惡性腫瘤為台灣十大死因之首，惡性腫瘤治療的困難之處並不僅限於腫瘤本身的消滅，更難以預防的是癌細胞轉移，常常使治療的時間拉長、難度提升。在腫瘤轉移時會產生「循環腫瘤細胞」（Circulating Tumor Cell, CTC），近年的研究表示循環腫瘤細胞的多寡與病情的嚴重性及預測方向有相當的關係，因此偵測與分析循環腫瘤細胞是近年來重要的研究方向之一，特別是對於早期癌症轉移以及接受藥物長期治療的病人而言。
　　本論文利用流體的特性，在玻璃基材上以SU-8做出結構。細胞在螢光抗體標定之後藉由重力和液體流動產生的側向拉力影響下產生單層緊密排列的自組裝陣列，在晶片中的觀測區域形成二維陣列，最後以螢光顯微鏡觀測循環腫瘤癌細胞並計算數量，並與醫院進行IRB計劃合作取得大腸癌及乳癌病患的血液檢體，並且與流式細胞儀及Isoflux系統的結果作比較。相較於實驗室細胞株的結果，檢體具有臨床意義且在大量的實驗後可以建立有效的臨床數據。
　　此研究方法製作成本較低，且結構相當簡易，檢體在晶片上十分鐘內即可完成沉降排列。本論文更建立自動化的檢測系統與細胞捉取系統統，除了縮短檢測的時間及減輕實驗人員負擔之外，更可以再檢測後取出循環腫瘤細胞做後續的細胞實驗。本研究的主要成果為：1.可運用於臨床上的檢測裝置。2.更快速的自動化檢測設備。此檢測方法未來更計畫與醫院合作進行癌症相關的檢測。3.細胞捉取設備。捕捉循環腫瘤細胞以利於後續培養或生化分析等實驗。

Malignant tumor is the first of top ten death causes in Taiwan. The difficulty is not only the treatment of malignant tumor eradication, and it is more difficult to prevent cancer metastasis. It makes treatment time stretched and hard. Circulating tumor cells (CTCs) are generated when tumor metastasis occurs. Recent research represents the severity of disease is related to the amounts of circulating tumor cells. So it is an important research direction which finds out and analysis circulating tumor cells for the clinical study.
This report uses a fluid feature for detecting CTCs. Self –Assembled Cell Array Chip makes up with SU-8 photoresist on a glass substrate. Cells produced by flow of liquid and gravity to form a high density monolayer array after fluorescent calibration. In the end CTCs can be count by the fluorescence microscope. It has IRB cooperated plan with hospital and allowed to get colorectal cancer and breast cancer patient blood sample to do the clinical research. The result of SACA Chip will be compared with flow cytometry and IsoFlux.
This report provides an inexpensive and simple way to make chip and the process costs less than ten minutes. It also builds up the automatic imaging and in-situ cell capturing process to shorten the process time and isolate CTCs after process.

摘要 ii
Abstract iii
誌謝 iv
目錄 v
圖目錄 viii
表目錄 xi
第一章 緒論 1
1.1 研究背景 1
1.2 研究目標 4
第二章 文獻回顧 5
2.1 細胞篩選方法 5
2.2免疫標定 5
2.2.1免疫抗體捕捉 5
2.2.2免疫磁珠分離 8
2.2.3流式細胞儀（Flow Cytometer） 9
2.2.4 CellSearch系統檢測循環腫瘤細胞 11
2.3非免疫標定 13
2.3.1細胞大小（Cell Size） 13
2.3.2密度梯度分離（Density） 18
2.3.3二維陣列 19
2.4文獻總結與研究目標 21
第三章 實驗設計 22
3.1 螢光檢測系統特色 22
3.2 實驗材料準備 22
3.2.1 細胞株介紹 22
3.2.2 藥品介紹 23
3.3藥品製備處理 25
3.3.1 細胞懸浮液 25
3.4 第一代微流道井式排列平台 26
3.4.1第一代平台設計及原理 27
3.4.2 第一代平台製成及組裝 29
3.4.3 第一代微流道井式細胞排列結果簡介 31
3.5 IRB癌症病患檢體測試 33
3.5.1 檢體分離及染色流程 33
3.6.2 循環腫瘤細胞影像辨識 35
3.6 自動影像檢測系統 36
3.6.1自動影像檢測系統設計 39
3.6.2 自動影像檢測系統控制流程 40
3.7 細胞自組裝排列沉降晶片 41
3.7.1 沉降晶片設計 41
3.7.2 晶片製作方式 42
3.8 細胞捉取裝置 44
3.8.1 細胞捉取裝置設計 45
3.8.2 玻璃針製作 46
第四章 結果與討論 47
4.1 IRB癌症病患檢測 47
4.1.1 循環腫瘤細胞統計 47
4.2 自動檢測系統影像拼接 51
4.2.1 自動影像檢測系統 53
4.2.2 影像拼接測試 54
4.3 細胞捉取裝置測試 60
4.3.1 玻璃針管製作 60
4.3.2 細胞捉取測試 61
第五章 結論 63
第六章 未來工作 65
參考文獻 69

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