In this work, we have investigated the in situ growth of Ag nanochains (AgNCs) on carboxyl-functionalized graphene oxide based on enzymatic metalization for the first time. The merit of this method is that the AgNCs could be generated directly on carboxyl-functionalized graphene oxide in a natural, environmentally friendly and sustainable way with high productivity. Furthermore, the Pt nanoparticles were wrapped around the AgNCs (Pt@AgNCs) which manifested some superior functions, such as excellent electrical conductivity and well-defined electrochemical redox activity. With Pt@AgNCs as the electrochemical redox probe, we developed a label-free electrochemical aptasensor for highly sensitive detection of platelet-derived growth factor. A linear response was obtained over a wide range from 6 pM to 40 nM with a detection limit of 1.97 pM.

In the present study, a gold nanoparticle-modified gold electrode (nanogold electrode) was used to develop a novel fluorescein electrochemical DNA biosensor based on a target-induced conformational change. The nanogold electrode was obtained by electrodepositing gold nanoparticles onto a bare gold electrode. This modification not only immobilized probe oligonucleotides, but also adsorbed fluorescein onto the surface of the gold nanoparticles to form an "arch-like" structure. This article compares the electrochemical signal changes caused by the hybridization of "arch-like" DNA on nanogold electrode and linear DNA on bare gold electrode. The results showed that the adsorption effect of nanogold can enhance the sensitivity of the sensor. The linear range of target ssDNA is from 2.0 × 10(-9)M to 2.0 × 10(-8)M with a correlation coefficient of 0.9956 and detection limit (3σ) of 7.10 × 10(-10)M. Additionally, the specificity and hybridization response of this simple sensor were investigated.

A novel signal amplification strategy of mimicking bi-enzyme synergetic catalysis to generate coreactant in situ was designed to fabricate an ultrasensitive peroxydisulfate electrochemiluminescence (ECL) immunosensor for detection of Streptococcus suis serotype 2 (SS2). It was the first time to detect SS2 by using ECL. Through the interaction between l-cysteine (l-cys) and hollow PtPd bimetal alloy nanoparticles (HPtPd) to form ((l-cys-HPtPd)n) nanocomposites, the loading amount of l-cys and HPtPd was greatly increased, which could greatly enhance the ECL signal of peroxydisulfate. At the same time, Glucose Oxidase (GOD), used to block nonspecific binding sites of (l-cys-HPtPd)n nanocomposites, could rapidly oxidize d-glucose in the detection solution into gluconic acid accompanying with the generation of H2O2, which was further catalyzed by HPtPd to generate O2. And O2, acted as the coreactant of peroxydisulfate, could greatly amplify the ECL signal. In the process, HPtPd could be regarded as mimicking enzyme, the effect of which was similar to horseradish peroxidase (HRP) in generating O2. With the several amplification factors of a sandwich-type structure we designed, a wide linear ranged from 0.0001 to 100ngmL−1 was acquired with a relatively low detection limit of 33fgmL−1 for SS2. The present work demonstrated that the novel strategy had the great advantages in sensitivity, selectivity and reproducibility which might hold a new promise for highly sensitive bioassays applied in clinical detection.

Herein, a strategy was put forward for sensitive label-free electrochemical immunosensor by using silver–graphene oxide (Ag–GO) as redox probe. Initially, a simple, soft reaction conditions and efficient route was designed to in situ reduce silver nanoparticles (AgNPs) onto graphene oxide (GO) by glucose as reducing agent. The obtained nanocomposite gave a pair of well-defined redox peaks with the advantages of good biocompatibility and chemical stability. Furthermore, the GO provided large surface area for assembly of abundant AgNPs with redox activity, which potentially paved sensitive way for immunodetection. Thus, the Ag–GO nanocomposites were used as redox probe to fabricate a sensitive label-free immunosensor for α-1-fetoprotein (AFP). In addition, gold nanoparticles (AuNPs) for immobilization of antibody (anti-AFP) could amplify the electrochemical signal and further enhance the sensitivity of the immunosensor due to its excellent conductivity and large surface area. Under the optimal condition, the anodic peak current linearly responded to the logarithm of AFP concentration in a wide range from 0.01 to 100 ng mL−1 with a low limit of detection of 3 pg mL−1 (S/N = 3). The designed immunosensor displayed the advantages of simple preparation, high sensitivity, good selectivity and satisfactory stability.

In the present study, we fabricated an ultrasensitive sandwich-type electrochemical aptasensor for thrombin (TB) based on a triplex signal amplification strategy. The hemin/G-quadruplex horseradish peroxidase-mimicking DNAzyme (HRP-DNAzyme) as well as blocking reagent-horseradish peroxidase (HRP) and iron telluride nanorods (FeTe NRs) could simultaneously amplify the electrochemical signal of thionine (Thi) in the presence of H2O2. Herein, FeTe NRs, as a newly discovered HRP-mimicking enzyme, were employed to construct an aptasensor for the first time. And, the FeTe NRs decorated by gold nanoparticles (abbreviated as AuNPs@FeTe NRs), were not only used as carriers of secondary thrombin aptamer (TBA 2), electron mediator thionine (Thi) and HRP, but also catalyzed the electrochemical reaction of Thi in the presence of H2O2. As can be seen from experiment results, with the triplex signal amplification strategy, the reduction peak current of the fabricated aptasensor was logarithmically related to the concentration of thrombin (TB) over a wide range from 1 pM to 20 nM, and a detection limit of 0.5 pM was obtained. Hence, the proposed aptamer-based sandwich sensing approach for amplified detection of TB could provide a promising way for highly sensitive determination of other analytes.

In the present study, an ultrasensitive electrochemical aptasensor for the simultaneous detection of thrombin (TB) and ochratoxin A (OTA) was fabricated by using exonuclease-catalyzed target recycling and DNA concatemers for signal amplification. The previously hybridized double-stranded DNAs (SH-cTBA/TBA and SH-cOBA/OBA) were self-assembled on a gold electrode. In the presence of targets, the formation of aptamer-target complexes would lead to not only the dissociation of aptamers (TBA and OBA) from the double-stranded DNAs but also the transformation of the complementary DNAs (SH-cTBA and SH-cOBA) into hairpin structures. Subsequently, owing to employment of RecJf exonuclease, which is a single-stranded DNA-specific exonuclease to selectively digest the appointed DNAs (TBA and OBA), the targets could be liberated from the aptamer-target complexes for recycling of the analytes. Thereafter, probe DNAs (T1 and T2) were employed to hybridize with SH-cTBA and SH-cOBA respectively to provide primers for the concatemer reaction. After that, when four auxiliary DNA strands S1, anthraquinone-2-carboxylic acid (AQ)-labeled S2, S3, S4, as well as hemin were introduced, extended dsDNA polymers with lots of AQ moieties and hemin-G-quadruplex complexes could form on the electrode surface. Then, based on the signal of the AQ and hemin-G-quadruplex complex, an electrochemical aptasensor for the simultaneous detection of TB and OTA was successfully fabricated.

In this work, an advanced sandwich-type electrochemical aptasensor for thrombin was proposed by integrating hemin/G-quadruplex with functionalized graphene-Pd nanoparticles composites (PdNPs-RGs). The hemin/G-quadruplex formed by intercalating hemin into thrombin binding aptamer (TBA), firstly acted as a NADH oxidase, assisting the oxidation of NADH to NAD(+) accompanying with the generation of H(2)O(2) in the presence of dissolved O(2). Subsequently, the hemin/G-quadruplex acted as HRP-mimicking DNAzyme that rapidly bioelectrocatalyze the reduction of the produced H(2)O(2). At the same time, the Pd nanoparticles supported on p-iodoaniline functionalized graphene were also adopted to catalyze the reduction of H(2)O(2). Thus, with the dual catalysis, a dramatically amplified electrochemical signal could be obtained. Besides, the avidin-biotin system for binding aptamer sequences on electrodes not only improved the sensitivity of thrombin analysis but also obtained an acceptable repeatability of the aptasensor. With several factors mentioned above, a wide linear ranged from 0.1 pM to 50 nM was acquired with a relatively low detection limit of 0.03 pM (defined as S/N=3). These excellent performances provided our approach a promising way for ultrasensitive assay in electrochemical aptasensors.

For the first time, a glucose oxidase-functionalized bioconjugate was prepared and served as a new trace label through its direct electrochemistry and electrocatalysis in a sandwich-type electrochemical aptasensor for ultrasensitive detection of thrombin.

In the present study, toluidine blue–graphene (Tb–Gra) nanocomposites were prepared to design a Lable-free electrochemical aptasensor for highly sensitive detection of thrombin based on layer-by-layer (LBL) technology. The nanocomposites with excellent redox electrochemical activities were first immobilized on the gold nanoparticles (nano-Au) modified glassy carbon electrodes (GCE). Then, the LBL structure was performed by electrostatic adsorption between the positively charged Tb–Gra and negatively charged nano-Au, which formed {Tb–Gra/nano-Au}n multilayer films for electroactive species enrichment and biomolecule immobilization. Subsequently, the thiolated thrombin binding aptamer (TBA) was assembled on the nano-Au surface through Au–S bond. In the presence of target thrombin (TB), the TBA on the multilayer could catch the thrombin onto the electrode surface, which resulted in a barrier for electro-transfer, leading to the decrease of the electrochemical signal of Tb–Gra nanocomposites. Under the optimal conditions, a wide detection range from 0.001nM to 80nM and a low detection limit of 0.33 pM (defined as S/N=3) for thrombin were obtained. In addition, the sensor exhibited excellent selectivity against other proteins.

Herein, we successfully fabricated a highly sensitive label-free electrochemical aptasensor for thrombin based on the amplification of graphene (Gra). The excellent electrochemical probe of nickel hexacyanoferrate nanoparticles (NiHCFNPs) was introduced to form Nafion-Graphene-NiHCFNPs (Nf-Gra-NiHCFNPs) nanocomposites membrane on the gold electrode. The employment of graphene not only enhanced the surface area of the electrode with increased NiHCFNPs immobilization, but also improved the conductivity of the electrode, which further effectively improved the sensitivity of this proposed aptasensor. Subsequently, AuNPs layer was formed to immobilize the thrombin aptamer (TBA) and enhance the stability of the composite monolayer mentioned above. Then, thiol-modified TBA was assembled onto the AuNPs layer. Thereafter, hexanethiol (HT) was employed to block the possible remaining active sites. With the dual amplification of Gra and AuNPs, the resulting aptasensor exhibited good current response to target thrombin with a wide linear range extended from 1 pM to 80 nM (the detection limit was 0.3 pM). Additionally, the morphologies of bare Au substrate, nickel hexacyanoferrate nanoparticles (NiHCFNPs) and nanocomposites were successfully characterized by atomic force microscopy (AFM).

A highly specific in situ amplification strategy was designed for ultrasensitive detection of thrombin by combining the layer-by-layer (LBL) assembled amplification with alkaline phosphatase (ALP) and gold nanoparticles (Au) mediated silver deposition. High-density carboxyl functionalized graphene oxide (FGO) was introduced as a nanocarrier for LBL assembling of alkaline phosphatase decorated gold nanoparticles (ALP-Au), which was further adopted to label thrombin aptamer II. After sandwich-type reaction, numerous ALP were captured onto the aptasensor surface and catalyzed the hydrolysis of ascorbic acid 2-phosphate (AAP), which in situ generated ascorbic acid (AA), reducing Ag+ to Ag nanoparticles (AgNPs) for electrochemical readout. Inspiringly, the in situ amplification strategy with ethanolamine as an effective blocking agent showed remarkable amplification efficiency, very little nonspecific adsorption, and low background signal, which was favorable to enhance the sensitivity of aptasensor. Our novel dramatic signal amplification strategy, with a detection limit of 2.7 fM, showed about 2–3 orders of magnitude improvement in the sensitivity for thrombin detection compared to other universal enzyme-based electrochemical assay.

Herein, a novel third-generation glucose biosensor based on unique hollow nanostructured Pt decorated multiwall carbon nanotubes (HPt-CNTs) composites was successfully constructed. The HPt-CNTs composites were successfully prepared and cast on the glassy carbon electrode (GCE) surface directly. With the help of electrostatic adsorption and covalent attachment, the negative l-cysteine (l-cys) and the positive poly(diallydimethylammonium) chloride (PDDA) protected gold nanoparticles (PDDA-Au) were modified on the resulting electrode surface subsequently, which provided further immobilization of glucose oxidase (GOD). Exploitation of the unique properties of HPt-CNTs composites led to the achievement of direct electron transfer between the electrode and the redox active centers of GOD, and the electrode exhibited a pair of well-defined reversible redox peaks with a fast heterogeneous electron transfer rate. In particular, the detection limit (4×10−7M) of this biosensor was significantly lower and the linear range (1.2μM–8.4mM) was much wider than similar carbon nanotubes (CNTs) and Pt-based glucose biosensors. The resulted biosensor also showed high sensitivity and freedom of interference from other co-existing electroactive species, indicating that our facile procedure of immobilizing GOD exhibited better response and had potential application for glucose analysis.

In this work, we have successfully demonstrated a facile strategy to incorporate high-quality hollow CoPt bimetal alloy nanoparticles (HCoPt) onto reduced graphene oxide sheet (HCoPt-RGs). An advanced sandwich-type electrochemical aptasensor for thrombin was proposed by using the HCoPt-RGs conjugates as secondary label. The formed conjugates provided large surface area for loading plentiful redox probe thionine (Thi), horseradish peroxidase (HRP) and secondary aptamer (Apt II) with good stability and friendly biocompatibility, indicating their superior properties in electroactive mediator enrichment and biomolecule immobilization. Furthermore, activated by glutaraldehyde (GA), the chitosan-hollow CoPt alloy nanoparticle (CS-HCoPt) film can greatly facilitate the capture of primary aptamer (Apt I) and dramatically reduce the nonspecific binding. Excellent sensitivity was obtained by detecting the conspicuously enhanced electrochemical signal of Thi, which was amplified by HCoPt alloy nanoparticles and HRP toward the catalytic reduction of H2O2. The aptasensor displayed excellent performance for thrombin with a wide linearity in the range from 1.0×10−12 to 5.0×10−8M and a relatively low detection limit of 3.4×10−13M. Moreover, the resulted aptasensor also exhibited good specificity, acceptable reproducibility and stability, indicating that the present strategy could pave a promising way for the wide application of graphene in clinical research.

In this work, a sandwich-type electrochemical aptasensor for simultaneous sensitive detection of platelet-derived growth factor (PDGF) and thrombin is fabricated. Reduced graphene oxide sheets (rGS) are used as matrices to immobilize the redox probes, which are subsequently coated with platinum nanoparticles (PtNPs) to form the PtNPs-redox probes-rGS nanocomposites. With the employment of the as prepared nanocomposites, a signal amplification strategy was described based on bienzyme (glucose oxidase and horseradish peroxidase) modified PtNPs-redox probes-rGS nanocomposites as the tracer labels for secondary aptamers (Apt II) through sandwiched assay. Gold nanoparticles functionalized single-walled carbon nanotubes (AuNPs@SWCNTs) as the biosensor platform enhance the surface area to capture a large amount of primary aptamers (Apt I), thus amplifying the detection response. The experiment results show that the multi-labeled PtNPs-redox probes-rGS nanocomposites display satisfying electrochemical redox activity and highly electrocatalytic activity of PtNPs and bienzyme, which exhibit high sensitivity for detection of proteins. The linear range of PDGF is 0.01-35 nM with a detection limit of 8 pM, while the linear ranges from 0.02 to 45 nM and a detection limit of 11 pM for thrombin are obtained. (C) 2011 Elsevier Ltd. All rights reserved.

Highly sensitive electrochemical platform based on Pt nanoparticles supported on carbon nanotubes (Ptnano-CNTs) and sugar–lectin biospecific interactions is developed for the direct electrochemistry of glucose oxidase (GOD). Firstly, Ptnano-CNTs nanocomposites were prepared in the presence of carbon nanotubes (CNTs), and then the mixture was cast on a glassy carbon electrode (GCE) using chitosan as a binder. Thereafter, concanavalin A (Con A) was adsorbed onto the precursor film by the electrostatic force between positively charged chitosan and the negatively charged Con A. Finally, the multilayers of Con A/GOD films were prepared based on biospecific affinity of Con A and GOD via layer-by-layer (LBL) self-assembly technique. The electrochemical behavior of the sensor was studied using cyclic voltammetry and chronoamperometry. The electrochemical parameters of GOD in the film were calculated with the results of the electron transfer coefficient (α) and the apparent heterogeneous electron transfer rate constant (ks) as 0.5 and 5.093 s−1, respectively. Experimental results show that the biosensor responded linearly to glucose in the range from 1.2 × 10−6 to 2.0 × 10−3 M, with a detection limit of 4.0 × 10−7 M under optimized conditions.

A novel tracer, platinum–gold alloy nanoparticles (Pt–AuNPs) and horseradish peroxidase (HRP) functionalized single-walled carbon nanotubes (SWCNTs) composite, is employed to label the secondary thrombin aptamer for constructing an ultrasensitive electrochemical aptasensor. Thionine, immobilized on functionalized SWCNTs, provides a pair of distinguished redox peak for electrochemical detection. Both the high-content Pt–AuNPs and HRP on SWCNTs amplify the electrochemical signal of thionine through electrocatalytic reduction of H2O2. Differential pulse voltammetry (DPV) is employed to detect thrombin with different concentrations. The reduction peak current is logarithmically related to the concentration of thrombin in an extremely wide range from 10fM to 5nM with a detection limit of 3.6fM. The dual signal amplification of Pt–AuNPs and HRP functionalized nanocomposite provides a promising way for ultrasensitive assay in electrochemical aptasensors.