Abstract: Metal nanoparticles and carbon nanotubes have been shown to possess high electrocatalytic activity. Indium tin oxide (ITO) is a popular electrode material, but the electro-catalytic properties of its nano-materials have not been reported. We demonstrate here for the first time facile electrocatalytic oxidation of ascorbic acid on ITO nanoparticle-modified electrodes. Compared to the conventional ITO thin film electrode, the voltammetric peak potential for ascorbic acid oxidation was lowered by 800 mV on ITO nanoparticle-modified electrodes to a potential similar to metal electrodes. The ITO nanoparticle was composed of 90% In2O3 and 10% SnO2. Since the electrocatalytic activity was also found on In2O3 nanoparticle electrodes but not on SnO2 nanoparticle electrodes, the In2O3 composition in ITO nanoparticle is mainly responsible for the high activity. In photoluminescence measurement, two intense emission peaks at 415 nm and 438 nm associated with surface oxygen vacancies were observed on the semiconductor electrodes. It was hypothesized that the oxygen vacancies could be the active sites for electrocatalytic reactions. A linear relationship between the oxidation current and ascorbic acid concentration was found in the range of 10μM - 5mM, with a lower detection limit of 5μM and 7.9% RSD (n=11). The high electro-catalytic activity and transmittance of In2O3 and ITO nanoparticle electrodes make them potentially very useful in opto-electronic devices and chemical/bio-sensors.
Abstract: Electrochemiluminescence (ECL) systems using (bpy = 2,2â²-bipyridyl) and tripropylamine (TPA) on metal and carbon electrodes are widely used in immunoassay and DNA hybridization detections. The large overpotential of TPA oxidation on indium tin oxide (ITO) electrodes has limited their ECL application. In this work, significant enhancement of ECL signal from the /TPA system was achieved on ITO nanoparticle-modified electrode. Due to the electrocatalytic activity of ITO nanoparticle electrodes, TPA oxidation potential was found to shift negatively by 600 mV, leading to 24-fold increases in ECL peak intensity over the conventional ITO electrode, and 2-fold increases in the integrated signal over the conventional gold electrode. As a result, the detection limit for in solution was 20-fold lower than the value on the conventional ITO electrode. Because the transmittance of ITO at 610 nm (the wavelength of emission maximum) was reduced only slightly after nanoparticle modification, it may offer a better alternative than metal and carbon electrodes in ECL measurements.
Abstract: A novel label-free electrochemical method for measuring the activity of protein tyrosine kinases (PTK) has been developed. Epidermal growth factor receptor (EGFR), a typical PTK associated with a large percentage of all solid tumors, was used as the model kinase. Poly(glu, tyr) (4:1) peptide, as a substrate of EGFR, was covalently immobilized on the surface of indium tin oxide (ITO) electrode by silane chemistry. The tyrosine (Tyr) residue in the polypeptide served as an electrochemical signal reporter. Its voltammetric current was catalyzed by a dissolved electron mediator Os(bpy)32+ (bpy = 2,2â²-bipyridine) for increased sensitivity. Phosphorylation of the Tyr led to a loss of its electrochemical current, thus providing a sensing mechanism for PTK activity. Experimental conditions for the silanization of ITO surface and immobilization of polypeptide were investigated in details to facilitate the generation of Tyr electrochemical signal. The proposed biosensor exhibited high sensitivity and excellent stability. The limit of detection for EGFR was 1 U mLâ1. Furthermore, this biosensor can also be used for quantitative analysis of kinase inhibition. On the basis of the inhibitor concentration dependent electrochemical signal, the half-maximal inhibition value IC50 of three EGFR inhibitors, PD-153035, OSI-774 and ZD-1839, and their corresponding inhibition constants Ki were estimated, which were in agreement with those obtained from the conventional kinase assay. This electrochemical biosensor can be implemented in an array format for the high throughput assay of in vitro PTK activity and PTK inhibitors screening for practical diagnostic application and drug discovery.
Abstract: Strategies for electrochemical sensing of DNA can be classified into label-free and label-based approaches, categories of which include enzyme-, nanomaterial- and redox labels that are attached to DNA either by covalent or non-covalent means. Metallointercalators represent one group of small molecule redox labels that non-covalently enter the groove of a DNA. The metallointercalator plays a dual-role in acting as a structure indicator (for hybridization) and a signal generator. Labeling is not needed, and electrochemical measurements can be carried out in a label-free solution of an electrolyte. However, such metallointercalators lack the option of catalytic signal generation as in the case of enzyme- and nanomaterial-based labels. Therefore, signal amplification becomes crucial. We first survey here recent progress in this area. A signal-amplifying system is presented that relies on the electroatalytic oxidation of a metallointercalator ruthenium(II)bipyridine/phenoxazine complex in the presence of electron donor species such as oxalate, DNA bases, or tripropylamine. Recent work on such DNA sensors is discussed. Results suggest that such metallointercalator-based DNA sensors represent a viable platform for developing high-throughput and automated PCR/lab-on-a-chip devices as well as visualized multifunctional DNA sensors.
Abstract: Development of the nanodevice that myosin-coated beads âwalkâ on actin filaments (F-actin) tracks for in vitro nanotransportation was hindered due to the difficulty of assembling large-area well-orientated F-actin tracks on the surface. In this work, we present a selective attachment of F-actin with controlled length on a patterned surface by employing biotinylated capped protein gelsolin as intermediate anchoring bridge. A patterned streptavidin layer was formed via coupling with a biotin layer that photo-actively attached to an amine-functionalized glass surface. The patterned film was found stable and homogenous compared to that obtained by microcontact printing method, according to the profiling with fluorescence microscopy. By a secondary blocking process, non-specific binding of F-actin to the patterned surface through electrostatic adsorption can be resisted. The length variation of F-actin as a function of gelsolin concentration was also investigated, implying that F-actin is appropriately of 2.5 μm in average length once F-actin/gelsolin molar ratio is 4:1. Finally, the selective attachment of F-actin was well characterized with quantifying the number of attached F-actin per unit area in the patterned areas over that in blocked areas. The density of F-actin was estimated at c.a. 2 μm2 per actin filament molecule so that the distance between one another actin filament is estimated as c.a. 1.41â1.97 μm. The unique properties of F-actin, e.g. well flexibility or electrical conductivity, make it feasible to lay them down and form unidirectional aligned tracks by fluidic flow or electrical field. This may open a possibility for the long-distant movement of myosin-coated beads, offering a novel discipline for the development of micro-biochip in vitro
Abstract: Electrochemical investigation on the interaction between small molecules and nucleic acids is reviewed, including the interaction of metal ions, metal complexes, drug molecules and organic pollutants with DNA, with the emphasis on the interaction of organic small molecules with DNA. In addition, the prospects of this filed in the future study are discussed
Abstract: A redox-labeled direct competitive electrochemical immunoassay for polycyclic aromatic hydrocarbons (PAHs) was developed. A ruthenium tris(bipyridine)-pyrenebutyric acid conjugate was synthesized as the redox-labeled tracer. Its recognition by an anti-PAH monoclonal antibody was confirmed by surface plasmon resonance. In the immunoassay, the antibody was immobilized on (3-glycidoxypropyl)-trimethoxysilane (GPTMS)-modified indium tin oxide (ITO) electrodes. The assay was quantified by measuring the electro-catalytic current of the redox label in an oxalate-containing electrolyte which served as a sacrificial electron donor to amplify the current signal. Formation of GPTMS film on ITO and subsequent antibody immobilization were characterized by X-ray photoelectron spectroscopy (XPS) and electrochemistry. Using a ruthenium tris(bipyridine)-conjugated IgG (IgG-Ru) as the surface-bound redox probe, the highest electrochemical signal was obtained on GPTMS electrodes with 1 h modification. Under the optimized conditions for ITO modification, antibody immobilization and tracer concentration, competition curves for benzo[a]pyrene and pyrenebutyric acid were obtained with a detection limit of 2.4 and 10 ng mLâ1, respectively. The redox-labeled electrochemical immunoassay with signal amplification mechanism offers a potential analytical method for the simultaneous detection of multiple environmental organic pollutants on antibody biochips.
Abstract: This paper presents an overview on the recent advance in the determination of environmental pollutants by immunosensors, which are classified as mass-based, optical and electrochemical sensors according to the signal transduction mechanism. New immunosensors based on total internal reflection fluorescence (TIRF), optical waveguide lightmode spectroscopy (OWLS) and surface plasma resonance (SPR), are described in detail. Advances in signal amplification, multiple-analyte determination, sensor regeneration, automation and miniaturization are discussed, followed by a perspective at the end of the paper.
Abstract: Binding and the redox reaction of the metallointercalator Ru(bpy)2(dppz)2+ (bpy = 2,2â-bipyridine, dppz = dipyrido[3,2-a:2â,3â-c]phenazine) with DNA was investigated by DNA film voltammetry. Calf-thymus DNA (CT-DNA) was assembled on a tin-doped indium oxide electrode by layer-by-layer electrostatic adsorption. Voltammetry of Ru(bpy)2(dppz)2+ (Ru-dppz) bound to the DNA film was measured in a redox-free electrolyte and showed strong dependence on the concentration of the metallointercalator. At low Ru-dppz concentrations, a single oxidation peak was observed, the potential of which shifted from 1.25 to 1.1 V with increasing Ru-dppz concentration (peak 1). At high metal chelate concentrations, an additional oxidation peak emerged with a potential of 1.25 V which was unaffected by the Ru-dppz concentration (peak 2). Three experiments were performed to investigate the mechanism and structural basis of the multiple peaks. First, voltammetry of Os(bpy)2(dppz)2+ bound to the CT-DNA film displayed only one peak at its oxidation potential of about 0.75 V. Second, the concentration dependence of Ru-dppz bound to a poly-(AU) film (which does not contain any guanine bases) exhibited only one oxidation peak at about 1.22 V that was independent of the Ru-dppz concentration. Third, when the guanine concentration in a mixed film of CT-DNA and poly-(AU) was changed and the bound Ru-dppz was kept constant, a pre-peak emerged and shifted to 1.1 V with increasing guanines. Based on these results, the appearance of two peaks in the voltammetric measurements of CT-DNA was rationalized by invoking two different DNA binding modes for the Ru-dppz complex:â intercalation and electrostatic association. Peak 2 arises from slow oxidation of guanines catalyzed by Ru-dppz electrostatically associated with the DNA film, since the addition of Mg2+ decreases the magnitude of peak 2. Peak 1 was not affected by Mg2+ ions, leading us to conclude that it is due to intercalated Ru-dppz. The intercalation positions the metal complex in close contact with the guanines inside DNA resulting in fast electrocatalytic reaction, giving rise to a catalytic pre-peak.
Abstract: DNA films immobilized on an indium tin oxide (ITO) electrode surface were detected and determined employing a high-affinity intercalator, Ru(bpy)2(dppz)2+ (bpy = 2,2â²-bipyridine, dppz = dipyrido[3,2-a:2â²,3â²-c]phenazine), as a redox indicator, and oxalate as a sacrificial electron donor in solution to chemically amplify the voltammetric signal of the indicator. Nucleic acids were immobilized on ITO by layer-by-layer electrostatic adsorption, using avidin as the first layer and nucleic acid as the second layer. In quartz crystal microbalance (QCM) measurements on an avidin-coated gold surface, the amount of adsorption from a 200âµgâmLâ1 nucleic acid solution was found to be 3.2âng mmâ2 for both double-stranded (ds-) and single-stranded (ss-) calf-thymus DNA as well as polycytidylic acid (Poly-C). After binding with Ru(bpy)2(dppz)2+ (Ru-dppz), voltammetry of the ds-DNA film on ITO was carried out in an indicator-free phosphate buffer. An anodic peak at about 1.15âV was observed, and it was assigned to Ru-dppz oxidation. When measured in an oxalate buffer, however, a catalytic current was observed due to the oxidation of oxalate by electrochemically generated Ru(bpy)2(dppz)3+, resulting in a 120-fold increase in the signal. Since oxalate itself produces a very low oxidation current on ITO, catalytic voltammetry produces about a 14-fold improvement in the signal-to-blank ratio over the non-amplified determination. As a result, ds-DNA adsorbed from 20ângâmLâ1 solution could be detected, which was estimated by QCM to be 160âpgâmmâ2 on the surface. The catalytic current of ds-DNA was substantially higher than that of ss-DNA and poly-C, indicative of selective binding of the redox indicator to ds-DNA. The results serve as a basis for the catalytic voltammetric detection of DNA hybridization in future work.
Abstract: Selective photoelectrochemical oxidation of DNA was achieved by ruthenium tris(bipyridine) immobilized on a tin oxide nanoparticle electrode. The metal complex was covalently attached to a protein, avidin, which adsorbed strongly on the tin oxide electrode by electrostatic interaction. Upon irradiation with 473-nm light, anodic photocurrent was generated in a blank electrolyte and was enhanced significantly after addition of poly(guanadylic acid) (poly-G) into the electrolyte. The current increased progressively with the nucleotide concentration, suggesting the enhancement effect was related to poly-G. The action spectrum indicates that the photocurrent was initiated by light absorption of the ruthenium compound immobilized on the electrode. Among the various polynucleotides examined, poly-G produced the largest photocurrent increase, followed by poly-A, single-stranded DNA, chemically damaged DNA, and double-stranded DNA, whereas poly-C and poly-U showed little effect. The combined experimental data support the hypothesis that the photoexcited Ru2+* species injects an electron into the semiconductor and produces Ru3+, which is then reduced back to Ru2+ by guanine and adenine bases in DNA, resulting in the recycling of the metal complex and enhanced photocurrent. The photoelectrochemical reaction can be employed as a new method for the detection of DNA damage.
