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In this work, titanate nanotubes (TNTs), polyaniline (PANI) and gold nanoparticles (GNPs) were assembled to form a ternary composite, which was then applied on an electrode as a scaffold of an electrochemical enzyme biosensor. The scaffold was constructed by oxidatively polymerising aniline to produce an emeraldine salt of PANI on TNTs, followed by gold nanoparticle deposition. A novel aspect of this scaffold lies in the use of the emeraldine salt of PANI as a molecular wire between TNTs and GNPs. Using horseradish peroxidase (HRP) as a model enzyme, voltammetric results demonstrated that direct electron transfer of HRP was achieved at both TNT-PANI and TNT-PANI-GNP-modified electrodes. More significantly, the catalytic reduction current of H2O2 by HRP was ∼75% enhanced at the TNT-PANI-GNP-modified electrode, compared to that at the TNT-PANI-modified electrode. The heterogeneous electron transfer rate constant of HRP was found to be ∼3 times larger at the TNT-PANI-GNP-modified electrode than that at the TNT-PANI-modified electrode. Based on chronoamperometric detection of H2O2, a linear range from 1 to 1200 μM, a sensitivity of 22.7 μA mM(-1) and a detection limit of 0.13 μM were obtained at the TNT-PANI-GNP-modified electrode. The performance of the biosensor can be ascribed to the superior synergistic properties of the ternary composite., (Copyright © 2016 Elsevier B.V. All rights reserved.)

In this work, a highly sensitive and stable sensing scaffold consisting of gold nanoparticle-encapsulated TiO2 nanotubes, the hydrophilic ionic liquid, 1-decyl-3-methylimidazolium bromide, and Nafion was developed for the fabrication of electrochemical enzyme biosensors. A significant aspect of our work is the application of 12-phosphotungstic acid as both a highly localized photoactive reducing agent to deposit well-dispersed gold nanoparticles on TiO2 nanotubes and an electron mediator to accelerate the electron transfer between an enzyme and the electrode. After characterizing the nanocomposite component of the scaffold by Fourier transform infrared spectroscopy, X-ray diffraction and transmission electron microscopy, thiolated horseradish peroxidase (as a model enzyme) was immobilized on the scaffold and the biosensor was applied to the detection of H2O2. The direct electron transfer between the enzyme and the electrode was promoted by the excellent biocompatibility and conductivity of the scaffold. In addition, a thiolated enzyme has significantly improved the stability and direct electron transfer of horseradish peroxidase on the biosensor, which could be ascribed to the strong affinity between the sulfhydryl group on the enzyme and gold nanoparticles on the biosensor surface. Cyclic voltammetry, chronoamperometry, and square wave voltammetry were used to study the electrochemistry and analytical performance of the biosensor. A dynamic range from 65 to 1600 μM, a limit of detection of 5 μM, and a sensitivity of (18.1 ± 0.43) × 10(-3) μA μM(-1) H2O2 were obtained. The sensing scaffold based on the nanocomposite was demonstrated to be effective and promising in developing enzyme biosensors.

An ultrasensitive electrochemical immmunosensor was demonstrated to be capable of detecting the hormone cortisol down to concentrations as low as 16 pg mL(-1). In addition, the immunosensor displayed a sensitivity of 1.6 μA pg(-1) mL(-1) and a linear range up to ∼2500 pg mL(-1) of cortisol. This immunosensor was constructed based on a Au nanoparticle|dimethyl 3,3'-dithiobispropionimidate·2HCl (DTBP)-Protein G scaffold-modified Au electrode. In this work, the Au nanoparticles were used to increase the electrochemically active surface area by 28% (with a standard deviation of 3%) to enhance the quantity of the Protein G scaffold on the electrode. Thiolation of Protein G by DTBP aided in avoiding the confirmation change of Protein G, while this Protein G-DTBP component offered an orientation-controlled immobilisation of the capture antibody on the Au electrode. In this immunosensor, a monoclonal anti-cortisol capture antibody was optimally aligned by the scaffold before a competitive immunoassay between sample cortisol and a horseradish peroxidase-labelled cortisol conjugate was conducted. For quantitative analysis, square wave voltammetry was used to monitor the reduction current of benzoquinone produced from a horseradish peroxidase catalysed reaction. The improved analytical performance of our immunosensor was attributed to the synergetic effect of Au nanoparticles and the Protein G-DTBP scaffold.

Low picograms of the hormone 17beta-estradiol were detected at an electrochemical immunosensor. This immunosensor features a gold nanoparticle|Protein G-(LC-SPDP)(1)-scaffold, to which a monoclonal anti-estradiol capture antibody was immobilised to facilitate a competitive immunoassay between sample 17beta-estradiol and a horseradish peroxidase-labelled 17beta-estradiol conjugate. Upon constructing this molecular architecture on a disposable gold electrode in a flow cell, amperometry was conducted to monitor the reduction current of benzoquinone produced from a catalytic reaction of horseradish peroxidase. This current was then quantitatively related to 17beta-estradiol present in a sample. Calibration of immunosensors in blood serum samples spiked with 17beta-estradiol yielded a linear response up to approximately 1200 pg mL(-1), a sensitivity of 0.61microA/pg mL(-1) and a detection limit of 6 pg mL(-1). We attribute these favourable characteristics of the immunosensors to the gold nanoparticle|Protein G-(LC-SPDP) scaffold, where the gold nanoparticles provided a large electrochemically active surface area that permits immobilisation of an enhanced quantity of all components of the molecular architecture, while the Protein G-(LC-SPDP) component aided in not only reducing steric hindrance when Protein G binds to the capture antibody, but also providing an orientation-controlled immobilisation of the capture antibody. Coupled with amperometric detection in a flow system, the immunosensor exhibited excellent reproducibility.

In this study, our goal was to produce a self-assembled layer on a gold electrode that would enable the capture of antibodies orientated for maximum binding to their specific antigen in an immunosensor. To achieve this, the amine groups from lysine residues in protein G were initially converted to thiol groups with 2-iminothiolane. The high affinity of thiols for a gold surface facilitates the direct formation of a self-assembled protein G layer. Following this, the coated gold electrode was exposed to a solution of capture antibody (mAb1) so that these antibodies could attach to the protein G layer through their nonantigenic regions, leaving antigen binding sites available with minimal steric hindrance for binding of target analyte. A comparative study between this method and the more conventional strategy of covalently attaching a layer of nonthiolated protein G on an alkanethiol self-assembled monolayer-coated gold electrode has been performed. Based on a reduced preparation time, and an enhanced capacity for immobilized capture antibody to bind its target analyte due to a more favorable orientation, the layer of thiolated protein G was found to be a more suitable backbone for an electrochemical immunosensor. [ABSTRACT FROM AUTHOR]