Direct and rapid electrochemical immunosensing system based on a conducting polymer

A system device using multifunctional conjugated copolymer poly(5-hydroxy-1,4-naphthoquinone-co-hydroxy-2-thioacetic acid-1,4-naphthoquinone) acting both as immobilizing and transducing element for reagentless immunosensor has been constructed. Its functionality was evaluated in an antigen-antibody interaction model using ovalbumin-anti-ovalbumin. It was shown that the system specifically detects via electrochemical signal the antigen-antibody immune interaction in a reagentless context. Comparison to the conventional ELISA technique relevant to performance and sensitivity was presented.     

Nanosilver and DNA-functionalized immunosensing probes for electrochemical immunoassay of alpha-fetoprotein

A simple and sensitive electrochemical immunosensor for a one-step immunoassay for alpha-fetoprotein (AFP) was designed using silver nanoparticles and double-stranded DNA as matrices. The detection was based on the change in the electron transfer resistance before and after the antigen-antibody reaction by using electrochemical impedance spectroscopy. Under optimal conditions, the resistance shift of the immunosensor is proportional to the AFP concentration in the range 3.5 –360 ng·mL^?1 with a detection limit of 1.5 ng·mL^?1 (at 3σ). The immunosensor exhibits high sensitivity, good reproducibility and stability. Results obtained for clinical serum samples by the immunosensor are in accordance with those determined by spectrophotometric enzyme-linked immunosorbent assays.     

Determination of trace cobalt(II) by adsorptive stripping voltammetry on disposable microfabricated electrochemical cells with integrated planar metal-film electrodes

This work reports the determination of trace Co(II) by adsorptive stripping voltammetry on disposable three-electrode cells with on-chip metal-film electrodes. The heart of the sensors was a bismuth-film electrode (BiFE) with Ag and Pt planar strips serving as the reference and counter electrodes, respectively. Metals were deposited on a silicon chip by sputtering while the areas of the electrodes were patterned via a metal mask. Co(II) was determined by square wave adsorptive stripping voltammetry (SWAdSV) after complexation with dimethylglyoxime (DMG). The experimental variables (the DMG concentration, the preconcentration potential, the accumulation time and the SW parameters), as well as potential interferences, were investigated. Using the selected conditions, the 3σ limit of detection was 0.09 μg l⁻¹ of Co(II) (for 90 s of preconcentration) and the relative standard deviation for Co(II) was 3.8% at the 2 μg l⁻¹ level (n = 8). The method was applied to the determination of Co(II) in a certified river water sample. These mercury-free electrochemical devices present increased scope for field analysis and μ-TAS applications.     

Development and evaluation of electrochemical glucose enzyme biosensors based on carbon film electrodes

Electrochemical glucose enzyme biosensors have been prepared on carbon film electrodes made from carbon film electrical resistors. Evaluation and characterisation of these electrodes in phosphate buffer saline solution has been carried out with and without pretreatment by cycling in perchloric acid or at fixed applied potential. Both pretreatments led to a reduction in the carbon surface oxidation peak and enabled better detection of hydrogen peroxide in the pH range of 5-7. Glucose oxidase enzyme was immobilised on the carbon surface by mixing with glutaraldehyde, bovine serum albumin and with and without Nafion. The performance of these two types of electrode was similar, that containing Nafion being more physically robust. Linear ranges were up to around 1.5 mM, with detection limits 60 μM, and pretreatment of the carbon film electrode at a fixed potential of +0.9 V versus SCE for 5 min was found to be the most beneficial. Michaelis-Menten constants between 5 mM and 10 mM were found under the different experimental conditions. Coating the immobilised enzyme layer with a thin layer of Nafion was found to give similar results in the determination of glucose to mixing it but with benefits against interferences for the analysis of complex matrices, such as wine. Potentialities, for a short-term-use or disposable sensors, are indicated.     

Composite electrodes of electrochemical capacitors based on carbon materials with different structure

For composite electrodes based on active carbon DCL Supra 30, ordered mesoporous carbon, and synthetic carbon material Sibunit, the electrical double layer capacitance is studied. The original carbon samples are characterized by the methods of gas adsorption, X-ray diffraction, and transmission electron microscopy. The mesoporous structure of the material synthesized by the template method provides the maximum rate of ion transport in pores and demonstrates an insignificant decrease in the specific capacitance (9.5% in an aqueous electrolyte and 1.1% in an nonaqueous electrolyte) with an increase in the polarizing current.     

Investigations on electrochemical supercapacitors using polypyrrole redox electrodes and PMMA based gel electrolytes

Polypyrrole based solid state electrochemical redox supercapacitors have been fabricated using the polymeric gel electrolytes comprising of poly methyl methacrylate (PMMA)-propylene carbonate (PC)-ethylene carbonate (EC)-perchlorate salts of different cations [Li⁺, Na⁺ and (C₂H₅)₄N⁺ (TEA⁺)]. A comparative study has been carried out using linear sweep reversal voltammetry, complex impedance spectroscopy and constant current charge-discharge tests. The capacitance values of the cells have been observed to be in the range of 15.3-22.5 mF cm⁻² (equivalent to single electrode specific capacitance of 120-178 F g⁻¹ of polypyrrole). This corresponds to the values of energy density 16.7-24.7 Wh kg⁻¹ and power density 1.6-2.8 kW kg⁻¹ calculated for the working voltage of 1.0 V limited for polypyrrole based redox capacitors. Substantial improvements in the coulombic efficiency of the cells have been observed (close to 100%) due to the application of gel electrolytes showing flexible and liquid like behaviour. Further, the types and sizes of the cations in the gel electrolytes do not play any dominant role in the capacitive behaviour of the redox cells.     

Microscopic in situ diffuse reflectance spectroelectrochemistry of solid state electrochemical reactions of particles immobilized on electrodes

An instrumental setup is described which allows electrochemical measurements to be performed with solid particles immobilized on the surface of a graphite electrode with in situ recording of diffuse reflectance spectra under an incident light microscope. The instrument used includes a special electrochemical cell and a microscope which is interfaced by a light␣guide to a transputer-integrated photodiode-array spectrometer allowing measurements ranging from 320 to 950 nm with a resolution of 3.2 nm/pixel and a PC-interfaced potentiostat. The ₀R₀ geometry of the optical arrangement and the use of crossed polarization filters for blocking specular reflectance makes it possible to use the Kubelka-Munk function for quantifying the optical measurements. The above instrument has been used for the study of the electrochromic system of solid silver octacyanomolybdate(IV/V) adjacent to a silver nitrate solution. The in situ diffuse reflec tance spectroelectrochemical measurements prove that the electrochemical reaction starts at the graphite/sample interface and then advances into the bulk of the sample towards the sample/electrolyte interface. The ratios Red:Ox determined by spectrometry and chronocoulometry as a function of electrode potentials are identical.     

Monolithic integration of optical waveguides for absorbance detection in microfabricated electrophoresis devices.

The fabrication and performance of an electrophoretic separation chip with integrated optical waveguides for absorption detection is presented. The device was fabricated on a silicon substrate by standard microfabrication techniques with the use of two photolithographic mask steps. The waveguides on the device were connected to optical fibers, which enabled alignment free operation due to the absence of free-space optics. A 750 microm long U-shaped detection cell was used to facilitate longitudinal absorption detection. To minimize geometrically induced band broadening at the turn in the U-cell, tapering of the separation channel from a width of 120 down to 30 microm was employed. Electrical insulation was achieved by a 13 microm thermally grown silicon dioxide between the silicon substrate and the channels. The breakdown voltage during operation of the chip was measured to 10.6 kV. A separation of 3.2 microM rhodamine 110, 8 microM 2,7-dichlorofluorescein, 10 microM fluorescein and 18 microM 5-carboxyfluorescein was demonstrated on the device using the detection cell for absorption measurements at 488 nm.     

Design and development of microfabricated capillary electrophoresis devices with electrochemical detection

Our efforts have been focused on developing a self-contained and transportable microfabricated electrophoresis (CE) system with integrated electrochemical detection (ED). The current prototype includes all necessary electrodes “on-chip” and utilizes miniaturized CE and ED supporting electronics custom designed for this purpose. State-of-the-art design/modeling tools and novel microfabrication procedures were used to create recessed platinum electrodes with complex geometries and the CE/ED device from two patterned ultra-flat glass substrates. The electrodes in the bottom substrate were formed by a self-aligned etch and deposition technique followed by a photolithographic lift-off process. The microchannels (20 μm deep×65 μm wide (average)) were chemically etched into the top substrate followed by thermal bonding to complete the microchip device. CE/ED experiments were performed using 0.02 M phosphate buffer (pH 6), an analyte/buffer solution (2.2 mM dopamine, 2.3 mM catechol) and varying separation voltages (0-500 V) with a custom electronics unit interfaced to a laptop computer for data acquisition. Detection limits (S/N=3) were found to be at the micromolar level and a linear detection response was observed up to millimolar concentrations for dopamine and catechol. The microchip CE/ED system injected 50 pl volumes of sample, which corresponded to mass detection limits on the order of 200 amol. For the first time, an integrated “on-chip” multi-electrode array CE/ED device was successfully designed, fabricated and tested. The majority of the electrodes (six out of eight) in the array were capable of detecting dopamine with the amplitude of the signal (i.e., peak heights) decreasing as the electrode distance from the channel exit increased.     

An integrated electrochemical fructose biosensor based on tetrathiafulvalene-modified self-assembled monolayers on gold electrodes

An integrated amperometric fructose biosensor based on a gold electrode (AuE) modified with a self-assembled monolayer (SAM) of 3-mercaptopropionic acid (MPA) on which fructose dehydrogenase (FDH) and the mediator tetrathiafulvalene (TTF) are co-immobilized by cross-linking with glutaraldehyde is reported. Variables concerning the behavior of the biosensor were optimized by taking the slope value obtained for the fructose calibration plot in the 0.1–1.0 mM concentration range as a criterion of selection. At an applied potential of +0.20 V, a good repeatability of such slope values (RSD=6.7%, n=10) was obtained with no need to apply a cleaning or pretreatment procedure to the modified electrode. Moreover, results from five different TTF-FDH-MPA-AuEs yielded a RSD of 5.8%. The useful lifetime of one single biosensor was approximately 30 days, exhibiting a 93% of the original response on the 33rd day. A linear calibration graph was obtained for fructose over the 1.0×10^–5–1.0×10^–3 M range, with a limit of detection of 2.4×10^–6 M. The effect of potential interferents was evaluated. The TTF-FDH-MPA-AuE also performed well in the flow-injection mode. The biosensor was used for the determination of fructose in real samples, and the results compared with those provided by using a commercial enzyme test kit.     

Transparent electrochemical capacitor based on electrodeposited MnO2 thin film electrodes and gel-type electrolyte

A new concept for a transparent electrochemical capacitor was demonstrated. It includes two MnO₂ electrodes separated with a gel electrolyte, making the whole device less than 1 mm thick and transparent. Preliminary electrochemical experiments revealed a capacitance of 2 mF cm^?2 over more than 200 cycles. The device exhibited only a small absorbance change during repeated cycling.     

Nanoparticle based enhancement of electrochemical DNA hybridization signal using nanoporous electrodes

A novel nanoparticle-based enhanced methodology for the detection of ssDNA using nanoporous alumina filter membranes, containing pores of 200 nm in diameter, is reported. The blockage of the pores due to the hybridization is detected by measuring the decrease in the differential pulse voltammetric response of the [Fe(CN)(6)](4-/3-) redox indicator and using screen-printed carbon electrodes as transducing platform. Furthermore, 20 nm gold nanoparticle (AuNPs) tags are used in order to increase the sensitivity of the assay. The enhancement mechanism of DNA detection is due to an additional blocking effect induced by hybridization reaction by bringing AuNPs inside the pores. The developed methodology can be extended to other biosensing systems with interest not only for DNA but also for proteins and cells. The developed nanochannel/nanoparticle biosensing system would have enormous potential in future miniaturized designs adapted to mass production technologies such as screen-printing technology.     

Sensitive and selective determination of tolterodine tartrate and its electrochemical investigation on solid carbon based electrodes

Tolterodine tartrate, a muscarinic receptor antagonist, was oxidized in various buffer media with different pH values using cyclic, differential pulse, and square wave voltammetric techniques on glassy carbon and boron-doped diamond electrodes. Two irreversible anodic peaks were obtained. The oxidation process of tolterodine tartrate was diffusion controlled depending on pH for both electrodes. A detailed oxidation mechanism was proposed and discussed. The dependences of the peak current and peak potentials on pH, concentration, nature of the buffer, and scan rate were investigated. A linear response between the peak current and the tolterodine tartrate concentration was obtained using differential pulse and square wave voltammetric techniques in the range of 0.4–8.0 μM for the peak at lower potential in acetate buffer at pH 5.7 and 0.4–40.0 μM for the peak at higher potential in 0.1 M H₂SO₄ on glassy carbon electrode and in the range of 0.4–40.0 μM in Britton-Robinson buffer at pH 11.0 on boron-doped diamond electrode. Limit of detection values varied between 0.04 and 0.13 μM for both techniques and electrodes. The repeatability, reproducibility, precision, and accuracy of the proposed methods were investigated. The recovery studies were also achieved to check selectivity, precision, and accuracy of the methods. The proposed methods were successfully applied to determine tolterodine tartrate from pharmaceutical dosage forms without any interference from inactive excipients.     

Catalysis of electrochemical reactions at redox polymer electrodes: Kinetic model for stationary voltammetric techniques

A kinetic model describing the catalytic activity of redox polymer film electrodes is presented and discussed for reaction schemes in which the primary reaction between the active form of the catalyst and the substrate is the rate-determining step. The derivations are given for stationary techniques, such as rotating disc electrode voltammetry. Besides the diffusion of the substrate from the bulk of the solution to the film—solution interface, three kinetic factors determine the magnitude of the catalytic current: “diffusion” of electrons from the electrode surface to the film—solution interface; diffusion of the substrate in the opposite direction; rate of the rate-determining step of the catalytic reaction. Correspondingly, the kinetics of the diffusion—reaction process can be entirely described by only three dimensionless parameters expressing the relative rates of the rate-controlling processes. Provision is made for the partition coefficient of the substrate between the film and the solution to be different from unity. A finite-difference resolution of the kinetics is developed, but attention is mainly focused on the search of characteristic behaviors depending upon a lesser number of parameters and expressed by simple closed-form equations which are particularly convenient to use in the processing of experimental data. These are essentially of four types.

• 1.(1) The electron and substrate diffusion of the film are so fast that the rate-controlling phenomenon in the film is the catalytic reaction.
• 2.(2) The cataytic reaction is so fast that the kinetics is controlled jointly by the two diffusion process.
• 3.(3) When “diffusion” of electron is faster than diffusion of substrate in the film a pure kinetic situation may arise by mutual compensation of the latter process and the catalytic reaction.
• 4.(4) In the opposite case a pure kinetic situation may again arise resulting this time from mutual compensation of electron “diffusion” and catalytic reaction.

The kinetics observed in intermediary situations is also described, as well as the variations of the parameters that are required to pass from one limiting behavior to the other. It is shown that, according to the kinetic situation prevailing in the film, a second wave way appear which features the direct reduction of the substrate at the electrode surface after it has diffused through the film. Diagnostic criteria of the various limiting situation are presented, being based on the variations of the height of the catalytic wave with the rotation speed and the solution concentration of the substrate, as well as on the existence and height of the second wave. The electron “diffusion” through the film can be characterized independently by means of experiments involving the oxido-reduction of the film in the absence of substrate. Conversely, the substrate diffusion in the film can be characterized using analogous system in wich catalysis is absent. With the help of these data, the catalytic reaction occuring in the film can be quantitatively characterized. It is shown that this does not require exact knowledge of the film thickness. Such application of the kinetic model to actual experimental systems are illustrated by the discussion of experimental data from the literature.     

Electrolyte effects on electrochemical properties of osmium complex polymer modified electrodes.

The electrolyte effects on the electrochemical behaviors of osmium complex polymer modified electrodes were investigated by a comparison between two osmium complexes, [Os(bpy)2(PVI)10Cl]Cl (Os-PVI10) and [Os(bpy)2(PVP)10Cl]Cl (Os-PVP10). The electrode process at Os-PVI10 modified electrodes is reaction-controlled, while a diffusion-controlled electrode process exists at Os-PVP10 modified electrodes. Both the cation and anion in supporting electrolytes strongly affect their electrochemical behaviors, such as the redox potential, wave shape and peak current. These phenomena are attributed to a change in the film structure and polymer swelling in various supporting electrolytes. The influence of electrolyte anions on the electrochemical behaviors is related to their hydrophobicity. The electrode process of Os-PVP10 depends on the pH value of solutions, exhibiting different electron transfer mechanisms.