Development and Evaluation of a Miniaturized Taste Sensor Chip

A miniaturized taste sensor chip was designed for use in a portable-type taste sensing system. The fabricated sensor chip (40 mm × 26 mm × 2.2 mm) has multiple taste-sensing sites consisting of a poly(hydroxyethyl methacrylate) hydrogel with KCl as the electrolyte layer for stability of the membrane potential and artificial lipid membranes as the taste sensing elements. The sensor responses to the standard taste substances showed high accuracy and good reproducibility, which is comparable with the performance of the sensor probe of the commercialized taste sensing system. Thus, the fabricated taste sensor chip could be used as a key element for the realization of a portable-type taste sensing system

Analisis Pola Keluaran Prototipe Sensor Rasa Portable Campuran Lipid Dioctyl Phosphate Dan Trioctyl Methyl Ammonium Chloride

Portable taste sensor prototype has been fabricated as two channel ion meter and one types of ion selecting membrane. Two channel ion meter were made for pre development of the taste sensor system or as a electronic tongue wich acted as an multi channel ion meter.ion meter was made using IC op-amp LF412 as its amplifier and microcontroller Atmega8535. meanwhile types of the selecting membrane ion were made using variation: mix of lipid diocthyl phosphate (DOP) and triocthyl methyl ammonium chloride (TOMA) with mass ratio respectively 3:7 and 9:1. On this sensor, lipid determines membrane selectivity to particular ions. Output from taste sensor is a electric potential and then it could be compared with electrode potential references. This portable taste sensor prototype is then tested is ability to sensing for 5 types of solution: glucose, KCl, HCl, quinine, and MSG. Those 5 types represent 5 basic taste: sweet by glucose, salty by KCL, sour by HCl, bitter by quinine, and umami by MSG. Every solution is made using 1mM KCl solution and it is variated to be 7 types of concentration. As a result, taste sensor with lipid membrane mixture of 3:7 mixing tend to respon to the anions sample, resulting potential which decreased for every increase in concentration. For membrane with a lipid mixture of 9:1 showed a tendency to respond to the cations sample, resulting potential which increased for every increase in concentration

Advanced Taste Sensors Based on Artificial Lipids with Global Selectivity to Basic Taste Qualities and High Correlation to Sensory Scores

Effective R&D and strict quality control of a broad range of foods, beverages, and pharmaceutical products require objective taste evaluation. Advanced taste sensors using artificial-lipid membranes have been developed based on concepts of global selectivity and high correlation with human sensory score. These sensors respond similarly to similar basic tastes, which they quantify with high correlations to sensory score. Using these unique properties, these sensors can quantify the basic tastes of saltiness, sourness, bitterness, umami, astringency and richness without multivariate analysis or artificial neural networks. This review describes all aspects of these taste sensors based on artificial lipid, ranging from the response principle and optimal design methods to applications in the food, beverage, and pharmaceutical markets

Electronic Noses and Tongues: Applications for the Food and Pharmaceutical Industries

The electronic nose (e-nose) is designed to crudely mimic the mammalian nose in that most contain sensors that non-selectively interact with odor molecules to produce some sort of signal that is then sent to a computer that uses multivariate statistics to determine patterns in the data. This pattern recognition is used to determine that one sample is similar or different from another based on headspace volatiles. There are different types of e-nose sensors including organic polymers, metal oxides, quartz crystal microbalance and even gas-chromatography (GC) or combined with mass spectroscopy (MS) can be used in a non-selective manner using chemical mass or patterns from a short GC column as an e-nose or “Z” nose. The electronic tongue reacts similarly to non-volatile compounds in a liquid. This review will concentrate on applications of e-nose and e-tongue technology for edible products and pharmaceutical uses

나노디스크 및 나노베지클에 내장된 후각 수용체의 최적의 생산과 냄새의 패턴 및 시각화에 대한 응용

학위논문(박사) -- 서울대학교대학원 : 공과대학 화학생물공학부, 2022. 8. 박태현.G protein-coupled receptors (GPCRs) are the most intensively studied for screening drug targets. Especially, class A GPCR including olfactory receptor (OR) which accounts for about 85 % of GPCR family is more important for codifying and screening target receptors. There are about 400 kinds of ORs in human olfactory system. The interactions between ORs and odorants generate signals which are transferred to brain as combinatorial codes. Humans can discriminate more than 1 trillion olfactory stimuli with a limited number of ORs because of widespread OR-driven modulation such as inhibition and enhancement in peripheral olfactory coding. Since the sense of smell perceives the complex external world as a pattern, many studies have been conducted to mimic the response of ORs. In particular, protein-based nanobiosensor is expected as a platform to mimic the olfaction because it has advantages such as mass production, ease of reuse, and low cost. However, reconstitution of the structure of GPCRs is challenging because almost all GPCRs produced in E. coli system are expressed as inclusion bodies. For this reason, reconstitution techniques have been developed to recover the functionality of GPCRs, such as the use of detergent micelles, nanovesicles, bicelles and nanodiscs (NDs). Among these materials, NDs have been considered the most effective reconstitution material because of their stability in various environments and their functional lifetimes. In this thesis, ORs were produced in E. coli system with high productivity and reconstituted to ND or nanovesicle forms. Then the functional reconstituted ORs were applied to monitoring meat freshness/spoilage, disease diagnosis and practical colorimetric sensor. First, OR was overexpressed by coexpressing effector genes, such as djlA, the membrane-bound DnaK cochaperone, and rraA, inhibitor of the mRNA-degrading activity of E. coli RNase E. The E. coli strains coexpressing DjlA or RraA suppressed protein-induced toxicity and overexpressed the ORs. By controlling the molar ratio of OR, membrane scaffold protein, and phospholipid, ND of appropriate size were made, and high-purity ND could be purified. OR-embedded NDs showed stability to various temperature and storage time. Second, Human ORs which bind to gastric cancer and halitosis biomarkers were successfully reconstituted to ND form and purified. The NDs had various patterns to artificial saliva samples because NDs had various binding affinities to target molecules. Through principal component analysis of various patterns for artificial saliva samples, it was possible to distinguish between healthy control samples and patient samples. Third, trace amine-associated receptors (TAARs), TAAR13c and TAAR13d, were successfully overexpressed in E. coli system and reconstituted to ND form. These NDs were utilized for development of ND-based BE-nose for monitoring meat freshness. The ND-based BE-noses was successfully performed towards diverse on-site and the various real samples and could be used to monitor freshness of meat. Lastly, human OR1A2 (hOR1A2) was reconstituted into detergent micelle and it was used for development of colorimetric sensor detecting geraniol. Polydiacetylene (PDA) was used as secondary transducer for visualization of responses of OR. The structural and functional properties of the hOR1A2 were maintained when it was embedded in PDA/lipid nanovesicles. The hOR1A2 embedded in PDA/lipid nanovesicle caused a color transition from blue to purple when it reacted with geraniol, whereas there was no color transition when it reacted with other molecules. In this study, various ORs were successfully reconstituted with ND or nanovesicles. The reconstructed OR is expected to be applied to food freshness monitoring, disease diagnosis by pattern analysis, and practical colorimetric sensors.G 단백질 연결 수용체 (GPCR)는 약물 표적 스크리닝을 위해 가장 집중적으로 연구되는 단백질이다. 특히, GPCR 계열의 약 85%를 차지하는 후각 수용체 (OR)를 포함하는 클래스 A GPCR은 표적 수용체를 코드화하고 스크리닝하는 데 더 중요하다. 인간의 후각 시스템에는 약 400 종류의 OR이 있다. OR과 냄새 물질 사이의 상호 작용은 패턴의 조합으로 뇌에 전달되는 신호를 생성한다. 인간은 말초 후각 코딩의 억제 및 향상과 같은 광범위한 OR에 의한 조절로 인해 제한된 수의 OR로도 1조개 이상의 후각 신호를 구별할 수 있다. 후각은 복잡한 외부 세계를 패턴으로 인식하기 때문에 OR의 반응을 모방하기 위해 많은 연구가 수행되었다. 특히, 단백질 기반 나노바이오센서는 양산성, 재사용 용이성, 저비용 등의 장점이 있어 후각을 모방하는 플랫폼으로 기대된다. 그러나 GPCR 구조의 재구성은 대장균 시스템에서 생성되는 거의 모든 GPCR이 봉입체로 발현되기 때문에 어려운 일이다. 이러한 이유로 세제 미셀, 나노베지클, 바이셀 및 나노디스크 (ND)와 같은 GPCR의 기능을 복구하기 위한 재구성 기술이 개발되었다. 이들 물질 중 ND는 다양한 환경에서의 안정성과 기능적 수명 때문에 가장 효과적인 재구성 물질로 여겨져 왔다. 본 논문에서는 대장균 시스템에서 높은 생산성으로 OR을 생산하여 나노디스크 또는 나노베지클 형태로 구조를 재구성하였다. 그런 다음 기능적으로 재구성된 OR을 육류 신선도/부패 모니터링, 질병 진단 및 실용적인 비색 센서에 적용했다. 첫째, OR은 djlA, 막 결합 DnaK cochaperone 및 rraA, E. coli RNase E의 mRNA 분해 활성 억제제와 같은 이펙터 유전자를 공동 발현함으로써 과발현되었다. DjlA 또는 RraA를 공동 발현하는 대장균 균주는 단백질 발현에 의한 독성을 억제하고 후각 수용체를 과발현했습니다. 후각 수용체, 막 지지체 단백질, 인지질의 몰비를 조절하여 적절한 크기의 ND를 만들고 고순도 ND를 정제할 수 있었다. 후각 수용체가 내장된 ND는 다양한 온도 및 보관 시간에 대해 안정성을 보였다. 둘째, 위암 및 구취 바이오마커에 결합하는 인간 OR은 ND 형태로 성공적으로 재구성되고 정제되었다. ND는 표적 분자에 대한 다양한 결합 친화성을 가졌기 때문에 인공 타액 샘플에 대한 다양한 패턴을 가졌다. 인공 타액 샘플에 대한 다양한 패턴의 주성분 분석을 통해 건강한 대조군 샘플과 환자 샘플을 구별할 수 있었다. 셋째, 미량 아민 관련 수용체 (TAAR), TAAR13c 및 TAAR13d가 대장균 시스템에서 성공적으로 과발현되었고 ND 형태로 재구성되었다. 이러한 ND는 육류 신선도를 모니터링하기 위한 ND 기반 전자 코의 개발에 활용되었다. ND 기반 전자 코는 다양한 현장 및 실제 샘플에 성공적으로 작동되었으며 육류의 신선도를 모니터링하는 데 사용할 수 있었다. 마지막으로 인간 OR1A2 (hOR1A2)를 세제 미셀로 재구성하여 제라니올을 감지하는 비색 센서 개발에 활용하였다. 폴리다이아세틸렌 (PDA)은 후각 수용체의 반응을 시각화하기 위한 2차 변환기로 사용되었다. hOR1A2의 구조적 및 기능적 특성은 PDA/지질 나노베지클에 내장되었을 때 유지되었다. PDA/지질 나노베지클에 내장된 hOR1A2가 geraniol과 반응할 때 파란색에서 보라색으로 색상 전이를 일으킨 반면 다른 분자와 반응할 때는 색상 전이가 없었습니다. 이 연구에서는 다양한 OR이 ND 또는 나노베지클로 성공적으로 재구성되었다. 재구성된 OR은 식품 신선도 모니터링, 패턴 분석에 의한 질병 진단 및 실용적인 비색 센서에 적용될 수 있을 것으로 기대가 된다.Chapter 1 Research background and objective 15 Chapter 2 Literature review 20 2.1 Olfaction 22 2.1.1 Olfactory system 21 2.1.2 Olfactory receptors 25 2.1.3 Patterns of odorants 29 2.2 Nanobiosensor 32 2.2.1 Nanobiosensor system 32 2.2.2 Components for the nanobiosensor 35 2.2.3 Nanobiosensor detecting smell 37 2.3 Biomarkers in disease and food spoilage 44 2.3.1 Gastric cancer 44 2.3.2 Halitosis 47 2.3.3 Meat spoilage 49 2.4 Expression of GPCR in E. coli system 50 Chapter 3 Experimental procedures 52 3.1 Materials 53 3.2 Gene Cloning . 54 3.3 Expression 54 3.3.1 Expression of olfactory receptors in E. coli. 54 3.3.2 Expression of membrane scaffold protein in E. coli 56 3.3.3 Expression of olfactory receptors in HEK-293T cell 57 3.4 Purification. 57 3.4.1 Purification of olfactory receptors. 57 3.4.2 Purification of membrane scaffold protein 58 3.5 Functional reconstitution of olfactory receptors 59 3.5.1 Nanodisc 59 3.5.2 Detergent micelle. 60 3.5.3 Polydiacetylene/Lipid nanovesicle 61 3.6 Characterization . 61 3.6.1 Nano-glo dual luciferase assay 61 3.6.2 SDS-PAGE analysis 62 3.6.3 Dynamic light scattering 63 3.6.4 Circular dichroism . 63 3.6.5 Tryptophan fluorescence quenching assay 63 3.7 Immobilization of olfactory receptor-embedded nanodisc on graphene 64 Chapter 4 Enhancement of olfactory receptor production in E. coli system and characterization of olfactory receptor-embedded nanodiscs. 66 4.1 Introduction 67 4.2 Expression and purification of olfactory receptor in E. coli system 69 4.3 Purification and size analysis of olfactory receptor-embedded nanodiscs. 73 4.4 Stability of immobilized olfactory receptor-embedded nanodiscs. 77 4.5 Conclusions 82 Chapter 5 Development of nanodisc-based bioelectronic nose using trace amine-associated receptors for monitoring meat freshness/spoilage . 84 5.1 Introduction 85 5.2 Characterization of TAARs 87 5.3 Performance of nanodisc-based bioelectronic nose in the liquid phase. 91 5.4 Gas sensing performance of bioelectronic nose and its application to meat spoilage 97 5.5 Measurements of real samples using TAAR nanodisc-based bioelectronic nose 99 5.6 Conclusions 104 Chapter 6 Pattern analysis for gastric cancer biomarkers using human olfactory receptor-embedded nanodiscs . 105 6.1 Introduction 106 6.2 Affinities of human olfactory receptor-embedded nanodiscs to gastric cancer biomarkers 108 6.3 Patterns for gastric cancer biomarkers in artificial saliva 112 6.4 Principal component analysis for artificial saliva samples 115 6.5 Conclusions 117 Chapter 7 Pattern analysis for halitosis biomarkers in artificial saliva using olfactory receptor-embedded nanodiscs 118 7.1 Introduction 119 7.2 Characterization of olfactory receptor-embedded nanodiscs. 120 7.3 Patterns for halitosis biomarkers in artificial saliva. 122 7.4 Principal component analysis for artificial saliva samples 125 7.5 Conclusions 127 Chapter 8 Visual detection of geraniol using human olfactory receptor embedded in polydiacetylene/lipid nanovesicle 128 8.1 Introduction 129 8.2 Functionality of hOR1A2 embedded in detergent micelle and PDA/lipid nanovesicle. 130 8.3 Structural assay of hOR1A2 embedded in detergent micelle and PDA/lipid nanovesicle. 133 8.4 Size analysis and morphology of hOR1A2 embedded in PDA/lipid nanovesicle. 135 8.5 Photoluminescence intensity of hOR1A2 embedded in PDA/lipid nanovesicle. 137 8.6 Conclusions 141 Chapter 9 Overall discussion and further suggestions. 142 Bibliography . 147 국문초록 157박

Fabrication Of An Electronic Nose And Its Application For The Verification Of Eurycoma Longifolia Extracts [TP159.C46 I82 2007 f rb].

Hidung elekronik yang berasaskan penderia penimbang mikro hablur kuarza menggunakan etil selulosa, lipid (dioktil fosfat (DOP), trioktil metil ammonium klorida (TOMA), olil amina (OAm)) dan bahan fasa pegun kromatografi gas (Apiezon L (APZL),polipropilin glikol 1200 (PPG 1200), polietilina glikol 1000 (PEG 1000), polietilina glikol 4000 (PEG 4000), poli(bissanopropil-siloksana) (OV-275) dan dietilina glikolsuksinat (DEGS) sebagai membran penderia untuk menganalisis ekstrak daripada Eurycoma longifolia (Tongkat Ali) telah dibina. An electronic nose based on a quartz crystal microbalance array sensor using ethylcellulose (EC), lipids ((dioctyl phosphate (DOP), trioctyl methyl ammonium chloride (TOMA), oleyl amine (OAm)) and gas chromatography (GC) stationary phase materials ((Apiezon-L (APZ-L), polypropylene glycol 1200 (PPG 1200), polyethylene glycol 1000 (PEG 1000), polyethylene glycol 4000 (PEG 4000), poly(biscyanopropyl-siloxane) (OV-275) and diethylene glycol succinate (DEGS)) as sensing membrane for the analysis ofextracts of Eurycoma longifolia (Tongkat Ali) was develope

Potential use of electronic noses, electronic tongues and biosensors, as multisensor systems for spoilage examination in foods

Development and use of reliable and precise detecting systems in the food supply chain must be taken into account to ensure the maximum level of food safety and quality for consumers. Spoilage is a challenging concern in food safety considerations as it is a threat to public health and is seriously considered in food hygiene issues accordingly. Although some procedures and detection methods are already available for the determination ofspoilage in food products, these traditional methods have some limitations and drawbacks as they are time-consuming,labour intensive and relatively expensive. Therefore, there is an urgent need for the development of rapid, reliable, precise and non-expensive systems to be used in the food supply and production chain as monitoring devices to detect metabolic alterations in foodstuff. Attention to instrumental detection systems such as electronic noses, electronic tongues and biosensors coupled with chemometric approaches has greatly increased because they have been demonstrated as a promising alternative for the purpose of detecting and monitoring food spoilage. This paper mainly focuses on the recent developments and the application of such multisensor systems in the food industry. Furthermore, the most traditionally methods for food spoilage detection are introduced in this context as well. The challenges and future trends of the potential use of the systems are also discussed. Based on the published literature, encouraging reports demonstrate that such systems are indeed the most promising candidates for the detection and monitoring of spoilage microorganisms in different foodstuff

ANALISIS POLA KELUARAN PROTOTIPE SENSOR RASA PORTABLE CAMPURAN LIPID DIOCTYL PHOSPHATE DAN TRIOCTYL METHYL AMMONIUM CHLORIDE

Portable taste sensor prototype has been fabricated as two channel ion meter and one types of ion selecting membrane. Two channel ion meter were made for pre development of the taste sensor system or as a electronic tongue wich acted as an multi channel ion meter.ion meter was made using IC op-amp LF412 as its amplifier and microcontroller Atmega8535. meanwhile types of the selecting membrane ion were made using variation: mix  of lipid diocthyl phosphate (DOP) and triocthyl methyl ammonium chloride (TOMA) with mass ratio respectively 3:7 and 9:1.  On this sensor, lipid determines membrane selectivity to particular ions. Output from taste sensor is a electric potential and then it could be compared with electrode potential references. This portable taste sensor prototype is then tested is ability to sensing for 5 types of solution: glucose, KCl, HCl, quinine, and MSG. Those 5 types represent 5 basic taste: sweet by glucose, salty by KCL, sour by HCl, bitter by quinine, and umami by MSG. Every solution is made using 1mM KCl  solution and it is variated to be 7 types of concentration. As a result, taste sensor with lipid membrane mixture of 3:7 mixing tend to respon to the anions sample, resulting potential which decreased for every increase in concentration. For membrane with a lipid mixture of 9:1 showed a tendency to respond to the cations sample, resulting potential which increased for every increase in concentration.Key words: taste sensor, ion meter, ion selecting membrane, lipid, diocthyl phosphate, methyl ammonium chloride

음식물 부패도 연구를 위한 높은 신뢰도를 갖는 인공 감각 시스템

학위논문 (박사)-- 서울대학교 대학원 자연과학대학 생물물리 및 화학생물학과, 2017. 8. 전헌수.An artificial sensor based on nanomaterials has been great interest in research on artificial sensory systems because of their excellent sensitivity and selectivity. The fundamental studies of mammalian sensory systems and the hybridization of bio- and nanomaterials are necessary for the development of the remarkable performance of sensors. In this dissertation, we have investigated on the activity of various sensory receptor proteins and related responses. Furthermore, we developed artificial sensory systems for the assessment of food quality with high reliability which could mimic the response of mammalian sensory system. First, we have developed a multiplexed bioelectronics sensor (MBS) that could distinguish various odorants and tastants indicating the food contaminations. We demonstrated that the MBS could monitor the responses of various sensory receptors, showing different binding characteristics. The MBS exhibited a human-like performance in a mixture solution of various target molecules of receptors with 1 pM detection limit. In addition, our sensor platform could recognize food contamination indicators from the real food samples via the combinations of responses of different receptors. Moreover, we developed a highly-stable and oriented nanodiscs (NDs)-based bioelectronic nose (ONBN) for the detection of CV. TAAR13c-embedded nanodiscs (T13NDs) were constructed with TAAR13c produced in E. coli. High-quality T13NDs efficiently mimic native binding pockets and lead highly sensitive and selective detections of CV. Here, the immobilization of T13NDs with a desired orientation on floating electrodes via linker molecules enabled the active binding site to recognize target molecules, which results in high sensitivity and selectivity of our sensor platform. In addition, an ONBN quantitatively detect CV in real food samples by spoilage periods. These results indicate that our ONBN platform based on GPCR-conjugated FET is a new method for the detection of death-associated odor and has a potential on practical bioelectronic sensor applications. Additionally, in the last part of this dissertation, we discussed about the control of enzymatic reaction via nanostructured conducting polymer. We reported a novel bio-chip strategy for control of enzymatic reaction in real-time via electrical stimuli. This technique is named as a bio-switch chip (BSC). We fabricated BSC structures using polypyrrole (Ppy) with entrapped glucose oxidase (GOx) and showed the switching performance of enzymatic reaction in real-time. The introduction of a negative bias voltage on the BSC structure resulted in the 20-folds increased glucose oxidation reaction than that without a bias voltage. Furthermore, we could control the enzymatic reaction on specific regions because the BSC structures could be fabricated on specific regions. In consideration of the fact that enzymes enable useful and versatile to bio-chemical reactions, the capability to control the enzymatic reactions using simple electrical signals could open up various applications in the field of biochips and biochemical industries.Chapter 1 Introduction 1 1.1 Carbon Nanotubes 2 1.2 G protein-coupled receptor 5 1.3 Theoretical Modeling for Bio Sensors 8 1.4 References 11 Chapter 2 Multiplexed Bioelectronic Sensor for Assessment of Food Quality 13 2.1 Introduction 14 2.2 Construction of Multiplexed Bioelectronic Sensor 16 2.3 Characterization of Human Olfactory and Taste Receptors 19 2.4 Operation of MBS Device 21 2.5 Assessment of Food Freshness Using Bioelectronic Sensor 23 2.6 Conclusions 25 2.7 References 27 Chapter 3 Nanodisc-based Bioelectronic Sensor for Assessment of Food Quality 32 3.1 Introduction 33 3.2 Fabrication of oriented nanodisc-functionalized bioelectronics nose using TAAR13c-embedded nanodiscs 36 3.3 Characterization of TAAR13c expressed in HEK-293 cell and high-quality TAAR13c-embedded nanodiscs 37 3.4 Characteristics of an oriented NDs-functionalized CNT-FET with floating electrodes 42 3.5 Operation of ONBN Device 45 3.6 Conclusions 49 3.7 References 51 Chapter 4 Other Works: Bio-switch Chip based on Nanostructured Conducting Polymer and Entrapped Enzyme 55 4.1 Introduction 56 4.2 Structure of Bio-switch Chip 59 4.3 Characterization of Bio-switch Chip 62 4.4 Operation of Bio-switch Chip 64 4.5 Control of Enzymatic Reaction via Bio-switch Chip 69 4.6 Conclusions 72 4.7 References 74 Chapter 5 Conclusion 79 Chapter 6 Abstract in Korean 81Docto

Development of carbon nanotube-based bioelectronic sensors using receptor proteins and peptides for to water and food quality assessment and disease diagnosis

학위논문 (박사)-- 서울대학교 대학원 : 협동과정 바이오엔지니어링전공, 2016. 8. 박태현.Bioelectronic sensors that combine biomolecules as primary recognition elements and electronics as signal transducers have been extensively developed.1, 2 Biologically-inspired technologies for detection of target molecules increased the functionality of sensors with the nature system. Nanomaterial-based electronic devices rapidly amplify biological signals and convert into intuitive forms.3, 4, 5 Furthermore, many parts of living organisms are operated by action potentials so that they can be applied to the sensing mechanism of biolectronic sensors. Recently, various nanomaterials, such as carbon nanotube (CNT), graphene, nanoparticle, nanofiber, nanowire and nanofilm have been used in the fabrication of the bioelectronic sensors. Such nanomaterials were functionalized with biomolecules, such as DNA, protein, peptide, and cell for the specific binding. Advantages of using bioelectronic sensors are low manufacturing cost, small size, fast response, high sensitivity and selectivity. The objective of this research is to develop a CNT-based bioelectronic sensor using receptor proteins and peptides for various applications. Receptors receive external stimuli from out of the cell membrane and activate signal transduction pathways.6 The Receptors play important functions in the human body such as sensory transduction, cellular communication, neural transmission and hormonal signaling.7 In this study, receptor proteins were produced in forms of cell-derived nanovesicles, purified proteins and synthetic peptides. Their biological signals were monitored by The CNT-based transducers and applied to environmental monitoring, disease diagnosis and food quality assessment. For the deorphanization of olfactory receptors, artificial olfactory cells were constructed using heterologous reporter gene systems.8 Specific receptors were screened for detection of potential biomarkers and evaluation of cell surface expression was performed. The cell-derived nanovesicles were constructed for the scalable signal transduction to the CNTs.9 A CNT-based field-effect transistor (FET) was fabricated via the conventional photolithography process.10 CNT channels were functionalized with nanovecsicles and the electrical properties of the CNT-FET were also maintained after the immobilization of nanovesicles. The nanovesicle-based bioelectronic sensor detected target biomarkers with high sensitivity and selectivity. The bioelectronic sensor using receptors presented simple rapid analytical methods for water quality monitoring and diabetes diagnosis. Developing a multiplexed platform and miniaturizing sensing device is necessary for practical applications of bioelectronic sensors. Four kinds of receptor proteins were expressed in Escherichia coli, purified and refolded with the detergent micelle methods. The proteins were immobilized onto the multi-type CNT channels which designed for simultaneous detection of the target molecules. A current monitoring device was customized for the multi-channel CNT-FET and operated by a laptop computer. Complex pattern recognitions of various molecules were available without any interference from non-target molecules. The simple portable bioelectronic device was suitable for efficient assessment of food quality and is expected to be used as a rapid on-site sensing platform. Receptor protein-derived peptides were synthesized based on the sequence of ligand binding sites and applied to the development of the bioelectronic sensor. The peptides do not require cell membrane-like environments so that they were easily stored.11 Furthermore, The synthesized peptides could directly immobilized onto the CNTs using the pi-pi interaction by simple modifications with additional phenylalanine residues.12, 13 The peptide-based bioelectronic sensor detected a serum biomarker of nerumyelitis optica with high specificity. Furthermore, the sensitivity was much higher so that the early-phase detection was available. In this research, bioelectronic sensor using carbon nanotubes and various types of receptors such as nanovesicles, proteins and peptides were developed for simple and rapid detection of the biomarkers. The developed sensors applied for the environmental monitoring, disease diagnosis and food quality assessment.Chapter 1. Research background and objective 1 Chapter 2. Literature review 5 2.1 Concept of bioelectronic sensor 6 2.2 Biomolecules as primary recognition elements 7 2.3 Nanomaterials as secondary signal transducers 9 2.4 Bioelectronic sensors using receptor proteins 12 Chapter 3. Experimental procedures 16 3.1 Gene cloning 17 3.2 Production of receptor proteins 17 3.3 Characterization of receptor proteins 19 3.4 Construction of in vitro disease models 21 3.5 Fabrication of bioelectronic sensor 23 3.6 Electrical measurement 26 Chapter 4. Development of bioelectronic sensor using human olfactory nanovesicles for the detection of odor compounds in water pollution 27 4.1 Introduction 28 4.2 Screening of human olfactory receptors 29 4.3 Characterization of olfactory nanovesicles 32 4.4 Construction of nanovesicle-based bioelectronic nose 34 4.5 Detection of GSM and MIB using bioelectronic nose 36 4.6 Detection of GSM and MIB from water samples 42 4.7 Conclusions 44 Chapter 5. Development of bioelectronic sensor using human olfactory nanovesicles for the detection of VOCs from in vitro diabetic models 45 5.1 Introduction 46 5.2 Construction of in vitro diabetic models 47 5.3 Screening of human olfactory receptors 50 5.4 Construction of nanovesicle-based bioelectronic nose 52 5.5 Detection of 17ODYA using bioelectronic nose 55 5.6 Conclusions 57 Chapter 6. Development of portable and multiplexed bioelectronic sensor using human olfactory and taste receptor proteins for the assessment of food quality 58 6.1 Introduction 59 6.2 Production of human sensory receptors 60 6.3 Fabrication of multi-channel bioelectronic sensor 63 6.4 Detection of odor and taste molecules using bioelectronic sensor 69 6.5 Assessment of food freshness using biotelectronic senso 74 6.6 Conclusions 77 Chapter 7. Development of peptide-based bioelectronic sensor using extracellular loops of aquaporin for the diagnosis of neuromyelitis optica 78 7.1 Introduction 79 7.2 Construction of peptide-based bioelectronic sensor 80 7.3 Electrical characteristics of peptide-based bioelectronic sensor 83 7.4 Detection of AQP4 antibody using bioelectronic sensor 85 7.5 Detection of AQP4 antibody in human serum using bioelectronic sensor 87 7.6 Conclusions 89 Chapter 8. Overall discussion and further suggestions 90 Bibliography 95 Abstract 116Docto