A strategy for constructing a hybrid bilayer membrane based on a carbon substrate

We construct a hybrid bilayer membrane (HBM) on a new substrate-carbon electrode. It is an extension of HBM based on other substrates. Primary alkylamine was chemically modified onto the surface of a carbon electrode by electrochemical scans; thus, a monolayer was formed on the electrode. Because the alkane chains section is toward the outside, a hydrophobic surface was constructed. Then a lipid monolayer was spread on the hydrophobic surface of the carbon electrode. The formed HBM was characterized by electrochemical and ATR-FT-IR methods. From ATR-FT-IR results, the lipid order parameter (S) of 0.73 was obtained. This kind of hybrid membrane has the advantages of a lipid/alkanethiol HBM. A potential application of this HBM as a biosensor (detecting K+) was given.

Flow injection electrochemical enzyme immunoassay based on the use of an immunoelectrode strip integrate immunosorbent layer and a screen-printed carbon electrode

A flow injection amperometric immunoassay system based on the use of screen-printed carbon electrode for the detection of mouse IgG was developed. An immunoelectrode strip, on which an immunosorbent layer and screen-printed carbon electrode were integrated, and a proposed flow cell have been fabricated. The characterization of the flow immunoassay system and parameters affecting the performance of the immunoassay system were studied and optimized. Amperometric detection at 0.0 V (versus Ag/AgCl) resulted in a linear detection range of 30-700 ng ml(-1), with a detection limit of 3 ng ml(-1). The signal variation among electrode strips prepared from variant batch did not exceed 8.5% (n = 7) by measuring 0.5 mug ml(-1) antigen standard solution.

Layer-by-layer assembly of multilayer films composed of avidin and biotin-labeled antibody for immunosensing

Protein multilayers composed of avidin and biotin-labeled antibody (bio-Ab) were prepared on gold surface by layer-by-layer assembly technology using the high specific binding constant (K-a: approximate to 10(15) M-1) between avidin and biotin. The assembly process of the multilayer films was monitored by using real-time BIA technique based on surface plasmon resonance (SPR). The multilayer films were also characterized by electrochemical impedance spectroscopy (EIS) and reflection absorption Fourier transform infrared spectroscopy (FTIR). The results indicate that the growth of the multilayer is uniform. From response of SPR for each layer, the stoichiometry S for the interaction between avidin and bio-Ab is calculated to be 0.37 in the multilayer whereas 0.82 in the first layer. The protein mass concentration for each layer was also obtained. The schematic figure for the multilayer assembly was proposed according to the layer mass, concentration and S value. The utility of the mutilayer films for immunosensing has been investigated via their subsequent interaction with hIgG. The binding ability of the multilayer increased for one to three layers of antibody, and then reach saturation after the fourth layer. These layer-by-layer constructed antibody multilayers enhance the binding ability than covalently immobilized monolayer antibody. This technology can be also used for construction of other thin films for immunosensing and biosensor.

Kinetic characteristics of the interaction between morphine and its antibody by SPR competitive immunoassay

It is impossible for surface plasmon resonance to measure directly the binding kinetics between a low-molecular-weight analyte interacting and its immobilized binding partner. Solution competition method was applied to the kinetic study of the interaction between morphine and its antibody. The affinity constant between the antibody of morphine and morphine-BSA immobilized on the sensor chip was also obtained. The result showed that the affinity of polyclonal antibody is stronger than that of monoclonal antibody. And it also indicated that the protein combined with the analyte affected the binding of antibody to antigen.

Electrochemical and Raman studies of the biointeraction between Escherichia coli and mannose in polydiacetylene derivative supported on the self-assembled monolayers of octadecanethiol on a gold electrode

Here, we describe a new method to study the biointeraction between Escherichia coli and mannose by using supramolecular assemblies composed of polydiacetylene supported on the self-assembled monolayer of octadecanethiol on a gold electrode. These prepared bilayer materials simply are an excellent protosystem to study a range of important sensor-related issues. The experimental results from UV-vis spectroscopy, resonance Raman spectroscopy, and electrochemistry confirm that the specific interactions between E. coli and mannose can cause conformational changes of the polydiacetylene backbone rather than simple nonspecific adsorption. Moreover, the direct electrochemical detection by polydiacetylene supramolecular assemblies not only opens a new path for the use of these membranes in the area of biosensor development but also offers new possibilities for diagnostic applications and screening for binding ligands.

Direct electrochemistry of hemoglobin in egg-phosphatidylcholine films and its catalysis to H2O2

Direct electrochemistry of hemoglobin was observed in stable thin film composed of a natural lipid (egg-phosphatidylcholine) and hemoglobin on pyrolytic graphite (PG) electrode. Hemoglobin in lipid films shows thin layer electrochemistry behavior. The formal potential Edegrees' of hemoglobin in the lipid film was linearly varied with pH in the range from 3.5 to 7.0 with a slope of -46.4 mV pH(-1) Hemoglobin in the lipid film exhibited elegant catalytic activity for electrochemical reduction of H202, based which a unmediated biosensor for H2O2 was developed.

Formation of a supported hybrid bilayer membrane on gold: A sterically enhanced hydrophobic effect

Adsorption of a monolayer of didecanoyl-L-alpha-phosphatidylcholine (DDPC) from dispersions of small unilamellar vesicles onto hydrophobic surfaces was investigated by mean of cyclic voltammetry and impedance spectroscopy. The hydrophobic surfaces were self-assembled monolayers of 2-mereapto-3-n-octylthiophene (MOT) on gold. One characteristic of the MOT monolayer is its permeability to organic molecules in aqueous solution, thus providing a more energetically favorable hydrophobic surface for the addition of phospholipid vesicles. The kinetics of the lipid monolayer formation were followed by measuring the time-dependent interfacial capacitance. Unusual values of thickness and capacitance of the MOT/ DDPC bilayers were observed. An interdigitating conformation of the bilayer structure was proposed to interpret the experimental results, The horseradish peroxidase reconstituted into the bilayer demonstrated the expected protein activity, showing practical use in research and in biosensor application.

Amplified immunoassay of human IgG using real-time biomolecular interaction analysis (BIA) technology

An automated biomolecular interaction analysis instrument (BI-Acore) based on surface plasmon resonance (SPR) has been used to determine human immunoglobulin G (IgG) in real time. Polyclonal anti-human IgG antibody was covalently immobilized to a carboxymethyldextran modified gold film surface. The samples of human IgG prepared in HBS buffer were poured over the immobilized surface. The signal amplification antibody was applied to amplify the response signal. After each measurement, the surface was regenerated with 0.1 mol/L H3PO4. The assay was rapid, requiring only 30 min for antibody immobilization and 20 min for each subsequent process of immune binding, antibody amplification and regeneration. The antibody immobilized surface had good response to human IgG in the range of 0.12-60 nmol/L with a detection limit of 60 pmol/L. The same antibody immobilized surface could be used for more than 110 cycles of binding, amplification and regeneration. The results demonstrate that the sensitivity, specificity and reproducibility of amplified immunoassay using real-time BIA technology are satisfactory.

Kinetic analysis of TNF immune complex by using surface plasmon resonane

The kinetic analysis of the interaction between tumor necrosis factor(TNF) and its monoclonal antibody was performed by surface plasmon resonance(SPR) technique. The monoclonal antibody was immobilized to the surface of CM5 sensor chip by amine coupling. TNF at different concentrations was injected across the mAb immobilized surface. The interaction was recorded in real time and could be seen on the sensorgram. One cycle, including association, dissociation and regeneration, lasted no more than 15 min. The interaction results was evaluated using 1 : 1 Langmuir binding model. The kinetic rate constants were calculated to be: k =1.68 X 10(3) L (.) mol(-1) (.) s(-1), k(d) = 1.73 X 10(-4) s(-1), and the affinity constants K-A = 9. 7 X 10(3) L (.) mol(-1), K-r)= 1. 03 X 10(-7) Mol (.) L-1. The X-2 was 3.47, which showed that the interaction is consistent with the 1 : I model. We can see from the results that although there are two binding sites in one mAb molecule, TNF reacts with each site in an independent and noncooperative manner.

Progress in deoxyribonucleic acid electroanalysis

The electroanalysis research of deoxyribonucleic acid (DNA) is an attractive project in life science. This paper reviews the recent progress of DNA electrochemical analysis, which includes DNA electroanalysis and DNA electrochemical biosensors. The prospects of the research are very bright. 77 papers are cited.

Enhanced surface plasmon resonance immunoassay for human complement factor 4

Using an enhanced surface plasmon resonance (SPR) immunosensor, we have determined the concentration of human complement factor 4 (C4). Antibody protein was concentrated into a carboxymethyldextran-modified gold surface by electrostatic attraction force and a simultaneous covalent immobilization of antibody based on amine coupling reaction took place. The sandwich method was applied to enhance the response signal and the specificity of antigen binding assay. The antibody immobilized surface had good response to C4 in the range of 0.02-20 mug/ml by this enhanced immunoassay. The regeneration effect by pH 2 glycine-HC1 buffer was also investigated. The same antibody immobilized surface could be used more than 80 cycles of C4 binding and regeneration. In addition, the ability to determinate C4 directly from serum sample without any purification was investigated. The sensitivity, specificity and reproducibility of the enhanced immunoassay are satisfactory. The results clearly demonstrate the advantages of the enhanced SPR technique for C4 immunoassay.

Electrochemical biosensing in extreme environment

This review presents recent developments of electrochemical biosensors in extreme working environments. After a brief introduction to the electrochemical biosensor, the applications of biocatalytic biosensors and bioaffinity biosensors in harsh working conditions, in organic solvent, in gas-phase, in vivo measurement and in toxic environments, are discussed by means of several examples. Methods for improving the stability and extending the biosensor application scope are suggested, and new trends about biosensor development are also discussed.

Organic-phase enzyme electrode for phenolic determination based on a functionalized sol-gel composite

An amperometric biosensor for monitoring phenols in the organic phase was constructed by the silica sol-gel immobilization of tyrosinase on a glassy carbon electrode. The organic-inorganic hybrid materials with different sol-gel precursors and polymers were optimized, and the experimental conditions, such as the effect of the solvent, operational potential and enzyme loading were explored for the optimum analytical performance of the enzyme electrode. The biosensor can reach 95% of steady-state current in about 18 s, and the trend in the sensitivity of different phenols is as follows: catechol > phenol >p-cresol. In addition, the apparent Michaelis-Menten constants (K-m(app)) and the stability of the enzyme electrode were discussed. (C) 2000 Elsevier Science S.A. All rights reserved.

Species of rare earth (III), calcium (II) and zinc (II) in the presence of L-histidine and L-typtophane

Rare earth(III)-histidine (His)- tryptophane (Trp). Ca(II)-His-Trp and Zn(II)-His-Trp systems were studied by potentiometric titration and computer simulation under physiological conditions. The species of the systems and their stability constants were determined. The distributions of species of rare earth(III), Ca(II) and Zn(II) were discussed.

Bilayer lipid membranes and their application in electrochemical biosensors

The preparation and characteristics of bilayer lipid membranes including conventional bilayer membrane, solid supported self-assembling bilayer lipid membrane, solid supported hybrid bilayer membrane are described in this paper, The applications of bilayer lipid membranes in electrochemical biosensors are reviewed and the future development of electrochemical biosensor based on bilayer lipid membranes is discussed.