神經壞死病毒與虹彩病毒皆為高傳染性及高致病性病毒，可感染多種魚類。以石斑魚魚苗為例，其感染神經壞死病毒或虹彩病毒，死亡率均高於百分之八十。高死亡率造成養殖漁業重大經濟損失。現有檢測技術如病毒培養、酵素免疫分析法、聚合酶鏈鎖反應/反轉錄聚合酶鏈鎖反應有些缺點，包含費時、操作過程複雜、需要大型專業儀器及訓練有素之專業人員操作實驗。此外，目前對已感染病毒的魚並沒有適當的治療方式。因此，快速及準確的診斷神經壞死病毒與虹彩病毒對於養殖漁業是非常重要的議題。若愈早診斷出受病毒感染的魚，可越早將其撲殺，以防止疾病的擴散，進而降低經濟損失。在本研究中，以整合式微流體系統，包括整合式機台、微流體晶片、專一性磁珠及專一性分子信標，能夠在三十分鐘內自動化診斷神經壞死病毒或虹彩病毒，大幅縮短檢測時間。在本研究中，偵測神經壞死病毒或虹彩病毒的最低偵測極限均為10 ng/μL。最重要的是，此整合式微流體系統可成功自魚苗魚體中偵測出受神經壞死病毒或虹彩病毒感染之魚苗。因此，本研究之微流體系統未來可運用於田間檢測魚類病原體的有效診斷工具。

Abstract I
中文摘要 III
誌謝 IV
Table of Content V
List of Figure X
List of Table XIII
Abbreviations XIV
Nomenclature XVI
Chapter 1: Introduction 1
1.1 MEMS and Bio-MEMS Technology 1
1.2 Molecular Beacon 3
1.3 Literature Review 4
1.3.1 Nervous Necrosis Virus 4
1.3.2 Iridovirus 5
1.3.3 Conventional Diagnosis Methods 6
1.4 Motivation and Objectives 7
Chapter 2: Theory 10
2.1 Microfluidic Control System 10
2.1.1 Suction-type Pneumatic-driven Micropump 10
2.1.2 Micro-mixer 12
2.2 Molecular Diagnosis 13
2.2.1 DNA and RNA 13
2.2.2 PCR and RT-PCR 14
2.2.3 Gel Electrophoresis 15
2.2.4 Fluorescence Resonance Energy Transfer 16
Chapter 3: Materials and Methods 23
3.1 Viral Strain 23
3.2 Probe Conjugated Magnetic Beads 25
3.2.1 Preparation of Probe Conjugated Magnetic Beads 25
3.2.2 Hybridization Condition of Probe Conjugated Magnetic Beads 26
3.3 Molecular Beacon 27
3.4 Chip Design 27
3.5 Fabrication of Microfluidic Chip 28
3.6 Experimental Procedure 29
3.7 Custom-made Control System 30
Chapter 4: Results and Discussion 36
4.1 NNV RNA2 Molecular Beacon Assays 36
4.1.1 Optimal Hybridization Temperature of Specific NNV RNA2 Probe Conjugated Magnetic Beads 37
4.1.2 Optimal Hybridization Temperature of Molecular Beacon 38
4.1.3 Optimal Hybridization Time of Molecular Beacon 39
4.1.4 Detection Limit of Molecular Beacon 40
4.1.5 Specificity Tests of Molecular Beacon 41
4.1.6 Diagnosed of Infected and Non-infected NNV Fish Sample by Specific Probe and Molecular Beacon 42
4.2 Iridovirus MCP Molecular Beacon Assays 44
4.2.1 Optimal Hybridization Temperature of Specific Iridovirus MCP Probe Conjugated Magnetic Beads 44
4.2.2 Optimal Hybridization Temperature of Iridovirus MCP Molecular Beacon 45
4.2.3 Optimal Hybridization Time of Molecular Beacon 46
4.2.4 Detection Limit of Molecular Beacon 47
4.2.5 Specificity Test of Molecular Beacon 48
4.2.6 Diagnosis of Iridovirus Infected and Non-infected Fish Sample 50
Chapter 5: Conclusions and Future Work 68
5.1 Conclusions 68
5.2 Future Work 69
References 71

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