Rapid, Sensitive Detection of Neurotransmitters at Microelectrodes Modified with Self-assembled SWCNT Forests

Carbon nanotube (CNT) modification of microelectrodes can result in increased sensitivity without compromising time response. However, dip coating CNTs is not very reproducible and the CNTs tend to lay flat on the electrode surface which limits access to the electroactive sites on the ends. In this study, aligned CNT forests were formed using a chemical self-assembly method, which resulted in more exposed CNT ends to the analyte. Shortened, carboxylic acid functionalized single-walled CNTs were assembled from a dimethylformamide (DMF) suspension onto a carbon-fiber disk microelectrode modified with a thin iron hydroxide-decorated Nafion film. The modified electrodes were highly sensitive, with 36-fold higher oxidation currents for dopamine using fast-scan cyclic voltammetry than bare electrodes and 34-fold more current than electrodes dipped in CNTs. The limit of detection (LOD) for dopamine was 17 ± 3 nM at a 10 Hz repetition rate and 65 ± 7 nM at 90 Hz. The LOD at 90 Hz was the same as a bare electrode at 10 Hz, allowing a 9-fold increase in temporal resolution without a decrease in sensitivity. Similar increases were observed for other cationic catecholamine neurotransmitters, and the increases in current were greater than for anionic interferents such as ascorbic acid and 3,4-dihydroxyphenylacetic acid (DOPAC). The CNT forest electrodes had high sensitivity at 90 Hz repetition rate when stimulated dopamine release was measured in Drosophila . The sensitivity, temporal resolution, and spatial resolution of these CNT forest modified disk electrodes facilitate enhanced electrochemical measurements of neurotransmitter release in vivo.     

杂多酸修饰多晶铂电极的制备及其电催化性能的研究

为了改善铂基催化剂氧化甲醇的催化活性,首先通过循环伏安扫描制备了硅钨酸、磷钨酸、磷钼酸三种杂多酸修饰铂电极。通过研究铂电极修饰前后在硫酸底液中的循环伏安行为可知,尽管杂多酸具有较大的分子构型,但仍能在铂电极上吸附．另外通过循环伏安曲线研究了杂多酸修饰铂电极对甲醇氧化的电催化作用及抗一氧化碳毒化作用．测试结果表明：铂电极经杂多酸修饰后,能够大大提高其对甲醇氧化反应的催化活性以及抗一氧化碳毒化作用．并且三种杂多酸修饰铂电极中,磷钼酸修饰铂电极的催化活性最高,抗毒化作用最强．     

Spatially and temporally resolved single-cell exocytosis utilizing individually addressable carbon microelectrode arrays

We report the fabrication and characterization of carbon microelectrode arrays (MEAs) and their application to spatially and temporally resolve neurotransmitter release from single pheochromocytoma (PC12) cells. The carbon MEAs are composed of individually addressable 2.5-mum-radius microdisks embedded in glass. The fabrication involves pulling a multibarrel glass capillary containing a single carbon fiber in each barrel into a sharp tip, followed by beveling the electrode tip to form an array (10-20 microm) of carbon microdisks. This simple fabrication procedure eliminates the need for complicated wiring of the independent electrodes, thus allowing preparation of high-density individually addressable microelectrodes. The carbon MEAs have been characterized using scanning electron microscopy, steady-state and fast-scan voltammetry, and numerical simulations. Amperometric results show that subcellular heterogeneity in single-cell exocytosis can be electrochemically detected with MEAs. These ultrasmall electrochemical probes are suitable for detecting fast chemical events in tight spaces, as well as for developing multifunctional electrochemical microsensors.     

Flame etching enhances the sensitivity of carbon-fiber microelectrodes

Small sensors are useful for in vivo measurements and probing small spaces. In this paper, we compare two methods of fabrication of small, cylindrical carbon-fiber microelectrodes: flame-etching and electrochemical etching. With both methods, microelectrodes can be fabricated with tip diameters of 1 to 3 microm. Electrodes were tested with fast-scan cyclic voltammetry. Flame etching resulted in electrodes that have larger S/N ratios and higher currents per unit area for 1 microM dopamine than normal carbon-fiber microelectrodes or electrochemically etched electrodes. Therefore, the increased sensitivity is not just a property of size. The flame-etched surfaces had nanometer-scale surface features that were not observed on the other electrodes and exhibited increased sensitivity for other electroactive compounds found in the brain, including ascorbic acid, DOPAC, and serotonin. Faster kinetics and a faster response to a step change in dopamine were also observed, when the applied waveform was -0.4 to 1.0 V and back at 400 V/s. The sensitivity of the flame-etched electrodes was enhanced by overoxidizing the surface. The flame-etched electrodes were used to detect dopamine release in anesthetized rats after a single stimulation pulse. The small flame-etched electrodes will facilitate measurements of low concentrations in discrete brain regions or small organisms.     

Effect of pH and surface functionalities on the cyclic voltammetric responses of carbon-fiber microelectrodes.

Carbon electrodes are useful for the detection of oxidizable species with cyclic voltammetry. In particular, carbon-fiber microelectrodes have been employed for the measurement of several neurotransmitters in brain tissue. However, during cyclic voltammetry with carbon-fiber electrodes the current varies with changes in concentration of some inorganic cations as a result of their interaction with surface functional groups. The electrode's response to the hydronium ion is a particular concern because its voltammetric response occurs over a broad range of potentials that overlap those of neurotransmitters of interest such as dopamine. This is especially a problem in vivo because simultaneous changes of dopamine and pH frequently occur in brain tissue. In this work, voltammetric current changes are shown to arise from pH dependent shifts in the peak potentials of background voltammetric waves that arise from species confined to the carbon-fiber electrode surface. Polishing the electrode with alumina suspended in cyclohexane in an environment containing lowered oxygen, a method previously demonstrated to remove oxides from the carbon surface, leads to a substantial reduction in the sensitivity to pH changes. However, this is accompanied by a loss in signal amplitude for dopamine. The dopamine response can be restored using the cation exchanger Nafion without significantly increasing the pH response. To investigate which oxide functional groups play a direct role in the electrode's current responses to changes in pH, surface-confined carbonyl and alcohol functionalities were chemically modified. In both cases, the modification did not affect the carbon-fiber electrode's responsiveness to changes in pH. Nonetheless, the polishing technique proved to be effective in reducing pH interferences in in vivo applications.     

Subsecond adsorption and desorption of dopamine at carbon-fiber microelectrodes

High-repetition fast-scan cyclic voltammetry and chronoamperometry were used to quantify and characterize the kinetics of dopamine and dopamine-o-quinone adsorption and desorption at carbon-fiber microelectrodes. A flow injection analysis system was used for the precise introduction and removal of a bolus of electroactive substance on a sub-second time scale to the disk-shaped surface of a microelectrode that was fabricated from a single carbon fiber (Thornel type T650 or P55). Pretreatment of the electrode surfaces consisted of soaking them in purified isopropyl alcohol for a minimum of 10 min, which resulted in S/N increasing by 200-400% for dopamine above that for those that were soaked in reagent grade solvent. Because of adsorption, high scan rates (2,000 V/s) are shown to exhibit equivalent S/N ratios as compared to slower, more traditional scan rates. In addition, the steady-state response to a concentration bolus is shown to occur more rapidly when cyclic voltammetric scans are repeated at short intervals (4 ms). The new methodologies allow for more accurate determinations of the kinetics of neurotransmitter release events (10-500 ms) in biological systems. Brain slice and in vivo experiments using T650 cylinder microelectrodes show that voltammetrically measured uptake kinetics in the caudate are faster using 2,000 V/s and 240 Hz measurements, as compared to 300 V/s and 10 Hz.     

Achievement of near-reversible behavior for the

Voltammetric studies in the absence of added supporting electrolyte are presently dominated by the use of near-steady-state microelectrode techniques and millimolar or lower depolarizer concentrations. However, with this methodology, large departures from conventional migration-diffusion theory have been reported for the [Fe(CN)6](3-/4-) process at both carbon fiber and platinum microdisk electrodes. In contrast, data obtained in the present study reveal that use of the transient cyclic voltammetric technique at glassy carbon, gold, or platinum macrodisk electrodes and K4[Fe(CN)6] or K3[Fe(CN)6] concentrations of 50 mM or greater provides an approximately reversible response in the absence of added electrolyte. It is suggested that the use of very high [Fe(CN)6](3-) and [Fe(CN)6](4-) concentrations overcomes problems associated with a diffuse double layer and that large electrode surface areas and faster potential sweep rates minimize electrode blockage and passivating phenomena that can plague voltammetric studies at microelectrodes. The cyclic voltammetry of the [Fe(CN)6](3-/4-) couple at a range of concentrations at macroelectrodes in the absence of added inert electrolyte is compared with that obtained in the presence of 1 M KCl. The enhanced influences of uncompensated resistance, migration, and natural convection arising from density gradients under transient conditions at macrodisk electrodes also are considered.     

Nanostructured carbon fiber disk electrodes for sensitive determinations of adenosine and uric acid

Nanostructured carbon fiber microdisk electrodes were prepared by a combination of mechanical polishing and electrolytic treatment, where the latter involved moderate oxidation of the surface, followed by a reduction. A high density of surface defects contributed to a high capacitance of the nanostructured electrodes. Facilitated proton transfer was observed at the nanostructured surface and was associated with cation-exchanged oxide defects. The nanostructured surfaces intercalated uric acid and adenosine and engaged in fast electron/proton transfer in the oxidation of both analytes. As a result, electrolytic treatment followed by fast-scan voltammetry determinations led to a sensitive response to both analytes in physiological buffers. The nanostructured electrodes showed remarkable stability and could be easily regenerated and reused. With long use, electrode activity decreased. Kinetic discrimination of the surface-mediated reaction of ascorbate was achieved at high scan rates.     

Electrochemical biosensing platforms using platinum nanoparticles and carbon nanotubes

Platinum nanoparticles with a diameter of 2-3 nm were prepared and used in combination with single-wall carbon nanotubes (SWCNTs) for fabricating electrochemical sensors with remarkably improved sensitivity toward hydrogen peroxide. Nafion, a perfluorosulfonated polymer, was used to solubilize SWCNTs and also displayed strong interactions with Pt nanoparticles to form a network that connected Pt nanoparticles to the electrode surface. TEM and AFM micrographs illustrated the deposition of Pt nanoparticles on carbon nanotubes whereas cyclic voltammetry confirmed an electrical contact through SWCNTs between Pt nanoparticles and the glassy carbon (GC) or carbon fiber backing. With glucose oxidase (GOx) as an enzyme model, we constructed a GC or carbon fiber microelectrode-based biosensor that responds even more sensitively to glucose than the GC/GOx electrode modified by Pt nanoparticles or CNTs alone. The response time and detection limit (S/N = 3) of this biosensor was determined to be 3 s and 0.5 microM, respectively.     

Oxidation-state speciation of

The analytical utility of chemically modified microelectrodes for oxidation-state speciation of redox couples by cyclic voltammetry has been explored. [Re(I)(DMPE)3]+/[Re(II)(DMPE)3]2+, where DMPE = 1,2-bis(dimethylphosphino)ethane, was studied at carbon-fiber microelectrodes of approximately 5 microm in radius coated with Nafion-entrapped solgel-derived silica (Nafion-silica) composite. The results are compared with cyclic voltammetry of [Fe(CN)6]3-/[Fe(CN)6]4- at bare carbon-fiber microelectrodes. At both microelectrodes, the cathodic and anodic limiting currents are linearly proportional to the concentrations of the reducible and oxidizable species of a redox couple, respectively. The shape of the cyclic voltammogram and the magnitude of the steady-state limiting current are not affected by the potential at which the scan starts. Speciation of both forms of a redox couple could be achieved voltammetrically at the microelectrodes. However, a considerably slower scan rate was required to achieve steady state at the modified electrode because of the smaller diffusion coefficients of [Re(I)(DMPE)3]+ and [Re(II)(DMPE)3]2+ in the Nafion-silica composite. The detection limit at the modified electrode was considerably lower (5 x 10(-9) M for [Re(I)(DMPE)3]+) than at the bare electrode (6 x 10(-5) M for [Fe(CN)6]3- and [Fe(CN)6]4-) because of the substantial preconcentration of [Re(I)(DMPE)3]+ by the Nafion-silica composite.     

Fabrication and characterization of sputtered-carbon microelectrode arrays

This paper describes a robust and reliable process for fabricating a novel sputter-deposited, thin-film carbon microelectrode array using standard integrated circuit technologies and silicon micromachining. Sputter-deposited carbon films were investigated as potential candidates for microelectrode materials. The surface properties and cross section of the microelectrode arrays were studied by atomic force microscopy and scanning electron microscopy, respectively. Electrical site impedance, crosstalk, and lifetime (dielectric integrity) of microelectrodes in the array were characterized. Electrochemical response of the microelectrodes to hexaammineruthenium(III) chloride and dopamine were investigated by fast-scan cyclic voltammetry and high-speed, computer-based chronoamperometry; results show that thin-film carbon microelectrodes are well-behaved electrochemically. The thin carbon films offer extremely good electrical, mechanical, and chemical properties and thus qualify as viable candidates for various electroanalytical applications, particularly acute neurophysiological studies.     

Carbon composite microelectrodes fabricated by electrophoretic deposition

Single-walled carbon nanotubes were deposited on one end of the etched carbon fiber electrodes by an electrophoretic method. The carbon nanotube bundles formed a dense network on the carbon fiber surface. The electrochemical properties of the composite carbon electrodes were studied in the buffered neutral solutions. The results in cyclic voltammetry's characteristic indicate that the electrons on the electrodes transfer very fast. Furthermore, the redox reactions of dopamine (DA) on the composite electrodes show good sensitivity. When the DA concentration was 0.02 mM, the peak current in differential pulse technique reached 1.33 microA after performing the background subtraction. In addition, the simultaneous detection of DA and ascorbic acid (AA) showed that the interference effect was not observed. It was suggested that the carbon composite microelectrodes have potential applications as electrochemical sensors inside a single cell.     

Coupled electrochemical reactions at bipolar microelectrodes and nanoelectrodes

Here we report the voltammetric study of coupled electrochemical reactions on microelectrodes and nanoelectrodes in a closed bipolar cell. We use steady-state cyclic voltammetry to discuss the overall voltammetric response of closed bipolar electrodes (BPEs) and understand its dependence on the concentration of redox species and electrode size. Much of the previous work in bipolar electroanalytical chemistry has focused on the use of an "open" cell with the BPE located in an open microchannel. A closed BPE, on the other hand, has two poles placed in separate compartments and has remained relatively unexplored in this field. In this work, we demonstrated that carbon-fiber microelectrodes when backfilled with an electrolyte to establish conductivity are closed BPEs. The coupling between the oxidation reaction, e.g., dopamine oxidation, on the carbon disk/cylinder and the reduction of oxygen on the interior fiber is likely to be responsible for the conductivity. We also demonstrated the ability to quantitatively measure voltammetric properties of both the cathodic and anodic poles in a closed bipolar cell from a single cyclic voltammetry (CV) scan. It was found that "secondary" reactions such as oxygen reduction play an important role in this process. We also described the fabrication and use of Pt bipolar nanoelectrodes which may serve as a useful platform for future advances in nanoscale bipolar electrochemistry.     

Fast-scan deposition-stripping voltammetry at carbon-fiber microelectrodes: real-time, subsecond, mercury free measurements of copper

Elevated concentrations of hazardous metals in aquatic systems are known to threaten human health. Mobility, bioavailability, and toxicity of metals are controlled by chemical speciation, a dynamic process. Understanding metal behavior is limited by the lack of analytical methods that can provide rapid, sensitive, in situ measurements. While electrochemistry shows promise, it is limited by its temporal resolution and the necessity for Hg modified electrodes. In this letter, we apply fast-scan deposition-stripping voltammetry at carbon-fiber microelectrodes for in situ measurements of Cu(II). We present a novel, Hg-free technique that can measure Cu(II) with ppb sensitivity at 100 ms temporal resolution.     

Measurement of nanomolar dopamine diffusion using low-noise perfluorinated ionomer coated carbon fiber microelectrodes and high-speed cyclic voltammetry

Several improvements in the fabrication and use of carbon fiber voltammetric microelectrodes (CFVMs) are described. These procedures did not involve oxidative treatment, but resulted in sensitivities and selectivities approaching those of treated CFVMs, without the inherent slow response times associated with the latter electrodes. To accomplish this we reduced CFVM noise by (1) improving the adhesive seal between the 8 microns o.d. carbon fiber and the glass insulation using vacuum, (2) snapping rather than cutting or beveling the fiber to be flush with the glass, and (3) using a concentrated electrolyte solution to make electrical contact with the fiber. System noise was reduced by digital smoothing and signal averaging. Selectivity of the CFVMs for dopamine over ascorbate was enhanced to better than 2000:1 by coating with Naflon, a perfluorinated cation exchange polymer, using a low (+0.5 V vs Ag/AgCl) electroplating potential. This low voltage also prevented electrode surface oxidation. To demonstrate the performance of our CFVMs, we used them in conjunction with high-speed cyclic voltammetry to accurately measure the diffusion coefficient of iontophoretically released dopamine at concentrations as low as 35 nM over distances of less than 200 microns in agarose gel.     

Simulation of two-electron homogeneous electrocatalysis for steady-state voltammetry at hemispherical microelectrodes

Expanded space grid digital simulation of second-order, two-electron homogeneous electrocatalysis was extended to slow scan voltammetry at hemispherical microelectrodes. Predictions of the simulations are examined for reversible and quasireversible heterogeneous charge transfer of catalyst for a range of homogeneous catalytic rate constants (k1) and electrode radii. Working curves of catalytic efficiency vs long k1 were generated assuming reacting species with equal diffusion coefficients. As electrode radii in the less than 10-microns range decrease, progressively larger homogeneous catalytic rates are needed to yield analytically significant amplification of limiting currents. Simulations using hemispherical radii of (2/pi)rd can be used to predict catalytic efficiencies for microdisk electrodes with radii rd. Simulated working curves were used to estimate a log k1 of 3.88 +/- 0.55 (M-1 s-1) for electron transfer from the anion radical of 9,10-diphenylanthracene to 4,4'-dibromobiphenyl from steady-state catalytic efficiencies obtained at carbon microdisk electrodes. This value was in good agreement with 3.90 +/- 0.16 M-1 s-1 found previously by cyclic voltammetry.     

Metallic nanoparticle-carbon nanotube composites for electrochemical determination of explosive nitroaromatic compounds

Metal nanoparticles (Pt, Au, or Cu) together with multiwalled and single-walled carbon nanotubes (MWCNT and SWCNT) solubilized in Nafion have been used to form nanocomposites for electrochemical detection of trinitrotoluene (TNT) and several other nitroaromatics. Electrochemical and surface characterization by cyclic voltammetry, AFM, TEM, SEM, and Raman spectroscopy confirmed the presence of metal nanoparticles on CNTs. Among various combinations tested, the most synergistic signal effect was observed for the nanocomposite modified glassy carbon electrode (GC) containing Cu nanoparticles and SWCNT solubilized in Nafion. This combination provided the best sensitivity for detecting TNT and other nitroaromatic compounds. Adsorptive stripping voltammetry for TNT resulted in a detection limit of 1 ppb, with linearity up to 3 orders of magnitude. Selectivity toward the number and position of the nitro groups in different nitroaromatics was very reproducible and distinct. Reproducibility of the TNT signal was within 7% (n = 8) from one electrode preparation to another, and the response signal was stable (+/-3.8% at 95% confidence interval) for 40 repeated analyses with 10 min of preconditioning. The Cu-SWCNT-modified GC electrode was demonstrated for analysis of TNT in tap water, river water, and contaminated soil.     

Facile electrochemical characterization of core/shell nanoparticles. Ag core/Ag(2)O shell structures

We report in this paper a facile approach for the formation and electrochemical characterization of silver-silver oxide core-shell nanoparticles (NPs). Thus, thermal treatment at temperatures between 200 and 360 degrees C of Ag NP, in the gas phase or in an organic solvent, has been used to achieve the formation Ag@Ag2O NP. The evidence of formation of such a core-shell structure was obtained by cyclic voltammetry using a Nafion modified electrode (where Nafion containing carbon particles is used as the matrix to encapsulate the core-shell NP). Initial positive scans measure free Ag. Initial negative scans measure Ag2O, with the following positive scan, compared to the initial one, providing a measure of "trapped" or core Ag. The results presented demonstrate the utility of this approach in characterizing core-shell structures, like Ag@Ag2O, which could be extended to other core-shell forms, such as bimetallic core-shell NP.     

Electrochemical determination of dopamine based on self-assembled peptide nanostructure

Self-assembled peptide nanostructures are electronically insulating as are most biomaterials derived from natural amino acids. To obtain additional properties and increase the applicability of peptide nanomaterials, some chemical modifications can be performed and materials can be functionalized to form hybrid compounds. In this work, we described the formation of L-diphenylalanine nanotubes (PNTs) with cyclic-tetrameric copper(II) species containing the ligand (4-imidazolyl)ethylene-2-amino-1-ethylpyridine [Cu(4)(apyhist)(4)](4+) in the Nafion membrane on a vitreous carbon electrode surface. This copper complex has been studied as structural and functional models for the active centers of copper containing redox enzymes. Scanning electron microscopy was used to confirm the formation of the nanostructures. The electrochemical properties of the PNT-[Cu(4)(apyhist)(4)](4+)/Nafion film on a glassy carbon electrode were characterized using cyclic voltammetry and square-wave voltammetry and showed high electrocatalytic activity toward the oxidation of dopamine (DA). The detection sensitivity was found to be enhanced by the use of copper(II) complex in the PNTs/Nafion films. Under the optimum conditions, the square-wave voltammetry peak height was linearly related to the DA concentration over two concentration intervals, viz., 5.0-40 μmol L(-1) and 40-1000 μmol L(-1). The detection limit was 2.80 μmol L(-1) (S/N = 3), and ascorbic acid did not interfere with the DA detection. These results suggested that this hybrid bioinorganic system provides an attractive advantage for a new type of electrochemical sensors. The detection sensitivity was found to be enhanced by use of PNTs.     

Temporal resolution in electrochemical imaging on single PC12 cells using amperometry and voltammetry at microelectrode arrays

Carbon-fiber-microelectrode arrays (MEAs) have been utilized to electrochemically image neurochemical secretion from individual pheochromocytoma (PC12) cells. Dopamine release events were electrochemically monitored from seven different locations on single PC12 cells using alternately constant-potential amperometry and fast-scan cyclic voltammetry (FSCV). Cyclic voltammetry, when compared to amperometry, can provide excellent chemical resolution; however, spatial and temporal resolution are both compromised. The spatial and temporal resolution of these two methods have been quantitatively compared and the differences explained using models of molecular diffusion at the nanogap between the electrode and the cell. A numerical simulation of the molecular flux reveals that the diffusion of dopamine molecules and electrochemical reactions both play important roles in the temporal resolution of electrochemical imaging. The simulation also reveals that the diffusion and electrode potential cause the differences in signal crosstalk between electrodes when comparing amperometry and FSCV.