臺灣博碩士論文加值系統

本論文提出一仿輸卵管之微流道介電泳晶片系統，為了降低對細胞的影響與破壞，本研究使用非接觸式介電泳力方式達到細胞的捕捉與研究。仿體外受精技術 (In Vitro Fertilization, IVF)，將卵母細胞透過介電泳力定位於微流道內再者利用介電泳力捕抓更多的精蟲於卵母細胞的周圍來提升自然受精的機會，而將卵母細胞定位後在微流道內精蟲隨著流體皆能朝同一方向前進且卵母細胞周圍有更多的精蟲即能提高兩者自然授精的機率。
介電泳力主要透過非均勻電場產生密度不均的電場分佈，根據粒子與溶液間之介電係數(permittivity)與導電度(conductivity)差異，且在電場誘導產生不同誘導偶極矩，藉由溶液與粒子不同極化能力的特性，產生正或負介電泳效應，使粒子因受介電泳力的影響，驅動往高電場密度或低電場密度的特定位置，達到卵母細胞定位的效果。實驗中發現在其在高頻1MHz之下卵母細胞產生正介電泳力，進而可將卵母細胞捕抓在強電場區域上。此外利用數值模擬軟體先了解流道內高與低電場的分佈位置，以便了解在實驗中細胞產生正或負介電泳現象時會驅動到達之位置。
本實驗研究分為兩階段，第一階段完成絕緣結構微流道介電泳晶片系統製作與架設，主要在流道內進行細胞的位置操控，其中包含晶片設計及製作、介電泳電場模擬、介電泳細胞操控等結果。第二階段則承接第一階段的研究進一步改良成電極式介電泳晶片，來提升在流場下卵母細胞被定位以及精蟲與卵母細胞達到受精的目的。

This study presents an imitation of oviduct microfluidic dielectrophoresis chip system. In order to reduce the impact and destruction of cells, this study adopts dielectrophoresis force to manipulate the cell. Imitation of IVF (In Vitro Fertilization), the oocyte through the DEP force positioned in the microfluidic channel, and then we trap much more sperms around the oocyte by dielectrophoresis force to enhance the probability of natural fertilization. Therefore able to oocyte positioning in the microchannel and sperm with the fluid also go the same direction, so that the much more sperms around the oocyte can enhance the probability of natural fertilization.
Dielectrophoresis force primarily through non-uniform electric field induces the electric field distribution. According to the differences between of particle and solutions’ dielectric coefficient and conductivity induce difference dipole moment in the electric field. Different polarization characteristics of ability of particles and solution to generate positive or negative dielectrophoresis effect, the particles will be affected by the DEP force to move to the region with high or low electric field density to achieve the goal of oocyte positioning. The positive dielectrophoresis response of oocyte was exhibited with the frequency at 1 MHz, the oocyte will be affected by the DEP force to move to the region with high electric field density. To understand the position with high and low electric field distribution in microchannel, the numerical commercial software, CFDRC-ACE+, was used to know where the high electric field is.
This study is divided into two parts, the first to complete the insulator structure of microchannel dielectrophoresis chip system fabrication and setup. Mainly we manipulate cell in the microchannel, which contains the chip design and fabrication, dielectrophoresis electric field simulations, dielectrophoresis cell manipulation results. The second part improves the dielectrophoresis chip, which enhance the probability of oocyte position and sperm collision oocyte.

誌謝 I
中文摘要 II
Abstract III
目錄 IV
圖目錄 VII
第一章 緒論 1
1.1 前言與研究背景 1
1.2 研究動機 2
第二章 文獻回顧 5
2.1 機械力 (Mechanical Force) 5
2.2 光鉗 (optical tweezer) 9
2.3 電磁力 (Electromagnetic Force) 12
2.4 電泳 (Electrophoresis，EP) 14
2.5 介電泳 (Dielectrophoresis，DEP) 15
2.6 研究方法 22
第三章 介電泳理論 23
3.1 介電泳理論 23
3.2 Clausious-Mossotti 因子 27
第四章 實驗架構與方法 29
4.1 晶片幾何結構設計 29
4.2 CFDRC數值模擬分析與結果 31
4.2.1 穩態(steady state)電場分佈 32
4.2.2 暫態(transient state)粒子捕捉現象 34
4.3 細胞樣本處理 37
4.3.1 肝癌細胞(Hep-G2)培養 38
4.3.2 ICR老鼠之卵母細胞 40
4.4 介電泳晶片製作 43
4.4.1 電極製作 44
4.4.2 SU8-3050微流道結構製作 47
4.4.3 PDMS翻模 49
4.4.4 氧電漿晶片接合 50
4.5 實驗儀器與設備 52
4.6 主體實驗流程 53
4.7 實驗結果 54
4.7.1 肝細胞(Hep-G2)在不同頻率下之介電泳效應 54
4.7.2 ICR老鼠之卵母細胞之介電泳效應 59
4.8 問題與討論 63
4.8.1 晶片電極對位 63
4.8.2 微流道內氣泡的影響 64
4.8.3 溶液置換 64
4.8.4 介電泳頻率 65
4.8.5 卵子型態 66
第五章 電極式介電泳提升精卵受精機會 67
5.1 晶片設計改善 67
5.2 CFDRC數值模擬分析與結果 68
5.2.1 穩態(steady state)電場分佈 68
5.3 細胞樣本處理 70
5.3.1 ICR老鼠之精蟲細胞 70
5.4 實驗結果 72
5.4.1 傳統培養對照組 72
5.4.2 晶片實驗控制組 73
5.5 問題與討論 77
5.5.1 鼠精頭部互勾 77
5.5.2 BSA coating改善流道內細胞沾黏 79
5.5.3 1MHz下精蟲為正介電泳 81
第六章 結論與未來展望 82
6.1 結論 82
6.2 精確定位卵子 83
6.3 整合微流道精蟲篩及電濕潤選機制以實現IVF 83
第七章 參考文獻 85

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