Biomedical subjects

Joseph Wang

Publications and source records attributed to Joseph Wang.

85 records · Page 5Linked to original sources

PubMed2002

Contactless conductivity detector for microchip capillary electrophoresis.

A microfabricated electrophoresis chip with an integrated contactless conductivity detection system is described. The new contactless conductivity microchip detector is based on placing two planar sensing aluminum film electrodes on the outer side of a poly(methyl methacrylate) (PMMA) microchip (without contacting the solution) and measuring the impedance of the solution in the separation channel. The contactless route obviates problems (e.g., fouling, unwanted reactions) associated with the electrode-solution contact, offers isolation of the detection system from high separation fields, does not compromise the separation efficiency, and greatly simplifies the detector fabrication. Relevant experimental variables, such as the frequency and amplitude of the applied ac voltage or the separation voltage, were examined and optimized. The detector performance was illustrated by the separation of potassium, sodium, barium, and lithium cations and the chloride, sulfate, fluoride, acetate, and phosphate anions. The response was linear (over the 20 microM-7 mM range) and reproducible (RSD = 3.4-4.9%; n = 10), with detection limits of 2.8 and 6.4 microM (for potassium and chloride, respectively). The advantages associated with the contactless conductivity detection, along with the low cost of the integrated PMMA chip/detection system, should enhance the power and scope of microfluidic analytical devices.

Anions↗

PubMed2002

Magnetically-induced solid-state electrochemical detection of DNA hybridization.

A magnetic triggering of a solid-state electrical transduction of DNA hybridization is described. Positioning of an external magnet below the thick-film electrode attracts the DNA/particle network and enables the solid-state electrochemical stripping detection of the silver tracer. TEM imaging indicates that the hybridization event results in a three-dimensional aggregate structure in which duplex segments link the metal nanoparticles and magnetic spheres, and that most of this assembly is covered with the silver precipitate. This leads to a direct contact of the metal tag with the surface (in connection to the magnetic collection) and enables the solid-state electrochemical transduction (without prior dissolution and subsequent electrodeposition of the metal), using oxidative dissolution of the silver tracer. No such aggregates (and hence magnetic "collection") are observed in the presence of noncomplementary DNA, that is, without the linking hybrid. The new method couples high sensitivity of silver-amplified assays with effective discrimination against excess of closely related nucleotide sequences (including single-base imperfections). Such direct electrical detection of DNA/metal-particle assemblies can bring new capabilities to the detection of DNA hybridization, and could be applied to other bioaffinity assays.

DNA↗

PubMed2002

Capillary electrophoresis-electrochemistry microfluidic system for the determination of organic peroxides.

A microfluidic analytical system for the separation and detection of organic peroxides, based on a microchip capillary electrophoresis device with an integrated amperometric detector, was developed. The new microsystem relies on the reductive detection of both organic acid peroxides and hydroperoxides at -700 mV (vs. Ag wire/AgCl). Factors influencing the separation and detection processes were examined and optimized. The integrated microsystem offers rapid measurements (within 130 s) of these organic-peroxide compounds, down to micromolar levels. A highly stable response for repetitive injections (RSD 0.35-3.12%; n = 12) reflects the negligible electrode passivation. Such a "lab-on-a-chip" device should be attractive for on-site analysis of organic peroxides, as desired for environmental screening and industrial monitoring.

Electrochemistry↗

PubMed2002

Optimal environment for glucose oxidase in perfluorosulfonated ionomer membranes: improvement of first-generation biosensors.

An optimal environment for glucose oxidase (GOx) in Nafion membranes is achieved using an advanced immobilization protocol based on a nonaqueous immobilization route. Exposure of glucose oxidase to water-organic mixtures with a high (85-95%) content of the organic solvent resulted in stabilization of the enzyme by a membrane-forming polyelectrolyte. Such an optimal environment leads to the highest enzyme specific activity in the resulting membrane, as desired for optimal use of the expensive oxidases. Casting solution containing glucose oxidase and Nafion is completely stable over 5 days in a refrigerator, providing almost absolute reproducibility of GOx-Nafion membranes. A glucose biosensor was prepared by casting the GOx-Nafion membranes over Prussian Blue-modified glassy carbon disk electrodes. The biosensor operated in the FIA mode allows the detection of glucose down to the 0.1 microM level, along with high sensitivity (0.05 A M(-1) cm(-2)), which is only 10 times lower than the sensitivity of the hydrogen peroxide transducer used. A comparison with the recently reported enzyme electrodes based on similar H2O2 transducers (transition metal hexacyanoferrates) shows that the proposed approach displays a dramatic (100-fold) improvement in sensitivity of the resulting biosensor. Combined with the attractive performance of a Prussian Blue-based hydrogen peroxide transducer, the proposed immobilization protocol provides a superior performance for first-generation glucose biosensors in term of sensitivity and detection limits.

Biosensing Techniques↗

PubMed2002

Single-channel microchip for fast screening and detailed identification of nitroaromatic explosives or organophosphate nerve agents.

A single-channel chip-based analytical microsystem that allows rapid flow injection measurements of the total content of organic explosive or nerve agent compounds, as well as detailed micellar chromatographic identification of the individual ones, is described. The protocol involves repetitive rapid flow injection (screening) assays--to provide a timely warning and alarm--and switching to the separation (fingerprint identification) mode only when harmful compounds are detected. While micellar electrokinetic chromatography, in the presence of sodium dodecyl sulfate (SDS), is used for separating the neutral nitroaromatic explosive and nerve agent compounds, an operation without SDS leads to high-speed measurements of the "total" explosives or nerve agent content. Switching between the "flow injection" and "separation" modes is accomplished by rapidly exchanging the SDS-free and SDS-containing buffers in the separation channel. Amperometric detection was used for monitoring the separation. Key factors influencing the sample throughput, resolution, and sensitivity have been assessed and optimized. Assays rates of about 360 and 30/h can thus be realized for the "total" screening and "individual" measurements, respectively. Ultimately, such development will lead to the creation of a field-deployable microanalyzer and will enable transporting the forensic laboratory to the sample source.

Chemical Warfare Agents↗

PubMed2002

Towards disposable lab-on-a-chip: poly(methylmethacrylate) microchip electrophoresis device with electrochemical detection.

A fully disposable microanalytical device based on combination of poly(methylmethacrylate) (PMMA) capillary electrophoresis microchips and thick-film electrochemical detector strips is described. Variables influencing the separation efficiency and amperometric response, including separation voltage or detection potential are assessed and optimized. The versatility, simplicity and low-cost advantages of the new design are coupled to an attractive analytical performance, with good precision (relative standard deviation RSD = 1.68% for n = 10). Applicability for assays of mixtures of hydrazine, phenolic compounds, and catecholamines is demonstrated. Such coupling of low-cost PMMA-based microchips with thick-film electrochemical detectors holds great promise for mass production of single-use micrototal analytical systems.

Catecholamines↗

PubMed2002

On-chip enzymatic assays.

This article reviews different possibilities for conducting enzymatic assays on microchip platforms, along with potential advantages, limitations, and selected examples of such biochips. Enzyme-based chips combine the analytical power and reagent economy of microfluidic devices with the selectivity and amplification features of biocatalytic reactions. "Lab-on-chip" devices thus allow enzymatic assays to be performed more rapidly, easily, and economically. Such assays usually rely on on-chip mixing and reactions (of the substrates and enzymes) in connection to separations (of the substrates or products). The realization of on-chip enzymatic assays thus requires understanding of how enzymatic reactions behave on a small scale and can be interfaced with separation microchips, and how the microfluidics can be tailored to suit the requirements of particular enzymatic assays. The goal is to obtain sufficient reaction times, without compromising the quality of the analytical separation. The versatility of such on-chip enzymatic assays offers great promise for decentralized testing of clinically or environmentally important substrates.

Electrophoresis, Capillary↗

PubMed2002

Microchip-based amperometric immunoassays using redox tracers.

A new chip-based electrochemical immunoassay protocol, based on the use of a ferrocene redox label, is described. Two reaction formats, based on direct (noncompetitive) and competitive modes of operation, were employed for illustrating the use of redox tracers in chip-based electrochemical immunoassays. The direct assay consisted of mixing the ferrocene-tagged antibody and the antigen analyte, a rapid electrophoretic separation of labeled free antibody and the labeled antigen/antibody complex, and a downstream anodic detection of the ferrocene tracer at gold-plated carbon screen-printed electrode detector. The competitive assay integrates precolumn reactions of the labeled antigen and the target antigen with the antibody with electrophoretic separation of the free and bound labeled antigens, along with amperometric detection of the redox tag. An internal standard has been used to normalize the peak area for the construction of calibration plots. Fundamental operating variables are examined and optimized. The use of a redox tracer offers the advantages of simplified protocol, wider linear range, higher stability, and higher separation efficiency compared to an analogous use of enzyme tags. The direct mouse-immunoglobulin G (IgG) assay and the competitive 3,3',5-triiodo-L-thyronine (T(3)) one were accomplished within less than 150 and 130 s (with field strengths of 256 and 192 V/cm), and offer minimum detectable concentrations of 2.5 x 10(-12) and 1 x1 0(-6) g/mL, respectively. Such use of redox labels for chip-based amperometric immunoassay protocols offers considerable promise for decentralized clinical or environmental testing.

Antigen-Antibody Complex↗

PubMed2002

Oxygen-independent poly(dimethylsiloxane)-based carbon-paste glucose biosensors.

Several silicone oils have been assessed and compared as an internal source of oxygen in connection to their use as binders for carbon-paste glucose biosensors. All four poly(dimethylsiloxane) (PDMS) oils tested a dramatic increase in the oxygen capacity of carbon-paste enzyme electrodes to allow convenient biosensing under severe oxygen-deficit conditions. The resulting oxygen independence is better than that exerted by perfluorocarbon binders or that displayed by mediator-based bioelectrodes. The resistance to oxygen effects is indicated from the identical response (observed in the presence and absence of oxygen) up to 2 x 10(-2) M glucose and the slight (12%) sensitivity loss at 4 x 10(-2) M. The influence of the viscosity of the PDMS binder upon the internal oxygen supply is examined. The PDMS carbon-paste enzyme electrode displays a stable glucose response over prolonged (15 h) operation in an oxygen-free solution. On-line continuous testing indicates favorable dynamic properties with no carry-over effects over the physiological and pathophysiological range (3-12 mM glucose).

Biosensing Techniques↗

PubMed2002

Real-time electrochemical monitoring: toward green analytical chemistry.

This Account presents a survey of recent advances in electrochemical sensing technology relevant to green analytical chemistry and examines the potential advantages, limitations, and applications of these monitoring devices. Stricter environmental control and effective process monitoring have created considerable demands for innovative analytical methodologies. New devices and protocols, with negligible waste generation or no hazardous substances, and in situ real-time monitoring capability are particularly needed for addressing the challenges of green analytical chemistry. The coupling of modern electrochemical detection principles with recent advances in molecular recognition, microelectronics, and microfabrication has led to powerful, compact, and "user-friendly" analytical devices. The unique features of such electrochemical monitoring systems make them particularly attractive for addressing environmental and industrial problems and the challenges of green chemistry. These developments allow the instrument to be taken to the sample (rather than the traditional way of bringing the sample to the laboratory) and hence to ensure effective process or pollution control.

Biosensing Techniques↗

PubMed2002

Amplified label-free electrical detection of DNA hybridization.

A new protocol is described for amplifying label-free electrochemical measurements of DNA hybridization based on the enhanced accumulation of purine nucleobases in the presence of copper ions . Such electrical DNA assays involve hybridization of the target to inosine-substituted oligonucleotide probes (captured on magnetic beads), acidic dipurinization of the hybrid DNA, and adsorptive chronopotentiometric stripping measurements of the free nucleobases in the presence of copper ions. Both amplified adenine and guanine peaks can be used for detecting the DNA hybridization. The dramatic signal amplification advantage of this type of detection has been combined with efficient magnetic removal of non-complementary DNA, use of microliter sample volumes and disposable transducers. Factors influencing the signal enhancement were assessed and optimized. A detection limit of 40 fmol (250 pg) was obtained with 10 min hybridization and 5 min adsorptive-accumulation times. The advantages of this procedure were demonstrated by its application in the detection of DNA segments related to the BRCA1 breast cancer gene. The copper enhancement holds great promise not only for the detection of DNA hybridization, but also for trace measurement of nucleic acids.

Animals↗

PubMed2002

A chip-based capillary electrophoresis-contactless conductivity microsystem for fast measurements of low-explosive ionic components.

A miniaturized analytical system for separating and detecting inorganic explosive residues, based on the coupling of a micromachined capillary electrophoresis (CE) chip with a contactless conductivity detector is described. The low electroosmotic flow (EOF) of the poly(methylmethacrylate) (PMMA) chip material facilitates the rapid switching between analyses of cations and anions using the same microchannel and run buffer (and without an EOF modifier), and hence offers rapid (< 1 min) measurement of seven explosive-related cations and anions. Experimental parameters relevant to the separation and detection processes have been optimized. Addition of a 18-crown-6 ether modifier has been used for separating the peaks of co-migrating potassium and ammonium ions. The ionic-explosive microchip system combines the distinct advantages of contactless conductivity detection with the attractive features of plastic CE microchips. The new microsystem offers great promise for monitoring explosive-related ions at the sample source, with significant advantages of speed/warning, efficiency, cost, or sample size.

Electrophoresis, Capillary↗

PubMed2002

Dual enzyme electrochemical coding for detecting DNA hybridization.

Enzyme-based hybridization assays for the simultaneous electrochemical measurements of two DNA targets are described. Two encoding enzymes, alkaline phosphatase and beta-galactosidase, are used to differentiate the signals of two DNA targets in connection to chronopotentiometric measurements of their electroactive phenol and alpha-naphthol products. These products yield well-defined and resolved peaks at +0.31 V (alpha-naphthol) and +0.63 V (phenol) at the graphite working electrode (vs. Ag/AgCl reference). The position and size of these peaks reflect the identity and level of the corresponding target. The dual target detection capability is coupled to the amplification feature of enzyme tags (to yield fmol detection limits) and with an efficient magnetic removal of non-hybridized nucleic acids. Proper attention is given to the choice of the substrates (for attaining well resolved peaks), to the activity of the enzymes (for obtaining similar sensitivities), and to the selection of the enzymes (for minimizing cross interferences). The new bioassay is illustrated for the simultaneous detection of two DNA sequences related to the BCRA1 breast-cancer gene in a single sample in connection to magnetic beads bearing the corresponding oligonucleotide probes. Prospects for electrochemical coding of multiple DNA targets are discussed.

Alkaline Phosphatase↗