Label-free electrochemical immunosensor for the carcinoembryonic antigen using a glassy carbon electrode modified with electrodeposited Prussian Blue, a graphene and carbon nanotube assembly and an antibody immobilized on gold nanoparticles

We described a sensitive, label-free electrochemical immunosensor for the detection of carcinoembryonic antigen. It is based on the use of a glassy carbon electrode (GCE) modified with a multi-layer films made from Prussian Blue (PB), graphene and carbon nanotubes by electrodeposition and assembling techniques. Gold nanoparticles were electrostatically absorbed on the surface of the film and used for the immobilization of antibody, while PB acts as signaling molecule. The stepwise assembly process was investigated by differential pulse voltammetry and scanning electron microscopy. It is found that the formation of antibody-antigen complexes partially inhibits the electron transfer of PB and decreased its peak current. Under the optimal conditions, the decrease of intensity of the peak current of PB is linearly related to the concentration of carcinoembryonic antigen in two ranges (0.2–1.0, and 1.0–40.0 ng·mL^?1), with a detection limit of 60 pg·mL^?1 (S/N?=?3). The immunosensor was applied to analyze five clinical samples, and the results obtained were in agreement with clinical data. In addition, the immunosensor exhibited good precision, acceptable stability and reproducibility.

Preparation of Fe3O4@C@CNC multifunctional magnetic core/shell nanoparticles and their application in a signal-type flow-injection photoluminescence immunosensor

We describe here the preparation of carbon-coated Fe₃O₄ magnetic nanoparticles that were further fabricated into multifunctional core/shell nanoparticles (Fe₃O₄@C@CNCs) through a layer-by-layer self-assembly process of carbon nanocrystals (CNCs). The nanoparticles were applied in a photoluminescence (PL) immunosensor to detect the carcinoembryonic antigen (CEA), and CEA primary antibody was immobilized onto the surface of the nanoparticles. In addition, CEA secondary antibody and glucose oxidase were covalently bonded to silica nanoparticles. After stepwise immunoreactions, the immunoreagent was injected into the PL cell using a flow-injection PL system. When glucose was injected, hydrogen peroxide was obtained because of glucose oxidase catalysis and quenched the PL of the Fe₃O₄@C@CNC nanoparticles. The here proposed PL immunosensor allowed us to determine CEA concentrations in the 0.005–50 ng?·?mL^-1 concentration range, with a detection limit of 1.8 pg?·?mL^-1.

Label-free sandwich type of immunosensor for hepatitis C virus core antigen based on the use of gold nanoparticles on a nanostructured metal oxide surface

We report on the construction of a label-free electrochemical immunosensor for detecting the core antigen of the hepatitis C virus (HCV core antigen). A glassy carbon electrode (GCE) was modified with a nanocomposite made from gold nanoparticles, zirconia nanoparticles and chitosan, and prepared by in situ reduction. The zirconia nanoparticles were first dispersed in chitosan solution, and then AuNPs were prepared in situ on the ZrO₂-chitosan composite. In parallel, a nanocomposite was synthesized from AuNPs, silica nanoparticles and chitosan, and conjugated to a secondary antibody. The properties of the resulting nanocomposites were investigated by UV-visible photometry and transmission electron microscopy, and the stepwise assembly process was characterized by means of cyclic voltammetry and electrochemical impedance spectroscopy. An sandwich type of immunosensor was developed which displays high sensitivity to the HCV core antigen in the concentration range between 2 and 512?ng?mL^?1, with a detection limit of 0.17?ng?mL^?1 (at S/N?=?3). This immunosensor provides an alternative approach towards the diagnosis of HCV.

Disposable electrochemical immunosensor for myeloperoxidase based on the indium tin oxide electrode modified with an ionic liquid composite film containing gold nanoparticles, poly(o-phenylenediamine) and carbon nanotubes

A disposable electrochemical myeloperoxidase (MPO) immunosensor was fabricated based on the indium tin oxide electrode modified with a film composed of gold nanoparticles (AuNPs), poly(o-phenylenediamine), multi-walled carbon nanotubes and an ionic liquid. The composite film on the surface of the electrode was prepared by in situ electropolymerization using the ionic liquid as a supporting electrolyte. Negatively charged AuNPs were then adsorbed on the modified electrode via amine-gold affinity and to immobilize MPO antibody. Finally, bovine serum albumin was employed to block possible remaining active sites on the AuNPs. The modification of the electrode was studied by cyclic voltammetry and scanning electron microscopy. The factors affecting the performance of the immunosensor were investigated in detail using the hexacyanoferrate redox system. The sensor exhibited good response to MPO over two linear ranges (from 0.2 to 23.4 and from 23.4 to 300 ng.mL^?1), with a detection limit of 0.05 ng.mL^?1 (at an S/N of 3).

A nanocomposite made from conducting organic polymers and multi-walled carbon nanotubes for the adsorption and separation of gold(III) ions

We describe a new method for the separation and preconcentration of traces of Au(III) in environmental samples. Sorbents made from modified multiwalled carbon nanotubes and conducting polymers (PANI and PEDOT) were used for solid-phase extraction. The Au(III) ions are adsorbed as a result of the interaction with the electron pairs of =N- and -S- groups. Effects of pH value, flow rate and volume of sample, type, volume and concentration of eluent, and the adsorption capacity were investigated. The maximum adsorption capacity of MWCNTs/PANI and MWCNTs/PEDOT are 159 and 176?mg?g^?1, and the detection limits of this method are below 0.3 and 0.5?ng?mL^?1, respectively. The procedure was successfully applied to the determination of traces of Au(III) in a reference material and in environmental samples.

Hydrogen peroxide sensor based on a stainless steel electrode coated with multi-walled carbon nanotubes modified with magnetite nanoparticles

Multi-walled carbon nanotubes (MWCNTs) were decorated with magnetite (Fe₃O₄) nanoparticles and then used to modify a stainless steel electrode. The Fe₃O₄/MWCNTs composite was characterized by scanning electron microscopy, transmission electron microscopy, Raman spectroscopy and X-ray diffraction patterns. Electrochemical properties of the modified electrode revealed a substantial catalytic activity for the reduction of hydrogen peroxide. The relationship between peak current and the concentration of hydrogen peroxide was linear in the range from 0.06?mmol?L^?1 to 0.36?mmol?L^?1, and the lowest detectable concentration is 0.01?mmol·L^?1 (S/N?=?3). The modified stainless steel electrode displays excellent stability.

Gold nanoparticle-coated multiwall carbon nanotube-modified electrode for electrochemical determination of methyl parathion

We report on an electrochemical sensor for the determination of methyl parathion. It is based on an electrode modified with multi-walled carbon nanotubes that were covered with gold nanoparticles (Au-NPs). The vertically aligned array of MWCNTs on a tantalum substrate was coated with Au-NPs by overhead magnetron sputtering deposition. Scanning and transmission electron microscopy and XRD were used to characterize the Au-NP-MWCNTs composite. Cyclic voltammetry and differential pulse voltammetry were employed to evaluate the suitability of the new electrode for the determination of methyl parathion. Under the optimal conditions, the current response of the electrode to methyl parathion is linear in the range from 0.50 to 16.0 mg mL^-1, with a detection limit of 50 ??g mL^-1 (signal/noise = 3), and the sensitivity is 4.5 times better than that of the plain MWCNTs electrode. We conclude that this method represents a simple, rapid, effective and sensitive approach for the detection of methyl parathion.

One-step electrochemical immunosensing for simultaneous detection of two biomarkers using thionine and ferrocene as distinguishable signal tags

We report on a new kind of electrochemical immunosensors for simultaneous determination of the biomarkers carcinoembryonic antigen (CEA) and alpha-fetoprotein (AFP). Thionine and ferrocene were applied as distinguishable electrochemical tags (and mediators) which were covalently conjugated on anti-AFP and anti-CEA antibodies, respectively, via carboxy groups. The resulting conjugates were co-immobilized on a glassy carbon electrode functionalized with gold nanoparticles. Finally, horseradish peroxidase (HRP) was immobilized onto the modified electrode. Labeled thionine and ferrocene, respectively, act as distinguishable tags for simultaneous determination of AFP and CEA due to the difference in the location of their voltammetric peaks. With a one-step immunoassay format, the analytes in the sample produced transparent immunoaffinity reaction with the corresponding antibodies on the electrode. Once the immunocomplex is formed, it partially inhibits the active center of the immobilized HRP, and this decreased the activity of HRP in terms of reduction of hydrogen peroxide. This immunosensor enables the simultaneous determination of AFP and CEA in a single run and within the same dynamic range (0.01–50?ng?mL^?1) and the same lower detection limit (0.01?ng?mL^?1). The reproducibility and stability of the immunosensors are acceptable. The dual immunosensor was applied to evaluate several specimens, and the assay results are in acceptable agreement with clinical data.

Electrochemical immunoassay for subgroup J of avian leukosis viruses using a glassy carbon electrode modified with a film of poly (3-thiophene boronic acid), gold nanoparticles, graphene and immobilized antibody

We have modified a glassy carbon electrode (GCE) with a film of poly(3-thiophene boronic acid), gold nanoparticles and graphene, and an antibody (Ab) was immobilized on its surface through the covalent bond formed between the boronic acid group and the glycosyl groups of the Ab. Subgroup J of avian leukosis viruses (ALV-J) were electrochemically determined with the help of this electrode. There is a linear relationship between the electron transfer resistance (R _et) and the concentration of ALV-J in the range from 527 to 3,162 TCID₅₀?mL^?1 (where TCID₅₀ is the 50?% tissue culture infective dose). The detection limit is 210 TCID₅₀?mL^?1 (at an S/N of 3), and the correlation coefficient (R) is 0.9964. The electrochemical immunoassay showed good selectivity, stability and reproducibility.

Highly sensitive deoxynivalenol immunosensor based on a glassy carbon electrode modified with a fullerene/ferrocene/ionic liquid composite

We describe a sensitive electrochemical immunosensor for the detection of deoxynivalenol (DON). It is based on a glassy carbon electrode modified with a composite made from fullerene (C₆₀), ferrocene and the ionic liquid. The components were immobilized on the surface of the electrode using chitosan cross-linked with epichlorohydrin. Then, the antibody to DON was covalently conjugated to the surface which then was blocked with serum albumin. The performance of the immunosensor was investigated by cyclic voltammetry and electrochemical impedance spectroscopy. It offers good repeatability (RSD?=?1.2%), selectivity, a stability of more than 180?days, an impedimetric response to DON in the range of 1?pgmL^?1 to 0.3?ng?mL^?1, and a detection limit (at S/N?=?3) of 0.3?pgmL^?1. The limit of detection is better than that of GC, HPLC, GC-MS, HPLC-MS and LC-MS-MS. The effects of omitting C₆₀ or the ionic liquid were also examined. The results indicate that the sensitivity of the biosensor is 2-fold better if C₆₀ and ionic liquids are used. This demonstrates that C₆₀ facilitates electron transfer on the surface of the modified electrode due to its unique electrochemical properties, while the ionic liquid provides a biocompatible microenvironment for the antibody. This results in increased sensitivity and stability. The method was satisfactorily applied to the determination of DON in food samples.

Prussian blue-doped nanogold microspheres for enzyme-free electrocatalytic immunoassay of p53 protein

We report on a new enzyme-free electrochemical immunoassay for the sensitive detection of the p53 protein (p53; a model analyte) by using a screen-printed carbon electrode modified with monoclonal mouse anti-human p53 antibody tagged with gold nanoparticles. First, nanogold microspheres doped with Prussian Blue were synthesized by a reverse micelle method. The resulting microspheres were used to label polyclonal anti-p53 antibody which then was applied in a sandwich immunoassay in pH 6.5 buffer solution using the Prussian Blue in the particles as the redox-active reporter. The electrochemical signal of the immunosensor is shown to increase with the concentration of the analyte (p53 protein) in the range from 0.5 to 80 U mL^?1, with a detection limit of 0.1 U mL^?1. No non-specific adsorption was observed. Coefficients of variation for intra-assay and inter-assay were below 8.5 % and 11.5 %, respectively. In addition, the method was applied to the analysis of 15 human serum samples, and a good relationship was found between the new immunoassay and the referenced electro-chemiluminescence method.

Biocompatible and label-free amperometric immunosensor for hepatitis B surface antigen using a sensing film composed of poly(allylamine)-branched ferrocene and gold nanoparticles

An amperometric immunosensor has been developed for sensitive determination of hepatitis B surface antigen as a model protein. A glassy carbon electrode was modified with an assembly of positively charged poly(allylamine)-branched ferrocene (PAA-Fc) and negatively charged gold nanoparticles (Au NPs). The formation of PAA-Fc effectively avoids the leakage of Fc, retains its electrochemical activity, and enhances the conductivity of the composite. The adsorption of Au NPs onto the PAA-Fc matrix provides sites for the immobilization of the antigen and a favorable micro-environment to maintain its activity. The morphologies and electrochemistry of the sensing film were investigated via scanning electron microscopy, electrochemical impedance spectroscopy, and cyclic voltammetry. Factors influencing the performance of the immunosensor were studied in detail. The concentration of the antigen can be quantitated (by measuring the decrease of the amperometric response resulting from the specific binding between antigen and antibody) in the range between 0.1 and 150?ng?mL^?C1, with a detection limit of 40?pg?mL^?C1 (S/N = 3). The method is economical, efficient, and potentially attractive for clinical immunoassays.

Anodic stripping voltammetry of silver(I) using a carbon paste electrode modified with multi-walled carbon nanotubes

We describe a silver(I)-selective carbon paste electrode modified with multi-walled carbon nanotubes and a silver-chelating Schiff base, and its electrochemical response to Ag(I). Effects of reduction potential and time, accumulation time, pH of the solution and the stripping medium were studied by differential pulse anodic stripping voltammetry and optimized. The findings resulted in a method for the determination of silver over a linear response range (from 0.5 to 235 ng?mL^?1) and with a detection limit as low as 0.08 ng?mL^?1. The sensor displays good repeatability (with the RSD of ±?2.75 % for 7 replicates) and was applied to the determination of Ag(I) in water samples and X-ray photographic films.

A lead(II) sensor based on a glassy carbon electrode modified with Fe3O4 nanospheres and carbon nanotubes

We have developed a highly sensitive and selective sensor for lead(II) ions. A glassy carbon electrode was modified with Fe₃O₄ nanospheres and multi-walled carbon nanotubes, and this material was characterized by scanning electron microscopy and X-ray diffraction. The electrode displays good electrochemical activity toward Pb(II) and gives anodic and cathodic peaks with potentials at ?496 mV and ?638 mV (vs. Ag/AgCl) in pH?6.0 solution. The sensor exhibits a sensitive and fairly selective response to Pb(II) ion, with a linear range between 20 pM and 1.6 nM, and a detection limit as low as 6.0 pM (at a signal-to noise ratio of 3). The sensor was successfully applied to monitor Pb(II) in spiked water samples.

Electrochemiluminescence immunoassay at a nanoporous gold leaf electrode and using CdTe quantun dots as labels

An electrochemiluminescence-based immunoassay using quantum dots (QDs) as labels for the carcinoembryonic antigen (CEA) was developed using an electrode modified with leafs of nanoporous gold. CEA was initially immobilized on the electrode via a sandwich immunoreaction, and then CdTe quantum dots capped with thioglycolic acid were used to label the second antibody. The intensity of the ECL of the QDs reflects the quantity of CEA immobilized on the electrode. Thus, in the presence of dithiopersulfate as the coreactant, the ECL serves as the signal for the determination of CEA. The intensity of the electroluminescence (ECL) of the electrode was about 5.5-fold higher than that obtained with a bare gold electrode. The relation between ECL intensity and CEA concentration is linear in the range from 0.05 to 200?ng.mL^-1, and the detection limit is 0.01?ng.mL^-1. The method has the advantages of high sensitivity, good reproducibility and long-term stability, and paves a new avenue for applying quantum dots in ECL-based bioassays.

Non-enzymatic analysis of glucose on printed films based on multi-walled carbon nanotubes

We report on the fabrication of an enzyme–free electrochemical sensor for glucose based on a printed film consisting of multi–walled carbon nanotubes (MWCNTs). The MWCNT–based film can be produced by means of a flexographic printing process on a polycarbonate (PC) substrate. The electrochemical response of the MWCNT–based film (referred to as MWCNT–PC) towards the oxidation of glucose at pH 7 was studied by means of cyclic voltammetry and electrochemical impedance spectroscopy. The MWCNT–PC film exhibits substantial electrocatalytic activity towards the oxidation of glucose at an anodic potential of 0.30?V (vs. Ag/AgCl). The findings reveal that the MWCNT–PC film enables non–enzymatic sensing of glucose with a detection limit as low as 2.16?μM and a sensitivity of 1045?μA?mM^?1?cm^?2.

Disposable immunoassay for hepatitis B surface antigen based on a graphene paste electrode functionalized with gold nanoparticles and a Nafion-cysteine conjugate

We report on the modification of a graphene paste electrode with gold nanoparticles (AuNPs) and a Nafion-L-cysteine composite film, and how this electrode can serve as a platform for the construction of a novel electrochemical immunosensor for the detection of hepatitis B surface antigen (HBsAg). To obtain the immunosensor, an antibody against HBsAg was immobilized on the surface of the electrode, and this process was followed by cyclic voltammetry and electrochemical impedance spectroscopy. The peak currents of a hexacyanoferrate redox system decreased on formation of the antibody-antigen complex on the surface of the electrode. Then increased electrochemical response is thought to result from a combination of beneficial effects including the biocompatibility and large surface area of the AuNPs, the high conductivity of the graphene paste electrode, the synergistic effects of composite film, and the increased quantity of HBsAb adsorbed on the electrode surface. The differential pulse voltammetric responses of the hexacyanoferrate redox pair are proportional to the concentration of HBsAg in the range from 0.5–800?ng?mL^?1, and the detection limit is 0.1?ng?mL^?1 (at an S/N of 3). The immunosensor is sensitive and stable.

An electrochemical immunosensor for the tumor marker α-fetoprotein using a glassy carbon electrode modified with a poly(5-formylindole), single-wall carbon nanotubes,and coated with gold nanoparticles and antibody

We report on a label-free electrochemical immunosensor for α-fetoprotein (α-FP). It is based on the use of a glassy carbon electrode that was first modified with conducting poly(5-formylindole) and single-walled carbon nanotubes (P5FIn/SWNTs), and then coated with gold nanoparticles and the respective antibody. The presence of aldehyde groups warrants direct immobilization of the antibody and results in a convenient method for fabricating of the immunosensor. Gold nanoparticles (GNPs) were deposited on the P5FIn/SWNTs composite material, and the modified electrode was applied to the detection of α-FP. The analytical signal is obtained by measuring the change of amperometric response at a typical working voltage of 100 mV before and after the immunoreaction. The detection limit is 200 fg mL^?1. The immunosensor is simple, sensitive, specific and reproducible. It has the potential for reliable point-of-care diagnosis of tumor or other diseases. Figure

A simple electrochemical immunosensor based on conducting poly(5-formylindole) and single-walled carbon nanotubes composite was fabricated to detect alpha-fetoprotein. The detection limit is 200 fg mL^?1. This immunosensor is simple, sensitive, specific and reproducible.     

Multilayer structured immunosensor based on a glassy carbon electrode modified with multi-wall carbon nanotubes, polythionine, and gold nanoparticles

An immunosensor for determination of salbutamol was developed. It based on glass carbon electrode (GCE) modified with a conductive multilayer film comprised of multi-wall carbon nanotubes, polythionine and gold nanoparticles. Salbutamol antibody was immobilized on the surface of the modified GCE which then was blocked with bovine serum albumin (BSA). The stepwise self-assembly process of the immunosensor was studied by cyclic voltammetry. The detection scheme is based on competitive binding of salbutamol to the sensor surface whose differential pulse voltammetric signal decreases after competitive binding of the salbutamol-BSA conjugate and free salbutamol to the salbutamol antibody. The sensor responds to salbutamol in 5 to 150 nM concentration range, with a detection limit of 1 nM. This method was applied to the precise and sensitive determination of salbutamol in spiked feed samples.

Performance of dye-sensitized solar cells with various carbon nanotube counter electrodes

Double-wall carbon nanotubes (DWCNTs), single-wall carbon nanotubes (SWCNTs), and multi-wall carbon nanotubes (MWCNTs) were investigated as an alternative for platinum in counter-electrodes for dye-sensitized solar cells. The counter-electrodes were prepared on fluorine-doped tin oxide glass substrates by the screen printing technique from pastes of carbon nanotubes and organic binder. The solar cells were assembled from carbon nanotubes counter-electrodes and screen printed anodes made from titanium dioxide. The cells produced with DWCNTs, SWCNTs or MWCNTs have overall conversion efficiencies of 8.0%, 7.6% and 7.1%, respectively. Electrochemical impedance spectroscopy measurements revealed that DWCNTs displayed the highest catalytic activity for the reduction of tri-iodide ions. The large surface area and superior chemical stability of the DWCNTs facilitated the electron-transfer kinetics at the interface between counter-electrode and electrolyte and yielded the lowest transfer resistance, thereby improving the photovoltaic activity. A short-term stability test at moderate conditions confirmed the robustness of solar cells based on the use of DWCNTs, SWCNTs or MWCNTs.