生醫微流體晶片應用於生殖系統方面已經越來越普及。以往用以分選的機制大致可以分為：1.化學梯度吸引 2.熱吸引 3.穩定層流分選 4.逆向流場分選。但這些分選方法中，除了穩定層流外，都會對精子本身活動力有一定的削減。而穩定層流的方法又無法有效地分別出精子的活動力。因此，本研究提出了一個漸擴流場的機制，隨著流道的逐漸增擴，流速將相對的減慢。在流場中的精子感受到流場會對其做抵抗。同時隨著流場的減弱而依照本身運動能力停留在不同區塊，得到分選的效果。在分選漸擴區域，開設一個側枝，準備將分選後的精子取出。由於側枝在分選時做了氣密處理，與入口並無壓力差，因此精子在經過時並不會有流場將之帶入，而是僅依靠擴散的方式進入。側枝流道可以成功的取出高效精子同時不影響分選。分選結束後，將側枝的氣密解除，使用滴管將高效精子取出。如此一來，操作簡易，移除了多數實驗所需要的幫浦，在臨床上也是一大優勢。經過分選的精子，數量約2000至15000隻，存活率達到77%，證實晶片確實可將死亡精子沖洗掉，篩檢出存活精子。活動力方面，Motile sperm由38%提升至70%，Progressive Sperm則是由11%提升至33%，可見經過精片作用，高效能精子成功被取出。最後將分選後的精子與本實驗室的研究” 建構三維結構之微流體裝置應用於鼠卵細胞培養與透明帶去除”所剝殼的老鼠卵子做結合後，與使用傳統方式剝殼並配上未分選精子後做比較，受孕潛力較傳統方式的40%提升到60%。另外，本晶片使用的是PMMA壓克力。配合熱壓模的製程方法，可以以便宜又快速的方式量產。流道入口、出口端接有文字做標示，協助使用者操作，避免人為疏失造成晶片失效。經過封裝後，可將PMMA塑膠晶片做成一可以快速量產的商業產品，應用於人工試管嬰兒。

This paper proposes a selection and extraction strategy to acquire high quality sperms from a biomimetic microfluidic device capable of generating a dual gradient flow field for motile sperm sorting. The sorter consists of a straight flow zone, a rapidly expansion sector for sperm sorting by gradient up-stream flow, and a dumbbell channel for dead sperms separation at the outlet. A collateral runner is set at the front area of the expansion sector without affecting the sorting process because the chamber is blocked while sorting. At the end of sorting, the collateral chamber is open and then using just pipette to derive high quality sperms instead of pump. Controllable numbers, ranging from 2000 to 10000, of sperms can be collected with a motility (>70μm) ratio higher than 70%, suitable for in-vitro fertilization (IVF). Hot embossing lithography process is presented. Microchannels are replicated on a Polymethylmetacrylate (PMMA) stamp with thickness of 1 mm. With low cost and high replication precision, we can make it a clinical chip.

摘要 I
ABSTRACT II
致謝 III
總目錄 IV
圖目錄 IX
表目錄 XII
第一章緒論 1
1.1研究背景 1
1.1.1微機電系統 1
1.1.2微流體生醫晶片 1
1.2研究動機 3
第二章文獻回顧 6
2.1.晶片內部微流體層流之特性 6
2.1.1以穩定層流分離精子 7
2.1.2利用精子本身活動力分選 9
2.2逆向流場 11
2.2.1利用精子逆流的特性分選具活動力的精子 11
2.2.2漸括流場 16
2.3熱壓模製程 18
2.3.1熱壓模 18
2.3.2脫模 19
2.4研究目的 21
第三章晶片分析 24
3.1流道設計 24
3.2流速計算 26
3.3流場內壓力計算 29
3.4 晶片製程 33
3.4.1黃光微影製程 33
3.4.1.1母模製作 34
3.4.1.2軟微影製作 36
3.4.2熱壓模 37
3.5實驗 39
3.5.1檢體來源 39
3.5.1.1 PDMS多孔薄膜 40
3.5.1.2人精對於聚二甲基矽氧烷(PDMS)沾黏性之測試 41
3.5.2晶片材料選擇 42
3.5.3實驗方法 44
3.5.3.1 PDMS晶片 44
3.5.3.1.1親水化 44
3.5.3.1.2主動式注入 46
3.5.3.1.3被動式載入 47
3.5.3.1.4精子取樣 47
3.5.3.2 PMMA壓克力晶片 48
3.5.3.2.1潤濕 49
3.5.3.2.2載入 49
3.5.3.2.3取樣 50
3.5.3.3晶片產品 50
3.5.4精子染色 51
3.5.5效能估算 52
3.5.5.1晶片可處理的精子數量 52
第四章結果與討論 53
4.1黃光顯影製作母模結果 53
4.2塑膠晶片製程結果 53
4.2.1熱壓模 53
4.2.2熱接合 56
4.3 PDMS晶片測試結果 58
4.3.1檢體載入 58
4.3.1.1主動式推入 58
4.3.1.2被動式載入 59
4.3.2推出精子結果 59
4.3.2.1SYRINGE PUMP協助推入 59
4.3.2.2手指按壓 61
4.4塑膠晶片測試結果 62
4.4.1流場 62
4.4.2多孔薄膜過濾效果 63
4.4.3氣密討論 64
4.4.4取樣方法 65
4.4.5分選效能 66
4.4.7精卵結合受孕潛力 69
第五章結論與展望 71
5.1結論 71
5.2未來工作 71
5.2.1PDMS多孔薄膜探討 71
5.2.2側枝角度討論 72
5.2.3精卵結合 72
5.2.4結合精子晶片與卵子晶片 72
第六章參考文獻 73

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