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We report a magnetically-focusing biochip structure enabling a single layered magnetic trap-and-release cycle for biosensors with an improved detection speed and selectivity. Here, magnetic beads functionalized with specific receptor molecules were utilized to trap target molecules in a solution and transport actively to and away from the sensor surfaces to enhance the detection speed and reduce the non-specific bindings, respectively. Using our method, we demonstrated the high speed detection of IL-13 antigens with the improved detection speed by more than an order of magnitude. Furthermore, the release step in our method was found to reduce the non-specific bindings and improve the selectivity and sensitivity of biosensors. This method is a simple but powerful strategy and should open up various applications such as ultra-fast biosensors for point-of-care services.
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We developed a magnetically-refreshable receptor platform structure which can be integrated with quite versatile nano-biosensor structures to build reusable nano-biosensor chips. This structure allows one to easily remove used receptor molecules from a biosensor surface and reuse the biosensor for repeated sensing operations. Using this structure, we demonstrated reusable immunofluorescence biosensors. Significantly, since our method allows one to place receptor molecules very close to a nano-biosensor surface, it can be utilized to build reusable carbon nanotube transistor-based biosensors which require receptor molecules within a Debye length from the sensor surface. Furthermore, we also show that a single sensor chip can be utilized to detect two different target molecules simply by replacing receptor molecules using our method. Since this method does not rely on any chemical reaction to refresh sensor chips, it can be utilized for versatile biosensor structures and virtually-general receptor molecular species.
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Técnicas Biossensoriais/instrumentação , Imunofluorescência/instrumentação , Imãs/química , Nanotubos de Carbono/química , Anticorpos Imobilizados/química , Desenho de Equipamento , Fluorimunoensaio/instrumentação , Humanos , Interleucina-10/análise , Interleucina-4/análise , Magnetismo/instrumentação , Transistores EletrônicosRESUMO
Ethyl pyruvate (EP), a simple aliphatic ester of pyruvic acid, has been shown to have antiinflammatory effects and to confer protective effects in various pathological conditions. Recently, a number of studies have reported EP inhibits high mobility group box 1 (HMGB1) secretion and suggest this might contribute to its antiinflammatory effect. Since EP is used in a calcium-containing balanced salt solution (Ringer solution), we wondered if EP directly chelates Ca(2+) and if it is related to the EP-mediated suppression of HMGB1 release. Calcium imaging assays revealed that EP significantly and dose-dependently suppressed high K(+)-induced transient [Ca(2+)]i surges in primary cortical neurons and, similarly, fluorometric assays showed that EP directly scavenges Ca(2+) as the peak of fluorescence emission intensities of Mag-Fura-2 (a low-affinity Ca(2+) indicator) was shifted in the presence of EP at concentrations of ≥7 mmol/L. Furthermore, EP markedly suppressed the A23187-induced intracellular Ca(2+) surge in BV2 cells and, under this condition, A23187-induced activations of Ca(2+)-mediated kinases (protein kinase Cα and calcium/calmodulin-dependent protein kinase IV), HMGB1 phosphorylation and subsequent secretion of HMGB1 also were suppressed. (A23187 is a calcium ionophore and BV2 cells are a microglia cell line.) Moreover, the above-mentioned EP-mediated effects were obtained independent of cell death or survival, which suggests that they are direct effects of EP. Together, these results indicate that EP directly chelates Ca(2+), and that it is, at least in part, responsible for the suppression of HMGB1 release by EP.
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Cálcio/metabolismo , Proteína HMGB1/antagonistas & inibidores , Piruvatos/farmacologia , Animais , Calcimicina/farmacologia , Linhagem Celular , Células Cultivadas , Proteína HMGB1/metabolismo , Camundongos , N-Metilaspartato/farmacologia , Neurônios/efeitos dos fármacos , FosforilaçãoRESUMO
We investigated the photoconductive characteristics of molybdenum disulfide (MoS2) field-effect transistors (FETs) that were fabricated with mechanically exfoliated multi-layer MoS2 flakes. Upon exposure to UV light, we observed an increase in the MoS2 FET current because of electron-hole pair generation. The MoS2 FET current decayed after the UV light was turned off. The current decay processes were fitted using exponential functions with different decay characteristics. Specifically, a fast decay was used at the early stages immediately after turning off the light to account for the exciton relaxation, and a slow decay was used at later stages long after turning off the light due to charge trapping at the oxygen-related defect sites on the MoS2 surface. This photocurrent decay phenomenon of the MoS2 FET was influenced by the measurement environment (i.e., vacuum or oxygen environment) and the electrical gate-bias stress conditions (positive or negative gate biases). The results of this study will enhance the understanding of the influence of environmental and measurement conditions on the optical and electrical properties of MoS2 FETs.
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We presented a scalable fabrication method for the preparation of multilayered nano-prism vertex (NV)-tips whose dimensions can be controlled for tip-enhanced Raman spectroscopy (TERS). The NV-tip had sharp vertices (diameter ~20 nm) originated from the chemical lift-off process after the angle-grinding process, enabling high resolution imaging. TERS measurements were performed on brilliant cresyl blue (BCB) molecules using a Ag/Au NV-tip, revealing the enhanced field localization at the vertices of the NV-tip. Furthermore, we could observe the polarization effect of the NV-tip. Our NV-tips should be a powerful tool for basic research on TERS experiments and SPM applications.
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Nanotecnologia/instrumentação , Nanotecnologia/métodos , Análise Espectral Raman/instrumentação , Análise Espectral Raman/métodosRESUMO
We investigated the effects of passivation on the electrical characteristics of molybdenum disulfide (MoS(2)) field effect transistors (FETs) under nitrogen, vacuum, and oxygen environments. When the MoS(2) FETs were exposed to oxygen, the on-current decreased and the threshold voltage shifted in the positive gate bias direction as a result of electrons being trapped by the adsorbed oxygen at the MoS(2) surface. In contrast, the electrical properties of the MoS(2) FETs changed only slightly in the different environments when a passivation layer was created using polymethyl methacrylate (PMMA). Specifically, the carrier concentration of unpassivated devices was reduced to 6.5 × 10(15) cm(-2) in oxygen from 16.3 × 10(15) cm(-2) in nitrogen environment. However, in PMMA-passivated devices, the carrier concentration remained nearly unchanged in the range of 1-3 × 10(15) cm(-2) regardless of the environment. Our study suggests that surface passivation is important for MoS(2)-based electronic devices.
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We developed an olfactory-nanovesicle-fused carbon-nanotube-transistor biosensor (OCB) that mimics the responses of a canine nose for the sensitive and selective detection of hexanal, an indicator of the oxidation of food. OCBs allowed us to detect hexanal down to 1 fM concentration in real-time. Significantly, we demonstrated the detection of hexanal with an excellent selectivity capable of discriminating hexanal from analogous compounds such as pentanal, heptanal, and octanal. Furthermore, we successfully detected hexanal in spoiled milk without any pretreatment processes. Considering these results, our sensor platform should offer a new method for the assessment of food quality and contribute to the development of portable sensing devices.
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Biomimética/métodos , Técnicas Biossensoriais/métodos , Análise de Alimentos/métodos , Nanotecnologia/métodos , Nanotubos de Carbono/química , Nariz , Transistores Eletrônicos , Aldeídos/química , Animais , Biomimética/instrumentação , Técnicas Biossensoriais/instrumentação , Cães , Análise de Alimentos/instrumentação , Contaminação de Alimentos/análise , Células HEK293 , Humanos , Leite/química , Nanotecnologia/instrumentação , Fatores de TempoRESUMO
INTRODUCTION: this study aimed to investigate the long-term clinical efficacy and satisfaction degree of integrative Korean medicine (KM) treatment for patients with failed back surgery syndrome (FBSS). METHODS: we performed a follow-up questionnaire survey and retrospective analysis of medical records for patients with FBSS who underwent inpatient treatment for ≥ 1 week. The primary evaluation indices were numeric rating scale (NRS) scores for low back pain (LBP) and leg pain at admission and discharge. Sub-evaluation indices included the Oswestry Disability Index (ODI) and EuroQol 5-dimension (EQ-5D) score. The follow-up questionnaire survey obtained information regarding previous surgeries; reasons for satisfaction/dissatisfaction with surgical and KM treatment; and current status. RESULTS: compared with at admission, there was a significant post-treatment decrease in the NRS scores for LBP and leg pain, as well as the ODI score. Further, there was a significant post-treatment increase in the EQ-5D score. Regarding the patients' global impression of change for KM treatment administered during admission and at the follow-up questionnaire survey, 101 (95.3%) patients selected "minimally improved" or better. CONCLUSION: integrative KM treatment could effectively reduce pain, as well as improve function and health-related quality of life, in patients with FBSS.
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Olfactory receptor (OR) genes are considered to be the largest superfamily of the mammalian genome, and in the case of humans, approximately 390 kinds of functional ORs play a role in perceiving odors. In spite of their significance in olfaction, the function of all ORs has not yet been fully revealed. In order to efficiently identify specific ligands of orphan ORs, methods that can generate olfactory signals in a reliable manner and that can convert the cellular signals into measurable responses are required. Here, we describe an OR screening assay method using olfactory sensors that are based on cell-derived nanovesicles combined with single-walled carbon nanotube field-effect transistors (SWNT-FETs). The nanovesicles contain ORs on their surface membrane and induce influx of calcium ions similar to olfactory signal transduction. This ion influx causes an electrical current change along the carbon nanotube, and then this change is measured by the SWNT-FET sensor. This technique facilitates the simple and rapid screening of OR functions.
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Técnicas Biossensoriais , Ensaios de Triagem em Larga Escala , Nanotubos de Carbono/química , Receptores Odorantes/genética , Proteínas Recombinantes de Fusão/genética , Rodopsina/genética , Animais , Bovinos , Citocalasina B/química , Expressão Gênica , Células HEK293 , Humanos , Microtecnologia , Plasmídeos/química , Plasmídeos/metabolismo , Receptores Odorantes/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Rodopsina/metabolismo , Silício/química , Transfecção , Transistores EletrônicosRESUMO
A simple but efficient measurement platform based on ion-channel-coupled receptors and nanovesicles was developed for monitoring the real-time activity of G-protein-coupled receptors (GPCRs). In this work, an olfactory receptor (OR), the most common class A GPCR, was covalently fused with a Kir6.2 channel so that the GPCR action directly induced the opening of the ion channels and changes in the electrical membrane potential without complex cellular signaling processes. This strategy reduced the measurement errors caused by instability of various cellular components. In addition, rather than using whole cells, a cell-surface-derived nanovesicle was used to preserve the membrane-integrated structure of GPCRs and to exclude case-dependent cellular conditions. Another merit of using the nanovesicle is that nanovesicles can be easily combined with nanomaterial-based field-effect transistors (FETs) to build a sensitive and stable measurement platform to monitor GPCR activities with high sensitivity in real-time. Using a platform based on carbon nanotube FETs and nanovesicles carrying Kir6.2-channel-coupled ORs, we monitored the real-time response of ORs to their ligand molecules. Significantly, since this platform does not rely on rather unstable cell signaling pathways, our platform could be utilized for a rather long time period without losing its functionality. This system can be utilized extensively for simple and sensitive analysis of the activities of various GPCRs and should enable various academic and practical applications.
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Nanotecnologia/métodos , Canais de Potássio Corretores do Fluxo de Internalização/química , Receptores Odorantes/química , Receptores Odorantes/metabolismo , Células HEK293 , Humanos , Ativação do Canal Iônico , Ligantes , Potenciais da Membrana , Modelos Moleculares , Nanotecnologia/instrumentação , Nanotubos de Carbono/química , Conformação Proteica , Fatores de Tempo , Transistores EletrônicosRESUMO
A bioelectronic nose based on a novel microfluidic system (µBN) was fabricated to detect gaseous trimethylamine (TMA) in real-time. Single-walled carbon nanotube-field effect transistors (SWNT-FETs) were functionalized with olfactory receptor-derived peptides (ORPs) that can recognize the TMA molecules. The ORP-coated SWNT-FETs were assembled with a microfluidic channel and were sealed with top and bottom frames. This simple process was used to complete the µBNs, and a well-defined condition was achieved to detect the gaseous molecules. The µBNs allowed us to detect gaseous TMA molecules down to 10 parts per trillion (ppt) in real-time and showed high selectivity when distinguishing gaseous TMA from other gaseous odorants. The sensor was used to determine the quality of seafood (oysters), and spoiled seafood and other types of spoiled foods were also successfully discriminated without any pretreatment processes. These results indicate that portable-scale platforms can be manufactured by using µBNs and can be applicable for real-time on-site gas analysis.
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Técnicas Biossensoriais/instrumentação , Análise de Alimentos/instrumentação , Gases/análise , Metilaminas/análise , Microfluídica/instrumentação , Alimentos Marinhos/análise , Sequência de Aminoácidos , Animais , Desenho de Equipamento , Qualidade dos Alimentos , Gases/metabolismo , Proteínas Imobilizadas/química , Proteínas Imobilizadas/metabolismo , Limite de Detecção , Metilaminas/metabolismo , Dados de Sequência Molecular , Nanotubos de Carbono/química , Peptídeos/química , Peptídeos/metabolismo , Receptores Odorantes/química , Receptores Odorantes/metabolismo , Transistores EletrônicosRESUMO
We report on the conformal surface passivation of photonic crystal (PC) laser devices with an ultrathin dielectric layer. Air-bridge-type Γ-point band-edge lasers (BELs) are fabricated by forming a honeycomb lattice two-dimensional PC structure into an InGaAsP multiple-quantum-well epilayer. Atomic layer deposition (ALD) is employed for conformal deposition of a few-nanometer-thick SiO2 layer over the entire device surface, not only on the top and bottom surfaces of the air-bridge membrane but also on the air-hole sidewalls. Despite its extreme thinness, the ALD passivation layer is found to protect the InGaAsP BEL devices from harsh chemicals. In addition, the ALD-SiO2 is compatible with the silane-based surface chemistry, which allows us to use ALD-passivated BEL devices as label-free biosensors. The standard streptavidin-biotin interaction shifts the BEL lasing wavelength by â¼1 nm for the dipole-like Γ-point band-edge mode. A sharp lasing line (<0.2 nm, full width at half-maximum) and a large refractive index sensitivity (â¼163 nm per RIU) produce a figure of merit as high as â¼800 for our BEL biosensor, which is at least an order of magnitude higher than those of more common biosensors that rely on a broad resonance peak, showing that our nanolaser structures are suitable for highly sensitive biosensor applications.
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Técnicas Biossensoriais , Nanotecnologia/métodos , Dióxido de Silício/química , Biotina/química , Cristalização , Desenho de Equipamento , Lasers , Luz , Nanoestruturas , Óptica e Fotônica , Fótons , Refratometria , Silanos/química , Estreptavidina/química , Propriedades de Superfície , VibraçãoRESUMO
A bioelectronic nose for the real-time assessment of water quality was constructed with human olfactory receptor (hOR) and single-walled carbon nanotube field-effect transistor (swCNT-FET). Geosmin (GSM) and 2-methylisoborneol (MIB), mainly produced by bacteria, are representative odor compounds and also indicators of contamination in the water supply system. For the screening of hORs which respond to these compounds, we performed CRE-luciferase assays of the two odorants in heterologous cell system. Human OR51S1 for GSM and OR3A4 for MIB were selected, and nanovesicles expressing the hORs on surface were produced from HEK-293 cell. Carbon nanotube field-effect transistor was functionalized with the nanovesicles. The bioelectronic nose was able to selectively detect GSM and MIB at concentrations as low as a 10 ng L(-1). Furthermore, detection of these compounds from the real samples such as tap water, bottled water and river water was available without any pretreatment processes.
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Técnicas Biossensoriais/instrumentação , Canfanos/análise , Nariz Eletrônico , Naftóis/análise , Odorantes/análise , Poluição da Água/análise , Canfanos/metabolismo , Desenho de Equipamento , Células HEK293 , Humanos , Proteínas Imobilizadas/metabolismo , Nanotubos de Carbono/química , Nanotubos de Carbono/ultraestrutura , Naftóis/metabolismo , Receptores Odorantes/metabolismo , Qualidade da ÁguaRESUMO
We report a reusable floating-electrode sensor based on aligned semiconducting single-walled carbon nanotubes for the quantitative monitoring of the electrophysiological responses from a nonadherent cell. This method allowed us to monitor and distinguish the real-time responses from normal and small-cell lung cancer (SCLC) cells to the addition of nicotine. The difference was attributed to the overexpressed nicotinic acetylcholine receptors (nAChRs) in the SCLC cells. The sensor was also utilized to monitor the effect of various drugs on the cells. The treatment with inhibitors such as genistin or daidzein was found to reduce Ca(2+) influx in SCLC cells. Moreover, tamoxifen, though known as the antiestrogen compound, was found to only partly block the binding of daidzein to nAChRs. Significantly, the activities of multiple individual cells could be measured repeatedly using a single sensor device, enabling statistically meaningful measurements without errors from the device-to-device variations of the sensor characteristics. This capability of the quantitative monitoring of nonadherent cells should be a major breakthrough for electrophysiology research and various biomedical applications such as drug screening and therapeutic monitoring.
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Eletrofisiologia/instrumentação , Neoplasias Pulmonares/patologia , Carcinoma de Pequenas Células do Pulmão/patologia , Linhagem Celular Tumoral , Eletrodos , Reutilização de Equipamento , Humanos , Nanotubos de Carbono/química , Nicotina/farmacologia , Receptores Nicotínicos/metabolismo , SemicondutoresRESUMO
A human nose-mimetic diagnosis system that can distinguish the odor of a lung cancer biomarker, heptanal, from human blood is presented. Selective recognition of the biomarker is mimicked in the human olfactory system. A specific olfactory receptor recognizing the chemical biomarker is first selected through screening a library of human olfactory receptors (hORs). The selected hOR is expressed on the membrane of human embryonic kidney (HEK)-293 cells. Nanovesicles containing the hOR on the membrane are produced from these cells, and are then used for the functionalization of single-walled carbon nanotubes. This strategy allows the development of a sensitive and selective nanovesicle-based bioelectronic nose (NvBN). The NvBN is able to selectively detect heptanal at a concentration as low as 1 × 10(-14) m, a sufficient level to distinguish the blood of a lung cancer patient from the blood of a healthy person. In actual experiments, NvBN could detect an extremely small increase in the amount of heptanal from human blood plasma without any pretreatment processes. This result offers a rapid and easy method to analyze chemical biomarkers from human blood in real-time and to diagnose lung cancer.
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Técnicas Biossensoriais/instrumentação , Nariz Eletrônico , Neoplasias Pulmonares/diagnóstico , Nanotubos de Carbono/química , Aldeídos/química , Aldeídos/isolamento & purificação , Biomarcadores/sangue , Desenho de Equipamento , Células HEK293 , Humanos , Odorantes/análise , Receptores Odorantes/metabolismoRESUMO
We investigated the effect of irradiation on molybdenum disulfide (MoS2) field effect transistors with 10 MeV high-energy proton beams. The electrical characteristics of the devices were measured before and after proton irradiation with fluence conditions of 10(12), 10(13), and 10(14) cm(-2). For a low proton beam fluence condition of 10(12) cm(-2), the electrical properties of the devices were nearly unchanged in response to proton irradiation. In contrast, for proton beam fluence conditions of 10(13) or 10(14) cm(-2), the current level and conductance of the devices significantly decreased following proton irradiation. The electrical changes originated from proton-irradiation-induced traps, including positive oxide-charge traps in the SiO2 layer and trap states at the interface between the MoS2 channel and the SiO2 layer. Our study will enhance the understanding of the influence of high-energy particles on MoS2-based nanoelectronic devices.
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Aquaporin-4 (AQP4) water channel protein transports water molecules across cell membranes bidirectionally and involves in a neurological disorder, neuromyelitis optica (NMO) caused by anti-AQP4 antibodies. Here, we developed a platform based on nanovesicle-carbon nanotube hybrid nanostructures for the real-time detection of anti-AQP4 antibodies and the electrophysiological monitoring of AQP4 activities. Using the hybrid device, we could detect anti-AQP4 antibodies with a high sensitivity and estimate the binding constants under different osmotic conditions. The results show AQP4 had a better affinity to anti-AQP4 antibodies under hyper-osmotic conditions than normal conditions. Furthermore, our device can be utilized to study the real-time cellular responses related with AQP4 such as those to different osmotic stresses. This nanovesicle-based platform can be a simple but versatile tool for basic research about AQP4 and related biomedical applications such as disease diagnostics.
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Anticorpos/análise , Aquaporina 4/análise , Técnicas Biossensoriais/instrumentação , Nanoestruturas/química , Nanotubos de Carbono/química , Anticorpos/imunologia , Aquaporina 4/imunologia , Membrana Celular , Fenômenos Eletrofisiológicos , Desenho de Equipamento , Células HEK293 , Humanos , Neuromielite Óptica/imunologia , Pressão OsmóticaRESUMO
We report the development of "nano-storage wires" (NSWs), which can store chemical species and release them at a desired moment via external electrical stimuli. Here, using the electrodeposition process through an anodized aluminum oxide template, we fabricated multisegmented nanowires composed of a polypyrrole segment containing adenosine triphosphate (ATP) molecules, a ferromagnetic nickel segment, and a conductive gold segment. Upon the application of a negative bias voltage, the NSWs released ATP molecules for the control of motor protein activities. Furthermore, NSWs can be printed onto various substrates including flexible or three-dimensional structured substrates by direct writing or magnetic manipulation strategies to build versatile chemical storage devices. Since our strategy provides a means to store and release chemical species in a controlled manner, it should open up various applications such as drug delivery systems and biochips for the controlled release of chemicals.
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Trifosfato de Adenosina/química , Nanotecnologia/métodos , Nanofios/química , Polímeros/química , Pirróis/química , Óxido de Alumínio/química , Técnicas Biossensoriais , Biotecnologia , Sistemas de Liberação de Medicamentos , Eletroquímica , Eletrodos , Galvanoplastia , Ouro/química , Magnetismo , Microscopia Eletrônica de Varredura , Níquel/química , Água/químicaRESUMO
We developed a "chemical-pain sensor" that could recognize chemical pain stimuli such as capsaicin and resiniferatoxin just like mammalian chemical pain sensory systems. Here, we first prepared nanovesicles containing rat pain sensory receptor, rat transient receptor potential vanilloid 1 (rTRPV1), which is activated by noxious heat and capsaicin. And the nanovesicles were immobilized on a single-walled carbon nanotube-based field effect transistor. The chemical-pain sensor could selectively detect chemical pain stimuli with a high sensitivity of a 1 pM detection limit. It also responded to different chemical pain stimuli in a manner similar as to that of mammalian chemical pain sensory systems. This sensor platform can be utilized for various practical applications such as food screening tools and artificial somesthetic sensors. Moreover, TRP families have been suggested as potential drug targets related to nerve and circulation disorders. Thus, the capability of monitoring TRP responses using our sensor platforms should provide a powerful means for the development of new drugs as well as the basic research about nerve and circulation systems.