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研究生:黎耀軒
研究生(外文):Yao-Hsuan Lai
論文名稱:氧化銦鎵鋅薄膜電晶體生物感測器應用於生物化學反應之分析
論文名稱(外文):Investigation of Biochemical Reaction Kinetics by Transient Signal Responses of an IGZO-TFT Biosensor
指導教授:黃建璋黃建璋引用關係
指導教授(外文):Jian-Jang Huang
口試委員:楊志忠林致廷黃念祖
口試委員(外文):Chih-Chung YangChih-Ting LinNien-Tsu Huang
口試日期:2020-07-21
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:光電工程學研究所
學門:工程學門
學類:電資工程學類
論文種類:學術論文
論文出版年:2020
畢業學年度:108
語文別:英文
論文頁數:49
中文關鍵詞:薄膜電晶體生物感測器微流道蘋果酸-天冬氨酸穿梭動態反應反應常數
外文關鍵詞:TFTbiosensormicrofluidic channelmalate-aspartate shuttlekinetic reactionreaction constant
DOI:10.6342/NTU202002574
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這篇論文介紹以氧化銦鎵鋅薄膜電晶體與感測金屬電極組成之生物感測器偵測生物分子擴散與混合狀態並探討蛋白質與配體之動態反應,此研究分兩部分:
第一部分,以溶菌酶及其適體三乙醯殼三糖作為動態反應分析標的物,使用薄膜電晶體結合微流道作為感測的平台。將溶菌酶跟三乙醯殼三糖溶液注入微流道中來感測電流訊號。首先,單獨通入溶菌酶溶液至流道中,以建立溶菌酶濃度與電流變化關係。接者,將三種濃度比例的溶菌酶與三乙酰殼三糖混合在離心管中,並控制混和的反應時間;對擷取之電流變化,考量屏蔽效應進行修正後,可將建立之溶菌酶濃度與電流變化關係將電流變化轉為剩餘溶菌酶濃度。以此,可建立剩餘溶菌酶濃度與反應時間的擬合曲線。透過化學公式,可從曲線中得到反應級數、結合速率常數與分解常數。其中,分解常數之結果為39.10μM,與其他團隊之研究所提出之數值十分接近。
第二部分, 以煙酰胺腺嘌呤二核苷酸,還原形式,蘋果酸脫氫酶和草酰乙酸之間的化學反應動力學為研究目標。首先,將不同濃度的煙酰胺腺嘌呤二核苷酸與煙酰胺腺嘌呤二核苷酸,還原形式溶液注入微流體通道,以觀察電流訊號。通過擬合適當的擴散模型,可以得出汲極電流隨時間變化的函數。接著,測量不同濃度倍數的NADH,OAA和MDH溶液混合物。通過生化公式的計算,量測出的曲線可提供解離常數等信息。值得注意的是,導出的平衡常數為(8.06±0.61)×104,與其他團隊的研究結果相近。
In this thesis, a biosensor consists of an Indium-Gallium-Zinc-Oxide (IGZO) thin-film transistor (TFT), and a gold sensing electrode is demonstrated for diffusion and mixing properties detection of biomolecules. The biochemical reaction kinetics are further investigated. The thesis includes two parts.
In the first part, interaction kinetics of lysozyme and tri-N-Acetylglucosamine (NAG3) are investigated and applied to a TFT biosensor integrated with a linear shape microfluidic channel. First, different concentrations of lysozyme solution were introduced into the microfluidic channel to construct the relationship between the lysozyme concentration and the change of drain current. Then, three mixed ratios of lysozyme and NAG3 solutions were incubated in a microcentrifuge for different reaction times. Considering the screen effect, the extracted drain current variations are calibrated by the revision factor and the revised current variations are converted into remained lysozyme concentration by the correlation of current variation and lysozyme concentration. Based on the converted information, the fitting curves of remained lysozyme concentration versus reaction time are presented. The curves provide information that can be exploited to calculate the partial orders, association rate constant, and dissociation constant by biochemical formulas. It is worth noted that the derived dissociation constant is 39.10 μM, which is close to the results reported by previous researches.
In the second part, biochemical reaction kinetics between NADH (Nicotinamide Adenine Dinucleotide, reduced form), MDH (Malate dehydrogenase), and OAA (Oxaloacetic Acid, the conjugate acid of oxaloacetate) are investigated. First, NADH and NAD+ (Nicotinamide Adenine Dinucleotide) solutions of several concentrations are introduced into the microfluidic channel to observe the transient current response. By fitting the appropriate diffusion model, functions of drain current change to time can be derived. Then, different mixtures of NADH, OAA, and MDH solution are measured. By taking drain current change functions of NADH and NAD+ into consideration, the drain current function of the mixtures can be derived. Thus, by fitting the function of the mixture to the drain current profile, the ratio of NAD+ to NADH can be obtained. The curves provide the information that can be utilized to calculate the ratio of NAD+ to NADH and dissociation constant by biochemical formulas. It is worth noting that the derived apparent equilibrium constant is (8.06±0.61) ×104, which is close to the results reported by previous researches.
致謝 2
中文摘要 3
ABSTRACT 4
CONTENTS 6
LIST OF FIGURES 8
LIST OF TABLES 11
Chapter 1 Introduction 1
1.1 Overview of Biochemical Detection 1
1.2 Introduction of FET-based Biosensors 2
1.3 Importance of Biochemical Reaction Kinetics 4
1.4 Thesis Outline 5
Chapter 2 IGZO-TFT Biosensors for Investigation of Lysozyme and tri-N-acetylglucosamine Reaction Kinetics 7
2.1 Introduction 7
2.2 Material and Methods 8
2.2.1 Fabrication of IGZO-TFT Biosensors Integrated with Microfluidic Channels 8
2.2.2 Introduction of lysozyme and tri-N-acetylglucosamine 11
2.2.3 Measurement and experiment flow 12
2.3 Results and Discussion 15
2.3.1 Confirmation of diffusion dominant or flow dominant 15
2.3.2 Real-time analysis of lysozyme and tri-N-acetylglucosamine 16
2.3.3 Detection of lysozyme and tri-N-acetylglucosamine kinetic reaction 20
2.3.4 Extraction of biochemical constant 25
2.4 Summary 28
Chapter 3 IGZO-TFT Biosensors for Investigation of Malate-Aspartate Shuttle Biochemical Reaction Kinetics 30
3.1 Introduction 30
3.2 Material and Methods 31
3.2.1 Introduction of reduced nicotinamide adenine dinucleotide, oxaloacetate and malate dehydrogenase 31
3.2.2 Measurement and experiment flow 32
3.3 Results and Discussions 34
3.3.1 Real-time analysis of reduced nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide 34
3.3.2 Detection of reduced mixed nicotinamide adenine dinucleotide and oxaloacetate kinetic reaction with malate dehydrogenase 38
3.3.3 Extraction of biochemical constant 41
3.4 Summary 43
Chapter 4 Conclusions 44
REFERENCE 46
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