Acetylcholine Sensor Research Articles

An electrochemical sensor for detection of neurotransmitter-acetylcholine using metal nanoparticles, 2D material and conducting polymer modified electrode

An essential biological sensor for acetylcholine (ACh) detection is constructed by immobilizing enzymes, acetylcholinesterase (AChE) and choline oxidase (ChO), on the surface of iron oxide nanoparticles (Fe2O3NPs), poly(3,4-ethylenedioxythiophene) (PEDOT)-reduced graphene oxide (rGO) nanocomposite modified fluorine doped tin oxide (FTO). The qualitative and quantitative measurements of nanocomposites properties were accomplished by scanning electron microscope (SEM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). This prepared biological sensor delineated a wide linear range of 4.0nM to 800μM with a response time less than 4s and detection limit (based on S/N ratio) of 4.0nM. The sensor showed perfect sensitivity, excessive selectivity and stability for longer period of time during storage. Besides its very high-sensitivity, the biosensor has displayed a low detection limit which is reported for the first time in comparison to previously reported ACh sensors. By fabricating Fe2O3NPs/rGO/PEDOT modified FTO electrode for determining ACh level in serum samples, the applicability of biosensor has increased immensely as the detection of the level neurotransmitter is first priority for patients suffering from memory loss or Alzheimer's disease (AD).

Improved enzyme immobilization for enhanced bioelectrocatalytic activity of choline sensor and acetylcholine sensor

In this work, we report the development of a highly sensitive and stable choline/acetylcholine sensor based on the synergetic effect of multi-walled carbon nanotubes and ZnO nanoparticles. This unique multi-layer structure provided a favorable microenvironment to maintain the bioactivity of choline oxidase and acetylcholinesterase, which led to a rapid amperometric response to choline and acetylcholine. This nanomaterials-based choline/acetylcholine sensor was highly sensitive and showed good stability over a relatively long-term storage (90 days). The sensor prepared by loading 1.0-unit choline oxidase and acetylcholinesterase showed a linear response range of 1.0μM to 0.8mM and 1.0μM to 1.0mM, a sensitivity of 178μAmM−1cm−2 and 180μAmM−1cm−2, and a detection limit of 0.3μM, respectively. The sensor prepared was used to detect choline in human blood plasma samples, and the obtained results were consistent with those of the HPLC–MS/MS assay.

An electrochemical acetylcholine sensor based on lichen-like nickel oxide nanostructure

Lichen-like nickel oxide nanostructure was synthesized by a simple method and characterized. The nanostructure was then applied to modify a carbon paste electrode and for the fabrication of a sensor, and the electrocatalytic oxidation of acetylcholine (ACh) on the modified electrode was investigated. The electrocatalytic efficiency of the nickel oxide nanostructure was compared with nickel micro- and nanoparticles, and the lichen-like nickel oxide nanostructure showed the highest efficiency. The mechanism and kinetics of the electrooxidation process were investigated by cyclic voltammetry, steady-state polarization curve and chronoamperometry. The catalytic rate constant and the charge transfer coefficient of ACh electrooxidation by the active nickel species, and the diffusion coefficient of ACh were reported. A sensitive and time-saving hydrodynamic amperometry method was developed for the determination of ACh. ACh was determined with a sensitivity of 392.4mAM−1cm−2 and a limit of detection of 26.7μM. The sensor had the advantages of simple fabrication method without using any enzyme or reagent and immobilization step, high electrocatalytic activity, very high sensitivity, long-term stability, and antifouling surface property toward ACh and its oxidation product.

Non-enzymatic acetylcholine sensor based on Ni–Al layered double hydroxides/ordered mesoporous carbon

In this work, a novel non-enzymatic acetylcholine (ACh) sensor was constructed based on electrodepoition of Ni–Al layered double hydroxides (Ni–Al LDHs) on ordered mesoporous carbon (OMC) modified glassy carbon (GC) electrode. The obtained Ni–Al LDHs/OMC nanocomposite was characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD) and electrochemical methods. A novel non-enzymatic acetylcholine (ACh) sensor based on Ni–Al LDHs/OMC was developed. Compared with the pristine OMC and Ni–Al LDHs modified electrodes, Ni–Al LDHs/OMC displayed higher electrocatalytic activity toward ACh and gave a wide concentration interval of 2–4922μM and a low limit of detection (42nM, R=0.999), which is better than some reports in the literatures. The sensor also exhibited long-term stability and remarkable antifouling property. The excellent electrocatalysis for ACh can be attributed to the combination of the unique properties of Ni–Al LDHs nanoparticles and the ordered mesostructure of OMCs matrix.

Optical acetylcholine sensor based on free base porphyrin as a chromoionophore

In this work, the possibility of application of free base porphyrin as a lipophilic pH chromoionophore for the preparation of optical cation-selective sensors was investigated. The properties of polymeric membranes, containing porphyrins of different structures, namely tetraphenylporphyrin (TPP) and octaethylporphyrin (OEP), were compared. Changes in equilibrium between protonated and deprotonated form of porphyrin, resulting from variations in ACh concentration, were evaluated. The influence of various factors (kind and quantity of anionic additive and porphyrin in the membrane phase, pH of sample solution) on initial equilibrium was studied. The best membrane composition was chosen as: TPP 3 wt.%, KTFPB 175 mol.% relative to ionophore, PVC:o-NPOE (1 : 4) and measuring buffer solution: 0.05 M MES, pH 4.5. Selectivity, response stability, reversibility and repeatability tests were carried out for chosen sensor. Developed sensor allowed for the determination of a model analyte, acetylcholine, at the concentration range of 10(-5) to 10(-2) M, both in stationary and flow-injection system. Sensor response was reversible and repeatable in the mentioned concentration range.

Amperometric biosensor based on enzymes immobilized in hybrid mesoporous membranes for the determination of acetylcholine

Acetylcholine sensor is successfully prepared by using immobilized enzymes, i.e., acetylcholinesterase and choline oxidase within separate hybrid mesoporous silica membranes with 12 nm pore diameter (F127M). The measurement was based on the detection of hydrogen peroxide produced by two sequential enzyme reactions. The determination range and the response time are 6.0–800 μM and within approximately 3 min, respectively. The sensor is very stable compared to free enzymes and 80% of the initial response was maintained even after storage for 80 days. These results show that two enzymes are successfully immobilized and well stabilized, and at the same time, two sequential enzyme reactions efficiently proceed within the separate hybrid mesoporous membranes. Further, we studied the possible detection of organophosphorus pesticides in terms of the inhibition of acetylcholinesterase activity, i.e., the decrease of current response, and demonstrated that the nanomolar concentrations of pesticide (DZN-oxon) can be detected with our sensor.

Development of a highly-sensitive acetylcholine sensor using a charge-transfer technique on a smart biochip

We have developed a highly-sensitive acetylcholine (ACh) sensor based on a charge-transfer technique (CTT) using a smart biochip. The CCT ACh sensor (CTTAS) involves a charge-accumulation technique for detecting choline and H + ions, the sensing signals being amplified without an external amplifier. The CTTAS exhibits excellent performance for ACh detection with high sensitivity (11.6 mV/mM), high signal-to-noise ratio, large span (537 mV), wide linear range (1.5∼12.5 mM), fast response time (<20 s) and very good reproducibility. We obtained a very low limit of detection (∼0.01 mM). Under optimal conditions, the CTTAS outstripped the performance of the widely-used enzyme field-effect transistor (ENFET) with sensitivity more than three times greater than that of the ENFET (3.6 mV/mM). The CTTAS exhibited good selectivity, sensitivity, reproducibility and stability over a large concentration range. Due to its high resolution, fast response, small size and low cost, the CTTAS has tremendous potential for applications in medical research and clinical diagnosis.

A thin-film transistor based acetylcholine sensor using self-assembled carbon nanotubes and SiO2 nanoparticles

Abstract A high sensitivity acetylcholine (ACh) sensor is developed using a nanomaterial-based thin-film transistor. The device fabrication combines the “bottom-up” layer-by-layer self-assembly and the “top-down” microfabrication techniques. The transistor uses a self-assembled single-walled carbon nanotube (SWNT) multilayer as the semiconducting film. Silicon dioxide (SiO 2 ) nanoparticles are deposited on the substrate as the dielectric material. Acetylcholinesterase (AChE) enzyme molecules are immobilized on the surface as the sensing film, which can induce the ACh hydrolysis reaction and release hydrogen ions to the solution. Because all the assembly steps of the multilayer films are done in solutions at room temperature, the fabrication complexity and the process cost are dramatically reduced. The transistor-based sensor demonstrates high sensitivity for ACh sensing and shows a good linearity in the high ACh concentration range. The sensitivity, resolution, and response time of the sensor are measured as 378.2 μA/decade, 10 nM, and 15 s, respectively. This work presents a promising technique to develop disposable and high-sensitivity biosensors for a wide range of applications.

A High-Resolution Amperometric Acetylcholine Sensor Based on Nano-Assembled Carbon Nanotube and Acetylcholinesterase Thin Films

We demonstrate a carbon nanotube based high-resolution biosensor for acetylcholine sensing. Carbon nanotubes are deposited on a silicon wafer in a repeated fashion with layer-by-layer nano self-assembly technique. With nano-assembled acetylcholinesterase molecules on the surface, the carbon nanotube biosensor is capable of detecting acetylcholine at an ultra-low concentration of 100 pM. The sensitivity of the acetylcholine sensor is measured as 7.2 µA/decade. The real-time measurement shows the response time of the biosensor is approximately 6 sec. Both the carbon nanotube film and the acetylcholinesterase film are crucial in the sensing process. Due to its high resolution, fast response, small size, and low cost, the carbon nanotube biosensor has tremendous potential for applications in medical research and clinical diagnosis.

Electrodeposited Nickel/Platinum Alloy as a Biosensor for Acetyl Choline

AbstractNickel and platinum find extensive use in preparation of biosensors. In the present work, Ni/Pt alloy was plated on graphite to make acetylcholine sensor. The microstructure, surface composition and electrochemical performance of the electrode was analyzed by different techniques. The sensing performance was evaluated by cyclic voltammetry. The prepared alloy plate exhibited very good linear relationship between acetyl choline concentration and response current. The sensitivity and reproducibility of the prepared electrode were found better than other nickel electrodes reported.

Amperometric acetylcholine sensor catalyzed by nickel anode electrode

An amperometric method was using a nickel catalytic electrode in aqueous base solution for detecting acetylcholine (ACh). A sensing mechanism was developed in which ACh was hydrolyzed in base aqueous solution to produce the acetic anion and choline. The alcohol group of choline was oxidized to the corresponding carboxylic acid by Ni(OH) 2/NiOOH catalytic system. The amperometric response resulted from the current generated by ACh oxidation in response to step changes in ACh concentration. The potential window of limiting current of ACh anodic oxidation at the Ni interface was determined in NaOH electrolyte. The effect of NaOH electrolyte concentration on sensitivity was also discussed. At the optimum operating condition, the method exhibits a good linear relationship between the response current and the ACh concentration. The response time of the ACh sensing system was 10 s. Scanning electrochemical microscopy (SECM) with platinum micro-tips was used to investigate the diffusion layer thickness of Ni electrode.

A Theoretical Model for Immobilized Enzyme Inhibition Biosensors

The development and experimental verification of a theoretical model for immobilized enzyme inhibition biosensors is reported. The model predicts that the percentage of inhibited enzyme (I %), after exposure to an inhibitor, is linearly related to both the inhibitor concentration ([I]B) and the square root of incubation time (t1/2): I%=Kt1/2[I]B×100 The model is based on diffusion limited inhibitor transport and takes into account the heterogeneous nature of enzyme inhibition that results from immobilization of an enzyme at the surface of a transducer. Experimental verification of the model was achieved using an electrochemical (amperometric) acetylcholine sensor. The immobilized enzyme used in this study was acetylcholinesterase, while the organophosphorus pesticide, paraoxon, was employed as the inhibitor. A second enzyme, choline oxidase, was co-immobilized to facilitate electrochemical detection of the substrate. To satisfy the conditions required by the proposed model a specially designed biosensing device was fabricated employing a microdisk array as the transducer. Acetylcholinesterase (0.00025 units) and choline (1.0 units) were co-immobilized onto the array using a water-based polyurethane/polyethylene oxide colloidal dispersion matrix. A dialysis membrane was used as an active barrier to prevent fouling of the array and to restrict enzyme leaching. The device was used for both batch and flow injection analysis and experimental results verifying the proposed model are presented using both modes.

On-line electrochemical sensor for selective continuous measurement of acetylcholine in cultured brain tissue.

An on-line acetylcholine (ACh) sensor was developed in order to determine extracellular ACh concentrations without interference from choline (Ch). The sensor is composed of a small-volume enzymatic prereactor (22-microL inner volume) in which choline oxidase and catalase are immobilized in series. Carbon electrodes were modified with an acetylcholine esterase (AChE), choline oxidase (ChOx), and osmium poly(vinylpyridine)-based redox polymer containing horseradish peroxidase (Os-gel-HRP). The sensor sensitivity was 43.7 nA/microM (+/- 0.15, n = 3) for ACh under optimized conditions. Almost no response was seen when 100 microM Ch was continuously injected. The detection limit for ACh with the sensor was comparable to that obtained using liquid chromatography with electrochemical detection combined with an enzymatic reactor. The electrical stimulation of cultured rat hippocampal tissue resulted in an extracellular ACh increase of 20 nM (+/- 11 nM, n = 3). This increase was observed continuously with our online sensor combined with a microcapillary sampling probe located very close to the tissue. The continuous measurement of ACh and Ch using a split disk carbon film dual electrode in which one electrode surface was modified with ChOx/Os-gel-HRP and the other with AChE-ChOx/Os-gel-HRP bilayer film was also demonstrated to improve the response time by eliminating the prereactor.

Avidin-Biotin System-Based Enzyme Multilayer Membranes for Biosensor Applications: Optimization of Loading of Choline Esterase and Choline Oxidase in the Bienzyme Membrane for Acetylcholine Biosensors

The loading of choline esterase (ChE) and choline oxidase (ChOx) in the enzyme membrane of an acetylcholine biosensor was optimized based on a layer-by-layer construction of the bienzyme layers on the surface of a platinum (Pt) and a Pt-black electrode. To this goal, ChE and ChOx were tagged with biotin residues, so that the enzymes could be built into the multilayer assemblies composed of monomolecular layers of the enzymes and avidin. The ChE/ChOx bienzyme multilayer-modified electrodes showed amperometric response to acetylcholine in solution, showing that ChE and ChOx catalyzed hydrolysis reaction of acetylcholine and an oxidation reaction of resulting choline, respectively, successively in the bienzyme layer. It was found that the acetylcholine sensor which is modified with 10-layer ChOx exhibited its maximum response to acetylcholine when additional 2 layers of ChE were added to the surface of the ChOx layer. This optimum ratio of ChE to ChOx in the bienzyme layer was reasonably explained in terms of the relative values of catalytic activity of the enzymes. The usefulness of the layer-by-layer deposition technique in optimizing the enzyme loading in bienzyme system is discussed in detail.

Voltammetric Acetylcholine Sensor using Mercaptohydroquinone Monolayer-Attached Gold Electrode as the Base Transducer

Voltammetric Acetylcholine Sensor using Mercaptohydroquinone Monolayer-Attached Gold Electrode as the Base Transducer

Trends in biosensor research and development

Trends in biosensor research and development are described. Micromachining techniques have been applied to construct biosensor systems. Several detection units for flow injection analysis have been fabricated. An electrochemical flow cell was fabricated, and both an enzyme immobilized column and an electrochemical detector were integrated. A chemiluminescence detector was also fabricated. Biosensor systems for clinical analyses in the toilet have been developed. A quartz crystal oscillator was used for protein detection, and a thermistor was employed as a thermal detector resulting from the enzyme reaction. An injectable biosensor for rapid measurement has been developed, and the determination of sugars in fruits was carried out. Glucose oxidase was immobilized for the determination of glucose, and three enzymes, invertase, mutarotase and glucose oxidase, were immobilized for the sucrose sensor. Microbiosensors using carbon fiber microelectrodes have been applied to neuroscience. Micro acetylcholine sensors based on acetylcholine esterase and choline oxidase were fabricated and characterized. Micro glutamate sensors were also fabricated using platinized carbon fiber electrodes which were modified with glutamate oxidase. The micro glutamate sensor was applied to the detection of glutamate released from the neurons.

電界効果型トランジスタを用いたバイオ化学センサによる農薬類の計測

Acetylcholine sensor based on ISFET as a pH sensor, using the immobilized acetylcholine esterase (AChE) in the BSA membrane are reported for the characteristics and for measuring chemicals. We attempted the application of an enzyme sensor based on ISFET as an effective and simple potentiometric sensor for the detection of chemicals, as irreversible and reversible inhibitors. New data on the analysis of chemicals are reported using the present acetylcholine sensor based on the inhibition of the immobilized AChE. Reactivation of immobilized AChE after inhibition with reversible and irreversible inhibitors, was carried out using a mixture of acetylcholine and pyridine-2-aldoxime methiodide (PAM). The detection for PHC and Nicotine as reversible inhibitors, and Malathion and Dimethoate as irreversible inhibitors was found to be in the ranges of 5×10-5-5×10-4 M, 10-5-10-4 M, 10-6-10-4 and 2.5×10-5-10-3 M, respectively.The present study was also developed for an electrochemical sensor for toxic substances which works by measuring the inhibition of the enzyme activity. We suggested that these inhibition rates (e.g., I50) by the biosensor may be important as a biological index to biological systems with regard to toxic matters.

ポリビニルブチラール膜を用いるＦＥＴバイオセンサー

The present review describes the FET-biosensor using polyvinylbutyral (PVB) membrane. The PVB membrane was applied to the surface of the silicon nitride which is the gate insulator of the IS-FET (Ion Sensitive Field Effect Transistor). The membrane exhibited good adhesive properties and was used for biomolecules immobilization, such as urease. H+-ATPase, etc.. The water content of the membrane was 27.6% with a thickness of 23, am. The surface structure of the membrane was observed by scanning electron microscopy, which revealed a regular porous microfilter arrangement with a mean pore diameter of 0.25 μm. Immobilized urease activity in this membrane was approximately 20% higher than when a urease-triacetylcellulose membrane was used. Using this polyvinylbutyral membrane, urea sensor, ATP sensor, acetylcholine sensor and HSA sensor were constructed. The new type device of ISFET was also constructed from amorphous silicon. From this device and PVB membrane, glucose sensor, hypoxanthine sensor and inosine sensor were constructed.

Kinetic model of pH-based potentiometric enzymic sensors. Part 2. Method of fitting

Methods of fitting theoretical calibration graphs, evaluated on the basis of a kinetic model of pH-based enzymic sensors, to experimental data obtained using glucose and acetylcholine sensors are presented. Bisection and simplex methods were used for this purpose. Good agreement between the theoretical predictions and the experimental results was achieved.

Ultramicro acetylcholine sensor based on an enzyme-modified carbon fibre electrode

Ultramicro acetylcholine sensor based on an enzyme-modified carbon fibre electrode