.4 SEM........................................................... 17
2.5 實驗材料介紹................................................... 17
電動力學........................................................... 22
3.1 電泳(Electrophoresis)......................................... 22
3.2 電偶極化(Dipole).............................................. 23
3.3 介電泳(Dielectrophoresis, DEP)................................ 25
3.4 電滲流(Electroosmosis flow, EOF).............................. 27
3.5 交直流電場合力................................................. 28
晶片製作與模擬分析................................................. 29
4.1 晶片製作...................................................... 29
4.2 玻璃黃光微影製程............................................... 31
4.3 曝光與顯影.................................................... 32
4.4 玻璃基板蝕刻與去光阻........................................... 33
4.5 壓克力加工與 PDMS 晶片製作..................................... 33
4.6 玻璃基板與 PDMS 改質接合....................................... 35
4.7 數值模型建模.................................................. 36
4.8 電場梯度...................................................... 38
4.9 電流......................................................... 40
4.10 介電泳力.................................................... 41
4.11 交流電與直流電模擬結果....................................... 44
實驗結果與討論.................................................... 48
5.1 實驗步驟..................................................... 48
5.2 蝕刻玻璃基板................................................. 49
5.3 實驗架構..................................................... 50
5.4 前列腺特異抗原螢光測試........................................ 51
5.5 介電泳實驗................................................... 53
結論與未來展望................................................... 55
6.1 結論........................................................ 55
6.2 未來展望.................................................... 55
參考文獻......................................................... 56
Extended Abstract .............................................. 61

[1] R. Martinez‐Duarte, P. Renaud, and M. J. Madou, “A novel approach to dielectrophoresis using carbon electrodes,” Electrophoresis,Vol 32, no 17, pp. 2385-2392, 2011
[2] L. Wang, L. A. Flanagan, N. L. Jeon, E. Monuki, and A. P. Lee,
“Dielectrophoresis switching with vertical sidewall electrodes for
microfluidic flow cytometry,” Lab Chip, Vol. 7, no. 9, pp. 1114-1120, 2007.
[3] R. Martinez-Duarte, R. A. Gorkin, K. Abi-Samrab, and M. J. Madou,
“The integration of 3D carbon-electrode dielectrophoresis on a CD-like
centrifugal microfluidic platform,” Lab Chip, Vol. 10, no. 8, pp.1030, 2010.
[4] V. Chaurey, A. Rohani, Y. H. Su, K. T. Liao, and C. F. Chou, “Scaling
down constriction‐based (electrodeless) dielectrophoresis devices for
trapping nanoscale bioparticles in physiological media of high‐
conductivity,” Electrophoresis, Vol 34, no 7, pp. 1097-1104, 2013.
[5] P. K. Thwar, J. J. Linderman, and M. A. Burns, “Electrodeless direct
current dielectrophoresis using reconfigurable field‐shaping oil barriers,”
Electrophoresis , Vol 28. no 24, pp. 4572-4581, 2007.
[6] S. V. Puttaswamy, C. H. Lin, S. Sivashankar,Y. S. Yang, and C. H.
Liu,“Electrodeless dielectrophoretic concentrator for analyte preconcentration on poly-silicon nanowire field effect transistor,” Sensors Actuators B Chem, Vol. 178, pp. 547-554, 2013.
[7] A. Kale, L. Song, X. Lu, L. Yu, G. Hu, and X. Xuan, “Electrothermal
enrichment of submicron particles in an insulator‐based dielectrophoretic microdevice,” Electrophoresis, Vol 39, Issue 5-6, pp. 887-896, 2018.
[8] S. Park, M. Koklu, and A. BeskoK, “Particle trapping in highconductivity media with electrothermally enhanced negative dielectrophoresis,” Anal. Chem, Vol. 81, no. 6, pp. 2303-2310, 2009.
[9] Z. Ma, J. Guo, Y. J. Liu, and Y. Ai, “The patterning mechanism of carbon
nanotubes using surface acoustic waves: The acoustic radiation effect or
the dielectrophoretic effect,” Nanoscale, Vol. 7, pp. 14047-14054, 2015.
[10]H. Y. Wang, C. Y. Chen, P. Y. Chu, Y. X. Zhu, C. H. Hsieh, J. J. Lu, and
M. H. Wu, “Application of an optically induced dielectrophoresis (ODEP)-based microfluidic system for the detection and isolation of bacteria with heterogeneity of antibiotic susceptibility,” Sens. Actuators B Chem, Vol. 307, pp.127540, 2020.
[11]T. K. Chiu, A. C. Chao, W. P. Chou, C. J. Liao, H. M. Wang, J. H. Chang,
P. H. Chen, and M. H. Wu, “Optically-induced-dielectrophoresis
(ODEP)-based cell manipulation in a microfluidic system for highpurity isolation of integral circulating tumor cell (CTC) clusters based
on their size characteristics,” Sens. Actuators B Chem, Vol. 258, pp. 1161-1173, 2018.
[12]X. Jiang, Y. Zhou, Y. Chen, Y. Shao, and J. Feng, “Etching-engineered low-voltage dielectrophoretic nanotweezers for trapping of single molecules,” Analytical Chemistry, Vol. 93.no. 37, pp.12549-12555,2021.
[13]P. Y. Chu, C. H. Hsieh, and M. H. Wu, “The Combination of immunomagnetic bead-based cell isolation and optically induced dielectrophoresis (ODEP)-based microfluidic device for the negative selection-based isolation of circulating tumor cells (CTCs),” Frontiers in Bioengineering and Biotechnology, Vol 8, no 921, 2020.
[14]C. J. Liao, C. H. Hsieh, T. K. Chiu, Y. X. Zhu, and H. M. Wang, “An optically induced dielectrophoresis (ODEP)-based microfluidic system
for the isolation of high-purity CD45neg/EpCAMneg cells from the blood samples of cancer patients—demonstration and initial exploration of the clinical significance of these cells,” Micromachines, Vol 9, no 11, pp.563, 2018.
[15]Y. J. Eo, G. Y. Yoo, H. Kang, Y. K. Lee, C. S. Kim, and Y. R.Do,“Enhanced DC-Operated Electroluminescence of Forwardly Aligned p/MQW/n InGaN Nanorod LEDs via DC Offset-AC Dielectrophoresis,” ACS applied materials & interfaces,Vol 9, no 43, pp.
37912-37920, 2017.
[16]H. A. Pohl, “The Motion and precipitation of suspensoids in divergent
electric fields,” Appl. Phys, Vol. 22, no. 7, pp. 869-871, 1951.
[17]X. Wang, Y. Xin, L. Ren, Z. Sun, P. Zhu, Y. Ji, C. Li, J. Xu, and B. O.
MA, “Positive dielectrophoresis–based Raman-activated droplet sorting
for culture-free and label-free screening of enzyme function in
vivo,” Science advances,vol 6, no 32,eabb3521, 2020.
[18]P. Y. Chu, C. J. Liao, C. H. Hsieh, H. M. Wang, W. P. Chou, P. H. Chen,
and M. H. Wu, “Utilization of optically induced dielectrophoresis in a
microfluidic system for sorting and isolation of cells with varied degree
of viability: Demonstration of the sorting and isolation of drug-treated
cancer cells with various degrees of anti-cancer drug resistance gene
expression,” Sensors and Actuators B: Chemical, Vol 283, pp. 621-631,
2019.
[19]Y. Takahashi, and S. Miyata, “Continuous ES/feeder cell-sorting device
using dielectrophoresis and controlled fluid flow,” Micromachines, Vol
11. no 8, pp. 734, 2020.
[20]T. Shibuya, S. Uchida, Y. Ishii, and H. Nishikawa et al, “Assembling
gold nanoparticles by dielectrophoresis with pit arrays on PMMA fabricated by proton beam writing,” Nuclear Instruments and Methods in Physics Research Section B: Beam Interactions with Materials and
Atoms, Vol 456, pp. 60-63, 2019.
[21]T. Shibuya, S. Uchida, Y. Ishii, and H. Nishikawa, “Trapping of a Single
Microparticle Using AC Dielectrophoresis Forces in a Microfluidic
Chip,” Micromachines, Vol 14, no 1, pp. 59, 2023.
[22]K. T. Liao, and C. F. Chou, “Nanoscale molecular traps and dams for
ultrafast protein enrichment in high-conductivity buffers,” Journal of the
American Chemical Society, Vol 134, no 21 , pp. 8742-8745, 2012.
[23]A. Sonnenberg, J. Y. Marciniak, E. A. Skowronski, S. Manouchehri, L.
Rassenti, E. M. Ghia, F. Widhopf, T. J. Kipps, and M. J. Heller,
“Dielectrophoretic Isolation and Detection of Cancer Related Circulating Cell Free DNA Biomarkers from Blood and Plasma,” Electrophoresis, Vol. 35, pp. 1828-1836, 2014.
[24]M. Mohammadi, H. Madadi, J. Casals-Terré, and J. Sellarés,
“Hydrodynamic and direct-current insulator-based dielectrophoresis (HDC-iDEP) microfluidic blood plasma separation,” Analytical and bioanalytical chemistry, Vol 407, pp. 4733-4744, 2015.
[25]A. Nakano, F. Camacho-Alanis, and A. Ros, “Insulator-based dielectrophoresis with β-galactosidase in nanostructured devices,”Analyst, Vol 140. no 3, pp. 860-868, 2015.
[26]M. Lin, and R. K. Anand , “High-throughput selective capture of single
circulating tumor cells by dielectrophoresis at a wireless electrode array,” Journal of the American Chemical Society, Vol 139, no 26, pp.8950-8959, 2017.
[27]J. Zhang, D. Yuan, Q. Zhao, S. Yan, and S. Y. Tang et al, “Tunable
particle separation in a hybrid dielectrophoresis (DEP)-inertial microfluidic device,” Sensors and Actuators B: Chemical, Vol 267, pp.14-25, 2018.
[28]Mohamed Zackria Ansar B .I, M. Y. Caffiyar, and M. K. A, “Microfluidic
device for Multitarget separation using DEP techniques and its applications in clinical research,” In: 2020 Sixth International Conference on Bio Signals, Images, and Instrumentation (ICBSII). IEEE,pp. 1-6, 2020.
[29]M. Hata, M. Suzuki, and T. Yasukawa, “Selective retrieval of antibodysecreting hybridomas in cell arrays based on the dielectrophoresis,”
Biosensors and Bioelectronics, Vol 209, no 114250, 2022.
[30]S. P. Balk, Y. J. Ko, and G. J. Bubley, “Biology of prostate-specific
antigen,” Journal of clinical oncology, Vol 21, no 2, pp.383-391, 2003.
[31]W. J. Catalona, D. S. Smith, and T. L. Ratliff et al, “Measurement of
prostate-specific antigen in serum as a screening test for prostate
cancer,” New England journal of medicine, Vol 324, no 17, pp. 1156-
1161, 1991.
[32]李振羽, “無電極式介電泳晶片交流電動現象之研究,” 義守大學機械與自動化工程學系碩士在職專班碩士論文. 2011.
[33]W. Zhang, H. Wu, R. Zhang, X. Fang, and W. Xu, “Structure and effective charge characterization of proteins by a mobility capillary electrophoresis based method,” Chemical science, Vol 10, no 33, pp.7779-7787, 2019.
[34]沈文駿, “單根氧化釩奈米線的電性及光學性質研究,” 國立交通大學應用化學研究所碩士班碩士論文, 2010.
[35]B. J. Sanghavi, W. Varhue, J. L. Chávez, C. F. Chou, and N. S. Swami,“Electrokinetic preconcentration and detection of neuropeptides at patterned graphene-modified electrodes in a nanochannel.” Analytical chemistry, Vol 86, no 9, pp. 4120-4125, 2014.
[36]A. Rohani, W. Varhue, K. T. Liao, C. F. Chou, and Swami.N. S, “Nanoslit
design for ion conductivity gradient enhanced dielectrophoresis for ultrafast biomarker enrichment in physiological media,”Biomicrofluidics, Vol 10, no 3, 033109, 2016.
[37]A. Rohani, B. J. Sanghavi, A. Salahi, K. T. Liao, and C. F. Chou,Swami.N. S, “Frequency-selective electrokinetic enrichment of biomolecules in physiological media based on electrical double-layer polarization,” Nanoscale, Vol 9, no 33, pp. 12124-12131, 2017.
[38]K. T. Liao, M. Tsegaye, V. Chaurey, C. F. Chou, and Swami. N. S,“Nano‐constriction device for rapid protein preconcentration in
physiological media through a balance of electrokinetic forces,”Electrophoresis, Vol 33, no 13, pp. 1958-1966, 2012.
[39]M. Azadi, and G. G. Lopez, “Spin Curves for MicroChem S1800 (1805, 1813, 1818) Series Positive Resist,” University of Pennsylvania Scholarly Commons , 2016.
[40]張揚愉, “三維微流道無電極式介電泳晶片於微量蛋白質富集及快速偵測,” 國立虎尾科技大學自動化工程系碩士班碩士論文, 2019.
[41]C. H. Lin, G. B. Lee, Y. H. Lin, and G. L. Chang, “A fast prototyping
process for fabrication of microfluidic systems on soda-lime glass,”
Journal of Micromechanics and Microengineering, 2001, Vol 11,no 6,
pp. 726, 2011.
[42]N. Mandal, V. Pakira, N. Samanta, N. Das, and S. Chakraborty et al , “PSA detection using label free graphene FET with coplanar electrodes based microfluidic point of care diagnostic device,” Talanta, vol 222,121581, 2021.
[43]楊政穎, “血液在微流道中受介電泳作用之數值模型探討研究成果報告,” 行政院國家科學委員會專題研究計畫成果報告, 2007.
[44]H. Fischer, I. Polikarpov and A. F. Craievich, “Average protein density
is a molecular-weight-dependent function,” Protein Sci, Vol. 13, no. 10, pp. 2825-2828, 2004.