Qunfang Li

Ultrasensitive Aptamer-Based Multiplexed Electrochemical Detection by Coupling Distinguishable Signal Tags with Catalytic Recycling of DNase I

This work reports an aptamer-based, disposable, and multiplexed sensing platform for simultaneous electrochemical determination of small molecules, employing adenosine triphosphate (ATP) and cocaine as the model target analytes. The multiplexed sensing strategy is based on target-induced release of distinguishable redox tag-conjugated aptamers from a magnetic graphene platform. The electronic signal of the aptasensors could be further amplified by coupling DNase I with catalytic recycling of self-produced reactants. The assay was based on the change in the current at the various peak potentials in the presence of the corresponding signal tags. Experimental results revealed that the multiplexed electrochemical aptasensor enabled the simultaneous monitoring of ATP and cocaine in a single run with wide working ranges and low detection limits (LODs: 0.1 pM for ATP and 1.5 pM for cocaine). This concept offers promise for rapid, simple, and cost-effective analysis of biological samples.

Magnetic mesoporous organic-inorganic NiCo2O4 hybrid nanomaterials for electrochemical immunosensors.

This study demonstrates a facile and feasible strategy toward the development of advanced electrochemical immunosensors based on chemically functionalized magnetic mesoporous organic-inorganic hybrid nanomaterials, and the preparation, characterization, and measurement of relevant properties of the immunosensor for detection of carcinoembryonic antigen (CEA, as a model analyte) in clinical immunoassays. The as-prepared nanomaterials composed of a magnetic mesoporous NiCo(2)O(4) nanosheet, an interlayer of Nafion/thionine organic molecules and a nanogold layer show good adsorption properties for the attachment of horseradish peroxidase-labeled secondary anti-CEA antibody (HRP-anti-CEA). With a sandwich-type immunoassay format, the functional bionanomaterials present good analytical properties to facilitate and modulate the way it was integrated onto the electrochemical immunosensors, and allows the detection of CEA at a concentration as low as 0.5 pg/mL. Significantly, the immunosensor could be easily regenerated by only using an external magnet without the need of any dissociated reagents. Importantly, the as-synthesized magnetic mesoporous NiCo(2)O(4) nanomaterials could be further extended for detection of other biomarkers or biocompounds.

Gold-silver-graphene hybrid nanosheets-based sensors for sensitive amperometric immunoassay of alpha-fetoprotein using nanogold-enclosed titania nanoparticles as labels.

A new sandwich-type electrochemical immunosensor with enhanced sensitivity was developed for detection of alpha-fetoprotein (AFP, as a model analyte) in biological fluids by using nanogold-enclosed titania nanoparticle (AuTi)-labeled secondary antibody on a gold-silver-graphene hybrid nanosheet (AuAgGP)-functionalized glassy carbon electrode (GCE). The presence of the AuAgGP nanosheets not only enhanced the immobilized amount of biomolecules, but also improved the electrochemical properties of the immunosensor. With the aid of AuTi nanolabels, the electrochemical signal was greatly amplified in comparison with pure nanogold or titania-based labels. Under optimal conditions, the sensitivity and dynamic range of the immunosensor were evaluated by using the labeled horseradish peroxidase on the AuTi as trace and H(2)O(2) as enzyme substrate, and exhibited a wide dynamic range of 0.001-200 ng mL(-1) with a low detection limit (LOD) of 0.5 pg mL(-1) AFP (at 3σ). Both the intra- and inter-assay coefficients of variation were less than 10%. The current of the immunosensor at 13th day was as much as 90% of the initial current. In addition, the methodology was evaluated for 8 positive serum specimens obtained from hepatocarcinoma patients and 19 negative sera, and validated with the commercially available Roche 2010 Electrochemiluminescent (ECL) Automatic Analyzer. No significant differences at the 95% confidence level were encountered between two methods.

Carbon nanotube-based symbiotic coaxial nanocables with nanosilica and nanogold particles as labels for electrochemical immunoassay of carcinoembryonic antigen in biological fluids.

A feasible and practicable amperometric immunoassay strategy for sensitive screening of carcinoembryonic antigen (CEA) in human serum was developed using carbon nanotube (CNT)-based symbiotic coaxial nanocables as labels. To construct such a nanocable, a thin layer of silica nanoparticles was coated on the CNT surface by sonication and sol-gel methods, and then colloidal gold nanoparticles were assembled on the amino-functionalized SiO(2)/CNTs, which were used for the label of horseradish peroxidase-anti-CEA conjugates (HRP-anti-CEA-Au/SiO(2)/CNT). In the presence of analyte CEA, the sandwich-type immunocomplex was formed on an anti-CEA/Au/thionine/Nafion-modified glassy carbon electrode by using HRP-anti-CEA-Au/SiO(2)/CNTs as detection antibodies. To embody the advantages of the protocol, the analytical properties of variously modified electrodes were compared in detail on the basis of different nanolabels. Under optimal conditions, the cathodic peak currents of the electrochemical immunosensor were proportional to the logarithm of CEA concentration over the range from 0.01 to 12 ng mL(-1) in pH 5.5 HAc-NaAc containing 5mM H(2)O(2). At a signal-to-noise ratio of 3, the detection limit (LOD) is 5 pg mL(-1) CEA. Intra- and inter-assay coefficients of variation were below 9.5%. Meanwhile, the selectivity and stability of the immunosensor were acceptable. In addition, the technique was evaluated by spiking CEA standards in pH 7.4 PBS and with 35 clinical serum specimens, receiving excellent accordance with results from commercially available electrochemiluminescent enzyme-linked immunoassay.

Sensitive electrochemical immunoassay of carcinoembryonic antigen with signal dual-amplification using glucose oxidase and an artificial catalase.

A new dual-amplification strategy of electrochemical signal based on the catalytic recycling of the product was developed for the antigen-antibody interaction by glucose oxidase (GOD)- conjugated gold-silver hollow microspheres (AuAgHSs) coupled with an artificial catalase, Prussian blue nanoparticles (PB), on a graphene-based immunosensing platform. The first signal amplification introduced in this study was based on the labeled GOD on the AuAgHSs toward the catalytic oxidation of glucose. The generated H(2)O(2) was catalytically reduced by the immobilized PB on the graphene nanosheets with the second amplification. With a sandwich-type immunoassay format, carcinoembryonic antigen (CEA) was monitored as a model analyte by using the synthesized AuAgHSs as labels in pH 6.0 phosphate buffer containing 10mM glucose. Under optimal conditions, the electrochemical immunosensor exhibited a wide dynamic range of 0.005-50 ng mL(-1) with a low detection limit (LOD) of 1.0 pg mL(-1) CEA (at 3σ). Both the intra- and inter-assay coefficients of variation (CVs) were lower than 10%. The specificity and stability of the immunosensor were acceptable. In addition, the assay was evaluated for clinical serum specimens, and received a good correlation with those obtained by the referenced electrochemiluminescent (ECL).

Magneto-controlled electrochemical immunosensor for direct detection of squamous cell carcinoma antigen by using serum as supporting electrolyte

A new magneto-controlled microfluidic device for direct electrochemical determination of squamous cell carcinoma antigen (SCC-Ag) in serum was designed by using anti-SCC antibody (SCC-Ab)-functionalized magnetic mesoporous nanogold/thionine/NiCo(2)O(4) hybrid nanostructures as immunosensing probes (P(1)-Ab) and horseradish peroxidase-SCC-Ab conjugates-labeled nanogold/graphene nanosheets as signal tags (P(2)-Ab). In the presence of the analyte SCC-Ag, the sandwich immunocomplex was formed between the immunosensing probes and the signal tags. With the aid of an external magnet, the formed immunocomplex was attached to the microfluidic device. The assay was implemented in newborn calf serum (NBCS) containing 2.5 mM H(2)O(2) based on the labeled peroxidase on the P(2)-Ab toward the catalytic reduction of H(2)O(2). Under optimal conditions, the increase in the current was proportional to the concentration of SCC-Ag from 2.5 pg/mL to 15 ng/mL. The detection limit (LOD) was 1.0 pg/mL SCC-Ag at 3s(B). The electrochemical immunoassay displayed an acceptable precision, selectivity and stability. Clinical serum specimens were assayed with the method, and the results were in acceptable agreement with those obtained from the referenced electrochemiluminescent method. Importantly, the method can be suitable for on-line use in the mass production of miniaturized lab-on-a-chip devices and open a new opportunity for protein diagnostics and biosecurity.

An enzyme-free quartz crystal microbalance biosensor for sensitive glucose detection in biological fluids based on glucose/dextran displacement approach

A sensitive and facile quartz crystal microbalance (QCM) biosensor for glucose detection in biological fluids was developed by means of a displacement-type assay mode between glucose and its analogy dextran for concanavalin A (ConA) binding sites on a graphene-based sensing platform. To construct such a displacement-based sensor, phenoxy-derived dextran (DexP) molecules were initially assembled onto the surface of graphene-coated QCM probe via π-π stacking interaction, and ConA molecules were then immobilized on the dextran through the dextran-ConA interaction. Upon addition of glucose, the analyte competed with the dextran for the ConA, and displaced it from the QCM probe, leading to a change in the frequency. Under optimal conditions, the frequency change relative to the basic resonant frequency was proportional to glucose concentration, and exhibited a dynamic range from 0.01 to 7.5 mM with a low detection limit (LOD) of 5.0 μM glucose (at 3σ). The relative standard deviations (RSDs) were below 6.2% and 9.0% for the reproducibility and selectivity of the QCM glucose sensors, respectively. In addition, the assay system was evaluated with glucose spiking samples into the distilled water and blank cattle serum, receiving in excellent correlation with the referenced values.

An organic–inorganic hybrid nanostructure-functionalized electrode for electrochemical immunoassay of biomarker by using magnetic bionanolabels

A new electrochemical immunoassay of alpha-fetoprotein (AFP) was developed on an organic-inorganic hybrid nanostructure-functionalized carbon electrode by coupling with magnetic bionanolabels. Multi-walled carbon nanotubes (CNTs), single-stranded DNA, thionine and AFP were utilized for the construction of the immunosensor, while the core-shell Fe(3)O(4)-silver nanocomposites were employed for the label of horseradish peroxidase-anti-AFP conjugates (HRP-anti-AFP-AgFe). Electrochemical measurement toward AFP was carried out by using magnetic bionanolabels as traces and H(2)O(2) as enzyme substrate with a competitive-type immunoassay mode. Experimental results indicated that the immunosensors with carbon nanotubes and DNA exhibited better electrochemical responses than those of without carbon nanotubes or DNA. Under optimal conditions, the electrochemical immunosensor by using HRP-anti-AFP-AgFe as signal antibodies exhibited a linear range of 0.001-200 ng mL(-1) AFP with a low detection limit of 0.5 pg mL(-1) at 3s(B). Both intra- and inter-assay coefficients of variation were 7.3%, 9.4%, 8.7% and 10.2%, 7.8%, 9.4% toward 0.01, 30, 120 ng mL(-1) AFP, respectively. The specificity and stability of the electrochemical immunoassay were acceptable. In addition, the methodology was validated for 12 clinical serum specimens including 9 positive specimens and 3 normal specimens, receiving a good correlation with the results obtained from the referenced electrochemiluminescence assay.

Target-induced biomolecular release for sensitive aptamer-based electrochemical detection of small molecules from magnetic graphene

A novel magneto-controlled electrochemical sensing strategy for simple, sensitive and rapid determination of small molecules was designed by using target-induced release of ferrocene-labeled aptamers from a magnetic graphene platform.

Specific immunoreaction-induced controlled release strategy for sensitive impedance immunoassay of a cancer marker.

A simple and facile impedance immunoassay strategy for sensitive detection of alpha-fetoprotein (AFP), as a model cancer marker, was developed by using target-induced release of nanogold particle-labelled anti-AFP antibodies from polyvinylpyrrolidone-coated magnetic carbon nanotubes.