Yuling Cui

Electrochemical immunosensor for carcinoembryonic antigen based on nanosilver-coated magnetic beads and gold-graphene nanolabels.

A novel redox-active magnetic nanostructure was synthesized by using a wet chemical method for high-efficiency electrochemical immunoassay of carcinoembryonic antigen (CEA, as a model analyte). The nanostructures based on the combination of a magnetic nanocore, a layer of electroactive poly(o-phenylenediamine) (PPD), and a silver metallic shell displayed good adsorption properties for the attachment of anti-CEA antibody selective to CEA. The magnetic nanostructure presented good redox behaviors to facilitate and modulate the way it was integrated into a magnetic carbon paste electrode. The assay was based on a sandwich-type immunoassay protocol by using nanogold-patterned graphene oxide nanoscales (AuNP-GO), conjugated with horseradish peroxidase-labeled anti-CEA, as secondary antibodies and biofunctionalized magnetic nanostructures as immunosensing probes. Under optimal conditions, the nanoparticle-based immunocomposites exhibited good electrochemical responses for the determination of CEA, and allowed the detection of CEA at a concentration as low as 1.0 pg mL(-1) at a signal-to-noise ratio of 3. In addition, the magnetic immunosensing had good reproducibility, and acceptable accuracy, and could be successfully applied for the detection of CEA in the clinical serum specimens. Significantly, by controlling the target biomolecules, this assay can be easily extended for use with other immunosensings, and thus represents a versatile design routine.

Multifunctional gold-silica nanostructures for ultrasensitive electrochemical immunoassay of streptomycin residues.

A facile and simple electrochemical immunoassay for ultrasensitive determination of streptomycin residues (STR) in food was designed by using nanogold-assembled mesoporous silica (GMSNs) as bionanolabels on a three-dimensional redox-active organosilica-functionalized sensing interface. To construct such a sensing interface, we initially synthesized organosilica colloids by using wet chemical method, and then utilized the prepared colloidal organosilica nanocomposites for the immobilization of monoclonal anti-STR antibodies on a glassy carbon electrode based on a sol-gel method. The bionanolabels were prepared based on coimmobilization of horseradish peroxidase (HRP) and STR-bovine serum albumin conjugates (STR-BSA) on the GMSNs. With a competitive-type immunoassay format, the assay toward STR analyte was carried out in pH 5.5 acetate acid buffer (ABS) by using redox-active organosilica nanocomposites as electron mediators, biofunctionalized GMSNs as traces, and hydrogen peroxide (H(2)O(2)) as enzyme substrate. Under optimal conditions, the reduction current of the electrochemical immunosensor decreased with the increase in STR level in the sample, and displayed a wide dynamic range of 0.05-50 ng mL(-1) with a low detection limit (LOD) of 5 pg mL(-1) at 3s(B). Intra- and interassay coefficients of variation were less than 8.7 and 9.3% for STR detection, respectively. In addition, the methodology was validated with STR spiked samples including honey, milk, kidney, and muscle, receiving a good correspondence with the results obtained from high-performance liquid chromatography (HPLC).

Graphene oxide-labeled sandwich-type impedimetric immunoassay with sensitive enhancement based on enzymatic 4-chloro-1-naphthol oxidation

A new sandwich-type impedimetric immunosensor based on functionalized graphene oxide nanosheets with a high ratio of horseradish peroxidase (HRP) and detection antibody was developed for the detection of carcinoembryonic antigen (CEA) by coupling with enzymatic biocatalytic precipitation of 4-chloro-1-naphthol (4-CN) on the captured antibody-modified glassy carbon electrode. Two molecular tags (with and without the graphene oxide nanosheets) were investigated for the detection of CEA and improved analytical features were acquired with the graphene-based labeling. With the labeling method, the performance and factors influencing the properties of the impedimetric immunosensors were also studied and evaluated. Under the optimal conditions, the dynamic concentration range of the impedimetric immunosensors spanned from 1.0pgmL(-1) to 80ngmL(-1) CEA with a detection limit (LOD) of 0.64pgmL(-1). Intra- and inter-assay coefficients of variation were less than 7.5% and 11%, respectively. Additionally, the methodology was evaluated for CEA analysis of 10 clinical serum samples and 5 diluted serum samples, receiving in a good accordance with the results obtained by the impedimetric immunoassay and the commercialized electrochemiluminescent method.

GoldMag nanocomposite-functionalized graphene sensing platform for one-step electrochemical immunoassay of alpha-fetoprotein

A new flow-through electrochemical immunosensor was designed for sensitive detection of alpha-fetoprotein (AFP) in human serum by using nanogold-functionalized magnetic graphene nanosheets as immunosensing probes. Initially, amino functionalized magnetic beads were covalently immobilized on the surface of graphene oxide nanosheets (MGPs), then nanogold particles were adsorbed on the amino groups of the MGPs to construct GoldMag nanocomposites functionalized graphene nanosheets (GMGPs), and then horseradish peroxidase-anti-AFP conjugates (HRP-anti-AFP) were assembled onto the surface of nanogold particles (bio-GMGP). With the aid of an external magnet, the formed bio-GMGPs were attached onto the base electrode in the flow system. With a non-competitive immunoassay format, the injected sample containing AFP antigens was produced transparent immunoaffinity reaction with the immobilized HRP-anti-AFP on the bio-GMGPs. The formed immunocomplex inhibited partly the active center of HRP, and decreased the labeled HRP toward the reduction of H(2)O(2). The performance and factors influencing the performance of the immunosensor were investigated in detail. Under optimal conditions, the electrochemical immunosensor displayed a wide working range of 0.01-200 ng mL(-1) with a low detection limit (LOD) of 1.0 pg mL(-1) AFP (at 3s(B)). Intra- and inter-assay coefficients of variation (CV) were below 10%. In addition, the methodology was validated with real serum samples, receiving a good correlation with the results obtained from commercially available electrochemiluminescence automated analyzer.

Nanogold-functionalized magnetic beads with redox activity for sensitive electrochemical immunoassay of thyroid-stimulating hormone.

A new electrochemical immunosensor for sensitive determination of thyroid-stimulating hormone (TSH) was designed by using redox-active nanogold-functionalized magnetic beads (GoldMag) as signal tags on the nanogold-graphene interface. To construct such GoldMag nanostructures, polyethyleneimine-functionalized magnetic beads (PEI-MBs) were initially prepared by using a wet chemical method, and the electroactive thionine molecules and gold nanoparticles were then alternately immobilized on the surface of PEI-MBs by using an opposite-charged adsorption technique and an in situ synthesis method, respectively. The synthesized GoldMag nanostructures were utilized as signal tags for the label of horseradish peroxidase-anti-TSH conjugates (HRP-anti-TSH). With a sandwich-type immunoassay format, the conjugated signal tags on the transducer were increased with the increasing TSH concentration in the sample, thus enhancing the signal of the electrochemical immunosensor due to the labeled HRP toward the catalytic reduction of H(2)O(2). Under optimal conditions, the current was proportional to the logarithm of TSH concentration ranging from 0.01 to 20 μIU mL(-1) in pH 6.0 HAc-NaAc containing 6 mM H(2)O(2). The detection limit (LOD) was 0.005 μIU mL(-1) TSH at 3s(B). The immunosensor displayed an acceptable reproducibility, stability and selectivity. In addition, the methodology was evaluated with human serum specimens, receiving good correlation with results from commercially available electrochemiluminescent analyzer.

Nanogold-enhanced graphene nanosheets as multienzyme assembly for sensitive detection of low-abundanceproteins

A new electrochemical immunosensing protocol was designed for detection of carcinoembryonic antigen (CEA, as a model protein) by using graphene-carried poly(o-phenylenediamine)/gold hybrid nanosheets (GNPGs) as signal tags on the hierarchical dendritic gold microstructures (HDGMs)-modified glassy carbon electrode. To prepare the signal tags, poly(o-phenylenediamine) molecules were initially immobilized on the surface of graphene nanosheets via the π-stacking interaction. Then gold nanoparticles were assembled onto the poly(o-phenylenediamine)-modified graphene nanosheets, which were used for the labeling of anti-CEA detection antibodies and horseradish peroxidase (HRP). The as-prepared GNPGs were characterized by using atomic force microscopy (AFM), transmission electron microscopy (TEM) and UV-vis absorption spectroscopy. The assay was carried out with a sandwich-type immunoassay format in pH 5.5 acetic acid-buffered saline solutions containing 2.5 mmol L(-1) H2O2. Under optimal conditions, the electrochemical immunoassay exhibited a wide dynamic range of 0.005-80 ng mL(-1) toward CEA standards with a low detection limit of 5.0 pg mL(-1). Intra- and inter-assay coefficients of variation were less than 11.5%. No significant difference at the 0.05 significance level was encountered in the analysis of 6 clinical serum specimens and 6 spiked blank new born cattle serum specimens between the developed immunoassay and commercially available electrochemiluminescent (ECL) method for the detection of CEA.

Au(III)-assisted core–shell iron oxide@poly(o-phenylenediamine) nanostructures for ultrasensitive electrochemical aptasensors based on DNase I-catalyzed target recycling

A redox-active Au(III)-assisted core-shell iron oxide@poly(o-phenylenediamine) nanostructure was designed as a sensing platform for ultrasensitive electrochemical detection of small molecules (ATP, used as a model here) by coupling with DNase I-catalyzed target recycling.

Poly(o-phenylenediamine)-carried nanogold particles as signal tags for sensitive electrochemical immunoassay of prolactin

A novel class of redox-active molecular tags, poly(o-phenylenediamine)-carried nanogold particles (GPPDs), was first synthesized and functionalized with horseradish peroxidase-anti-prolactin conjugates (HRP-anti-PRL). Thereafter, a specific sandwich-type electrochemical immunoassay was designed for determination of prolactin (PRL) by using GPPD-labeled HRP-anti-PRL conjugates as molecular tags on anti-PRL antibody-modified glassy carbon electrode. Compared with pure gold nanoparticles and poly(o-phenylenediamine) microspheres, the as-prepared GPPDs increased the surface coverage of the nanostructures, and enhanced the immobilization amount of biomolecules. Several labeling protocols compromising GPPD-labeled HRP-anti-PRL, nanogold particles-labeled HRP-anti-PRL and poly(o-phenylenediamine) microspheres-labeled HRP-anti-PRL, were investigated for detection of PRL, and improved analytical features were obtained with the GPPD-based strategy. With the GPPD labeling method, dependence of the electrochemical signals on the incubation time and pH of the assay solution were also studied. The strong attachment of HRP-anti-PRL to the GPPDs resulted in a good repeatability and intermediate reproducibility down to 9.8%. The dynamic concentration range spanned from 0.5 to 180 ng mL(-1) PRL with a detection limit of 0.1 ng mL(-1) at the 3S(blank) level. No significant differences at the 95% confidence level were encountered in the analysis of 10 spiked blank cattle serum samples between the developed immunoassay and enzyme-linked immunosorbent assay method for determination of PRL.

Multiarmed star-like platinum nanowires with multienzyme assembly for direct electronic determination of carcinoembryoninc antigen in serum.

A new electrochemical immunoassay strategy for direct detection of carcinoembryoninc antigen (CEA) in serum was developed by using multiarmed star-like platinum nanowires (PtNWs) with biomolecular assembly as signal tags on an anti-CEA-functionalized graphene sensing platform. Initially, the PtNWs were synthesized via a wet chemical method, and then the synthesized PtNWs were used for the co-immobilization of CEA and horseradish peroxidase (HRP). Compared with platinum nanoparticles, the prepared PtNWs could provide a large room for the conjugation of HRP and CEA. With a competitive-type immunoassay format, the assay was performed in two types of supporting electrolytes including new born cattle serum (NBCS) and acetate buffer solution (ABS, pH 5.5), respectively. Similar detection limit (LOD) of 5.0 pg mL(-1) vs. 1.0 pg mL(-1) but narrower dynamic working linear range of 0.01-60 ng mL(-1) vs. 0.002-80 ng mL(-1) was obtained toward CEA standards in the NBCS compared to the ABS. The intra-assay coefficients of variation (CVs) were 4.3%, 8.6%, and 6.2% at 0.05, 10, and 40 ng mL(-1) CEA, respectively, while the inter-assay CVs were 7.6%, 10.5%, and 8.9% at the above-mentioned levels, respectively. In addition, the selectivity and stability of the electrochemical immunosensor were acceptable. Importantly, the developed method was used to assay clinical serum specimens, receiving a good relation with those obtained from the referenced method.

Nanogold–polyaniline–nanogold microspheres-functionalized molecular tags for sensitive electrochemical immunoassay of thyroid-stimulating hormone

Methods based on nanomaterial labels have been developed for electrochemical immunosensors and immunoassays, but most involved low sensitivity. Herein a novel class of molecular tags, nanogold-polyaniline-nanogold microspheres (GPGs), was first synthesized and functionalized with horseradish peroxidase-conjugated thyroid-stimulating hormone antibody (HRP-Ab(2)) for sensitive electrochemical immunoassay of thyroid-stimulating hormone (TSH). X-ray diffraction, confocal Raman spectroscopy, scanning electron microscope and transmission electron microscope were employed to characterize the prepared GPGs. Based on a sandwich-type immunoassay format, the assay was performed in pH 5.0 acetate buffer containing 6.0mmolL(-1) H(2)O(2) by using GPG-labeled HRP-Ab(2) as molecular tags. Compared with pure polyaniline nanospheres and gold nanoparticles alone, the GPG hybrid nanostructures increased the surface area of the nanomaterials, and enhanced the immobilized amount of HRP-Ab(2). Several labeling protocols comprising HRP-Ab(2), nanogold particle-labeled HRP-Ab(2), and polyaniline nanospheres-labeled HRP-Ab(2), were also investigated for determination of TSH and improved analytical features were obtained by using the GPG-labeled HRP-Ab(2). With the GPG labeling method, the effects of incubation time and pH of acetate buffer on the current responses of the immunosensors were also studied. The strong attachment of HRP-Ab(2) to the GPGs resulted in a good repeatability and intermediate precision down to 7%. The dynamic concentration range spanned from 0.01 to 20μIUmL(-1) with a detection limit (LOD) of 0.005μIUmL(-1) TSH at the 3s(B) criterion. Significantly, no significant differences at the 0.05 significance level were encountered in the analysis of 15 spiking serum samples between the developed electrochemical immunoassay and the commercially available enzyme-linked immunosorbent assay (ELISA) method for determination of TSH.