Zuanguang Chen

Direct electron transfer of glucose oxidase and biosensing for glucose based on PDDA-capped gold nanoparticle modified graphene/multi-walled carbon nanotubes electrode

In this work, poly (diallyldimethylammonium chloride) (PDDA)-capped gold nanoparticles (AuNPs) functionalized graphene (G)/multi-walled carbon nanotubes (MWCNTs) nanocomposites were fabricated. Based on the electrostatic attraction, the G/MWCNTs hybrid material can be decorated with AuNPs uniformly and densely. The new hierarchical nanostructure can provide a larger surface area and a more favorable microenvironment for electron transfer. The AuNPs/G/MWCNTs nanocomposite was used as a novel immobilization platform for glucose oxidase (GOD). Direct electron transfer (DET) was achieved between GOD and the electrode. Field emission scanning electron microscopy (FESEM), UV-vis spectroscopy and cyclic voltammetry (CV) were used to characterize the electrochemical biosensor. The glucose biosensor fabricated based on GOD electrode modified with AuNPs/G/MWCNTs demonstrated satisfactory analytical performance with high sensitivity (29.72mAM(-1)cm(-2)) and low limit of detection (4.8 µM). The heterogeneous electron transfer rate constant (ΚS) and the apparent Michaelis-Menten constant (Km) of GOD were calculated to be 11.18s(-1) and 2.09 mM, respectively. With satisfactory selectivity, reproducibility, and stability, the nanostructure we proposed offered an alternative for electrode fabricating and glucose biosensing.

Ultrasensitive electrochemical detection of microRNA based on an arched probe mediated isothermal exponential amplification.

In this work, a simple and label-free electrochemical biosensor is developed for microRNA (miRNA) detection on the basis of an arched probe mediated isothermal exponential amplification reaction (EXPAR). The arched probe assembled on the electrode surface consists of two strands that are partially complementary to each other at both ends. The target can hybridize with the complementary sequence of the arched structure, leading to the cleavage of the probe. The strand fixed on the surface of the electrode self-assembles, in the presence of hemin, to G-quadruplex unit, yielding electrochemical signals. The other strand liberated into the solution triggers the EXPAR to recycle and regenerate targets. This method exhibits ultrahigh sensitivity toward miRNA with detection limits of 5.36 fM and a detection range of 3 orders of magnitude. The biosensor is capable of discriminating a single-nucleotide difference between concomitant miRNA and performs well in analyzing crude extractions from cancer cell lines.

Sensitive electrochemical aptamer cytosensor for highly specific detection of cancer cells based on the hybrid nanoelectrocatalysts and enzyme for signal amplification

Human cancer is becoming a leading cause of death in the world and the development of a straightforward strategy for early detection of cancer is urgently required. Herein, a sandwich-type electrochemical aptamer cytosensor was developed for detection of human liver hepatocellular carcinoma cells (HepG2) based on the hybrid nanoelectrocatalysts and enzyme for signal amplification. The thiolated TLS11a aptamers were used as a selective bio-recognition element, attached to the gold nanoparticles (AuNPs) modified the glassy carbon electrode (GCE) surface. Meanwhile, the electrochemical nanoprobes were fabricated through the G-quadruplex/hemin/aptamer complexes and horseradish peroxidase (HRP) immobilized on the surfaces of Au@Pd core-shell nanoparticle-modified magnetic Fe3O4/MnO2 beads (Fe3O4/MnO2/Au@Pd). After the target cells were captured, the hybrid nanoprobes were further assembled to form an aptamer-cell-nanoprobes sandwich-like system on the electrode surface. Then, hybrid Fe3O4/MnO2/Au@Pd nanoelectrocatalysts, G-quadruplex/hemin HRP-mimicking DNAzymes and the natural HRP enzyme efficiently catalyzed the oxidation of hydroquinone (HQ) with H2O2, amplifying the electrochemical signals and improving the detection sensitivity. This electrochemical cytosensor delivered a wide detection range of 1×10(2)-1×10(7)cellsmL(-1), high sensitivity with a low detection limit of 15cellsmL(-1), good selectivity and repeatability. Finally, an electrochemical reductive desorption method was performed to break gold-thiol bond and desorb the components on the AuNPs/GCE for regenerating the cytosensor. These results have demonstrated that the electrochemical cytosensor has the potential to be a feasible tool for cost-effective cancer cell detection in early cancer diagnosis.

Ultrasensitive electrochemical detection of avian influenza A (H7N9) virus DNA based on isothermal exponential amplification coupled with hybridization chain reaction of DNAzyme nanowires

In this work, a simple and label-free electrochemical biosensor with duel amplification strategy was developed for DNA detection based on isothermal exponential amplification (EXPAR) coupled with hybridization chain reaction (HCR) of DNAzymes nanowires. Through rational design, neither the primer nor the DNAzymes containing molecular beacons (MBs) could react with the duplex probe which were fixed on the electrode surface. Once challenged with target, the duplex probe cleaved and triggered the EXPAR mediated target recycle and regeneration circles as well as the HCR process. As a result, a greater amount of targets were generated to cleave the duplex probes. Subsequently, the nanowires consisting of the G-quadruplex units were self-assembled through hybridization with the strand fixed on the electrode surface. In the presence of hemin, the resulting catalytic G-quadruplex-hemin HRP-mimicking DNAzymes were formed. Electrochemical signals can be obtained by measuring the increase in reduction current of oxidized 3.3,5.5-tetramethylbenzidine sulfate (TMB), which was generated by DNAzyme in the presence of H2O2. This method exhibited ultrahigh sensitivity towards avian influenza A (H7N9) virus DNA sequence with detection limits of 9.4 fM and a detection range of 4 orders of magnitude. The biosensor was also capable of discriminating single-nucleotide difference among concomitant DNA sequences and performed well in spiked cell lysates.

A repeatable assembling and disassembling electrochemical aptamer cytosensor for ultrasensitive and highly selective detection of human liver cancer cells.

In this work, a repeatable assembling and disassembling electrochemical aptamer cytosensor was proposed for the sensitive detection of human liver hepatocellular carcinoma cells (HepG2) based on a dual recognition and signal amplification strategy. A high-affinity thiolated TLS11a aptamer, covalently attached to a gold electrode through Au-thiol interactions, was adopted to recognize and capture the target HepG2 cells. Meanwhile, the G-quadruplex/hemin/aptamer and horseradish peroxidase (HRP) modified gold nanoparticles (G-quadruplex/hemin/aptamer-AuNPs-HRP) nanoprobe was designed. It could be used for electrochemical cytosensing with specific recognition and enzymatic signal amplification of HRP and G-quadruplex/hemin HRP-mimicking DNAzyme. With the nanoprobes as recognizing probes, the HepG2 cancer cells were captured to fabricate an aptamer-cell-nanoprobes sandwich-like superstructure on a gold electrode surface. The proposed electrochemical cytosensor delivered a wide detection range from 1×10(2) to 1×10(7) cells mL(-1) and high sensitivity with a low detection limit of 30 cells mL(-1). Furthermore, after the electrochemical detection, the activation potential of -0.9 to -1.7V was performed to break Au-thiol bond and regenerate a bare gold electrode surface, while maintaining the good characteristic of being used repeatedly. The changes of gold electrode behavior after assembling and desorption processes were investigated by electrochemical impedance spectroscopy and cyclic voltammetry techniques. These results indicate that the cytosensor has great potential in disease diagnostic of cancers and opens new insight into the reusable gold electrode with repeatable assembling and disassembling in the electrochemical sensing.

One-step synthesis of potassium ferricyanide-doped polyaniline nanoparticles for label-free immunosensor.

A novel, label-free and inherent electroactive redox immunosensor for ultrasensitive detection of carcinoembryonic antigen (CEA) was proposed based on gold nanoparticles (AuNPs) and potassium ferricyanide-doped polyaniline (FC-PANI) nanoparticles. FC-PANI composite was synthesized via oxidative polymerization of aniline, using potassium ferricyanide (K3[Fe(CN)6]) as both oxidant and dopant. FC-PANI acting as the signal indicator was first fixed on a gold electrode (GE) to be the signal layer. Subsequently, the negatively charged AuNPs could be adsorbed on the positively charged FC-PANI modified GE surface by electrostatic adsorption, and then to immobilize CEA antibody (anti-CEA) for the assay of CEA. The CEA concentration was measured through the decrease of amperometric signals in the corresponding specific binding of antigen and antibody. The wide linear range of the immunosensor was from 1.0 pg mL(-1) to 500.0 ng mL(-1) with a low detection limit of 0.1 pg mL(-1) (S/N=3). The proposed method would have a potential application in clinical immunoassays with the properties of facile procedure, stability, high sensitivity and selectivity.

A novel non-enzymatic hydrogen peroxide sensor based on poly-melamine film modified with platinum nanoparticles

A novel hydrogen peroxide (H2O2) sensor was fabricated using electrodepositing platinum nanoparticles (PtNPs) on a glassy carbon electrode (GCE) modified with a highly stable poly-melamine film. Since the formation of the poly-melamine layer significantly increased the surface area, high-density PtNPs were homogeneously loaded onto the nanostructured matrix of poly-melamine. The PtNPs/poly-melamine heterostructure-based H2O2 sensor synergized the advantages of both the conducting film and the nanoparticle catalyst. This non-enzymatic sensor exhibited high electrocatalytic activity toward the reduction of H2O2, and provided a linear response for H2O2 in the concentration range from 5 μmol L−1 to 1650 μmol L−1 with a limit of detection (LOD, S/N = 3) of 0.65 μmol L−1. In addition, this proposed H2O2 sensor displayed high sensitivity, good anti-interference ability, excellent reproducibility and long-term stability. The excellent analytical performances and facile preparation process made this novel hybrid electrode promising for the development of H2O2 sensors.

Molecularly imprinted coated graphene oxide solid-phase extraction monolithic capillary column for selective extraction and sensitive determination of phloxine B in coffee bean

A method was developed to sensitively determine phloxine B in coffee bean by molecularly imprinted polymers (MIPs) coated graphene oxide (GO) solid-phase extraction (GO-MISPE) coupled with high-performance liquid chromatography and laser-induced fluorescence detection (HPLC-LIF). The GO-MISPE capillary monolithic column was prepared by water-bath in situ polymerization, using GO as supporting material, phloxine B, methacrylic acid (MAA), and ethylene dimethacrylate (EDMA) as template, functional monomer, and cross-linker, respectively. The properties of the homemade GO-MISPE capillary monolithic column, including capacity and specificity, were investigated under optimized conditions. The GO-MIPs were characterized by scanning electron microscopy (SEM) and Fourier transform-infrared spectroscopy (FT-IR). The mean recoveries of phloxine B in coffee bean ranged from 89.5% to 91.4% and the intra-day and inter-day relative standard deviation (RSD) values all ranged from 3.6% to 4.7%. Good linearity was obtained over 0.001-2.0 μg mL(-1) (r=0.9995) with the detection limit (S/N=3) of 0.075 ng mL(-1). Under the selected conditions, enrichment factors of over 90-fold were obtained and extraction on the monolithic column effectively cleaned up the coffee bean matrix. The results demonstrated that the proposed GO-MISPE HPLC-LIF method can be applied to sensitively determine phloxine B in coffee bean.

Zebrafish on a Chip: A Novel Platform for Real-Time Monitoring of Drug-Induced Developmental Toxicity

Pharmaceutical safety testing requires a cheap, fast and highly efficient platform for real-time evaluation of drug toxicity and secondary effects. In this study, we have developed a microfluidic system for phenotype-based evaluation of toxic and teratogenic effects of drugs using zebrafish (Danio rerio) embryos and larvae as the model organism. The microfluidic chip is composed of two independent functional units, enabling the assessment of zebrafish embryos and larvae. Each unit consists of a fluidic concentration gradient generator and a row of seven culture chambers to accommodate zebrafish. To test the accuracy of this new chip platform, we examined the toxicity and teratogenicity of an anti-asthmatic agent-aminophylline (Apl) on 210 embryos and 210 larvae (10 individuals per chamber). The effect of Apl on zebrafish embryonic development was quantitatively assessed by recording a series of physiological indicators such as heart rate, survival rate, body length and hatch rate. Most importantly, a new index called clonic convulsion rate, combined with mortality was used to evaluate the toxicities of Apl on zebrafish larvae. We found that Apl can induce deformity and cardiovascular toxicity in both zebrafish embryos and larvae. This microdevice is a multiplexed testing apparatus that allows for the examination of indexes beyond toxicity and teratogenicity at the sub-organ and cellular levels and provides a potentially cost-effective and rapid pharmaceutical safety assessment tool.

A label-free hemin/G-quadruplex DNAzyme biosensor developed on electrochemically modified electrodes for detection of a HBV DNA segment

In this work, a label-free DNAzyme biosensor was proposed for the detection of oligonucleotides related to a hepatitis B virus (HBV) DNA segment. The novel DNAzyme biosensor was based on the electrochemical reduction of graphene oxide–carboxyl multi-walled carbon nanotube composites (GO–CMWCNTs) and the electrodeposition of gold nanoparticles (AuNPs) on a glassy carbon electrode (GCE). The thiolate hairpin capture probes with a caged G-quadruplex configuration were self-assembled on AuNPs through the well-known Au–thiol binding. Only after hybridization with target DNA, the hairpin configuration could be opened and released the particular guanine-rich nucleic acid sequences which could assemble to G-quadruplexes. In the presence of hemin and K+, the hemin/G-quadruplex DNAzyme was generated on the electrode surface, triggering the electrochemical H2O2-mediated oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB). With the advantages of amplification effects of AuNPs and hemin/G-quadruplex, a linear concentration range from 10 pM to 10 nM with a limit of detection of 0.5 pM was obtained for the target DNA. This method exhibited excellent discrimination of target DNA from one or two-base mismatched DNA and non-complementary DNA sequences. In addition, the proposed biosensor was successfully applied to the determination of target DNA in human serum samples.