Observing Mesoscopic Nucleic Acid Capacitance Effect and Mismatch Impact via Graphene Transistors.

This work reports a molecular-scale capacitance effect of the double helical nucleic acid duplex structure for the first time. By quantitatively conducting large sample measurements of the electrostatic field effect using a type of high-accuracy graphene transistor biosensor, an unusual charge-transport behavior is observed in which the end-immobilized nucleic acid duplexes can store a part of ionization electrons like molecular capacitors, other than electric conductors. To elucidate this discovery, a cascaded capacitive network model is proposed as a novel equivalent circuit of nucleic acid duplexes, expanding the point-charge approximation model, by which the partial charge-transport observation is reasonably attributed to an electron-redistribution behavior within the capacitive network. Furthermore, it is experimentally confirmed that base-pair mismatches hinder the charge transport in double helical duplexes, and lead to directly identifiable alterations in electrostatic field effects. The bioelectronic principle of mismatch impact is also self-consistently explained by the newly proposed capacitive network model. The mesoscopic nucleic acid capacitance effect may enable a new kind of label-free nucleic acid analysis tool based on electronic transistor devices. The in situ and real-time nucleic acid detections for virus biomarkers, somatic mutations, and genome editing off-target may thus be predictable.

Gas Sensor Based on 3-D WO3 Inverse Opal: Design and Applications.

A three-dimensional inverse opal (3DIO) WO3 architecture has been synthesized via a simple sacrificial template method. Morphology features of the 3DIO were characterized by scanning electron microscope (SEM) and its structure was characterized by X-ray diffraction (XRD). The shrinking ratio of the PMMA spheres was ~28.2% through measuring the distribution of the PMMA spheres and 3DIO WO3 center-to-center distance between the spheres and macropores, respectively. Beyond that, the 3DIO gas sensing properties were investigated systematically and the sensing mechanism of 3DIO WO3 was proposed. The results indicated that the response of the 3DIO sensor possessed excellent sensitivity to acetone gas, especially at trace levels. The 3DIO gas sensor response was ~7 to 5 ppm of acetone and could detect acetone low to 0.2 ppm effectively, which was in close proximity to the theoretical low detection limit of 0.14 ppm when Ra/Rg ≥ 1.2 was used as the criterion for reliable gas sensing. All in all, the obvious satisfaction of the gas-sensing properties was ascribed to the structure of the 3DIO, and the sensor could be a promising novel device in the future.

A rapid bioassay for recombinant interleukin-22.

Interleukin-22 is a novel cytokine produced mainly in activated T cells. The elaborate biological functions of IL-22 in vivo are still widely unknown. In this report, we describe a rapid, simple, and reproducible in vitro cell-based bioassay for measuring the bioactivity of recombinant interleukin-22 (IL-22) to study the primary function of IL-22 in vivo. Human hepatocyte cell line (HepG2) was transfected with pSTAT3-Luc, a plasmid carrying the luciferase gene under the control of STAT3. After screening and selection, one stable clone was established which generates a strong response to recombinant human IL-22 (rhIL-22) stimulation in a dose-dependent manner. The cell showed ED50 of 17.0 +/- 1.4 ng/mL (n = 15) to recombinant human IL-22. Pre-incubation of anti-IL-22 mAb with IL-22 recombinant proteins completely blocked the bioactivities. The assay can be completed within one day. The current assay provides a rapid analytical method to measure the biological activity of IL-22 in vitro.

[Antioxidants inhibit the oxidative modification of high density lipoproteins].

OBJECTIVE: To study the role and mechanism of antioxidants on inhibiting oxidative modification of high density lipoproteins (HDL). METHODS: Freshly prepared human plasma HDL was treated by incubation with copper ion, hyperchlorite or arterial wall cells. Compared to control, the test groups were treated with addition of different concentration of butylhydroxytoluene (BHT), vitamin C and vitamin E. Then, the relative electrophoretic mobility (REM), thiobarbituric acid-reactive substances (TBARS), ratio of lysolecithin to lecithin (LPC/PC), and lipoprotein moieties were investigated. RESULTS: BHT, vitamin C and vitamin E can significantly inhibit the increasing REM, TBARS, LPC/PC ratio and lipoprotein variation that induced by copper ion and hyperchlorite and arterial wall cells. But these antioxidants act on different manner. CONCLUSION: BHT, vitamin C and vitamin E can inhibit the oxidative modification of HDL and hence could be potential nutrients to prevent atherosclerosis.

[Study on the detection method for determining the bioactivity of recombinant human interleukin-22].

OBJECTIVE: To develop a method for determining the bioactivity of recombinant human IL-22. METHODS: pSTAT3-TA-Luc and pcDNA3. 1 plasmids were co-transfect to HepG2 cells to generate stable HepG2/ STAT3 cell line with G418 screen. After treating cell with IL-22, the luciferase activity was assayed. Orthogonal test was used to optimize the assay condition, and then the reproducibility and specificity of the assay was checked. RESULTS: The best condition for this assay are: cell density as 4 x 10(5)/mL, stimulating time as 4 hours, luciferase substrate as 50 microL. 50% effective dose (ED50) of IL-22 assayed by this method is 16.89 ng/mL, relative standard deviation (RSD) is 7.09%. Neutralizing antibody test shows the high specificity. Comparing with ELISA, the method described here has more advantages, including higher stability, easier performance and less cost. CONCLUSION: Luciferase reporter gene assay method is a fast, sensitive, reproducible method for IL-22 bioactivity determination.

Wire-in-Tube IrOx Architectures: Alternative Label-Free Immunosensor for Amperometric Immunoassay toward α-Fetoprotein.

A sensitive, label-free immunosensor based on iridium oxide (IrOx, 0≤x≤2) nanofibers, which were synthesized through a simple one-spinneret electrospinning method, was first developed for immunoassay of the cancer biomarker α-fetoprotein (AFP). The specific wire-in-tube nanostructure could be obtained and the composition of IrOx nanofibers also could be controlled through changing the annealing temperature. The unique structure and properties of IrOx nanofibers obtained at 500 °C not only led to increased electrode surface area and accelerated electron transfer kinetics but also could provide a highly stable matrix for the convenient conjugation of biomolecules together with chitosan (CS). The good electrochemical properties of the IrOx-nanofiber-modified immunosensor allowed one to detect AFP over a wide concentration range from 0.05 to 150 ng/mL, with a detection limit of 20 pg/mL. The proposed immunosensor also has been used to determine AFP in human serum with satisfactory results. The present protocol was shown to be quite promising for clinical screening of cancer biomarkers and point-of-care diagnostics applications.

Au-modified three-dimensional In2O3 inverse opals: synthesis and improved performance for acetone sensing toward diagnosis of diabetes.

Analyzing the volatile organic compounds (VOCs) in exhaled breath effectively is crucial to medical treatment, which can provide a fast and noninvasive way to diagnose disease. Well-designed materials with controlled structures have great influence on the sensing performance. In this work, the ordered three dimensional inverse opal (3DIO) macroporous In2O3 films with additional via-hole architectures were fabricated and different amounts of gold nanoparticles (Au NPs) were loaded on the In2O3 films aiming at enhancing their electrical responses. The gas sensing to acetone toward diabetes diagnosis in exhaled breath was performed with different Au/In2O3 electrodes. Representatively, the best 3DIO Au/In2O3 sensor can detect acetone effectively at 340 °C with response of 42.4 to 5 ppm, the actual detection limit is as low as 20 ppb, and it holds a dynamic response of 11 s and a good selectivity. Moreover, clinical tests proved that the as-prepared 3DIO Au/In2O3 IO sensor could distinguish acetone biomarkers in human breath clearly. The excellent gas sensing properties of the Au/In2O3 electrodes were attributed to the "spillover effects" between Au and In2O3 and the special 3DIO structure. This work indicates that 3DIO Au/In2O3 composite is a promising electrode material for actual application in the monitoring and detection of diabetes through exhaled breath.

A sensitive label-free amperometric immunosensor for alpha-fetoprotein based on gold nanorods with different aspect ratio.

A simple and accurate label-free amperometric immunosensor for α-fetoprotein (AFP) detection is developed based on gold nanorods (GNRs) with different aspect ratio and compared with gold particles (GNPs). The positively charged GNRs and GNPs due to the surface immobilized cetyltrimethyl ammonium bromide (CTAB) can adsorb the negatively charged AFP antibody (Ab) directly. The presence of the GNRs not only enhanced the immobilized amount of biomolecules, but also improved the electrochemical properties of the immunosensor. With the aid of GNRs, the electrochemical signal was greatly enhanced in comparison with GNPs. Under optimal conditions, the proposed immunosensor could detect AFP in a linear range from 0.1 to 200 ng/mL with a detection limit of 0.04 ng/mL (signal-to-noise ratio = 3), and it also possessed good reproducibility and storage stability. Moreover, the detection of AFP in five human serum samples also showed satisfactory accuracy. The proposed methodology was potentially attractive for clinical immunoassay.

Preparation and Gas Sensing Properties of In2O3/Au Nanorods for Detection of Volatile Organic Compounds in Exhaled Breath.

A series of In2O3/Au nanorods (NRs) were fabricated and characterized by scanning electron microscope (SEM), transmission electron microscope (TEM), X-ray diffractometer (XRD) and X-ray photoelectron spectroscopy (XPS). The length to diameter ratios of In2O3/Au NRs was periodically modulated in the range of 2.9-4.5 through controlling the initial content of indium salt and reaction time. Their gas sensing properties to volatile organic compounds (VOCs) were carefully studied and then applied in exhaled breath detection. The results demonstrate that In2O3/Au NRs gas sensor can effectively detect acetone at 250 °C and ethanol at 400 °C. The corresponding actual detection limit is as low as 0.1 ppm to acetone and 0.05 ppm to ethanol, respectively. Moreover, by using humidity compensation method, In2O3/Au NRs gas sensor can clearly distinguish the acetone and ethanol biomarkers in human breath. The main reason of the enhanced gas sensing properties was attributed to the "spillover effects" between Au and In2O3 NRs. The excellent sensing performance indicates that In2O3/Au NRs is a promising functional material to actual application in monitoring and detecting diabetes and safe driving area in a noninvasive and more accurate way.

Unglycosylated recombinant human glutathione peroxidase 3 mutant from Escherichia coli is active as a monomer.

Glutathione peroxidase 3 (GPx3) is a glycosylated member of GPx family and can catalyze the reaction of different types of peroxides with GSH to form their corresponding alcohols in vitro. The active center of GPx3 is selenocysteine (Sec), which is incorporated into proteins by a specific mechanism. In this study, we prepared a recombinant human GPx3 (rhGPx3) mutant with all Cys changed to Ser from a Cys auxotrophic strain of E. coli, BL21(DE3)cys. Although lacking post-translational modification, rhGPx3 mutant still retained the ability to reduce H2O2 and PLPC-OOH. Study on the quaternary structure suggested that rhGPx3 mutant existed as a monomer in solution, which is different from native tetrameric GPx3. Loss of the catalytic activity was considered to be attributed to both the absence of glycosylation and the failure of the tetramer. Further analysis was performed to compare the structures of rhGPx3 and GPx4 mutant, which were quite similar except for oligomerization loop. The differences of amino acid composition and electrostatic potentials on the oligomerization loop may affect the binding of large substrates to rhGPx3 mutant. This research provides an important foundation for biosynthesis of functionally selenium-containing GPx3 mutant in E.coli.

Ultrasensitive non-enzymatic glucose sensor based on three-dimensional network of ZnO-CuO hierarchical nanocomposites by electrospinning.

Three-dimensional (3D) porous ZnO-CuO hierarchical nanocomposites (HNCs) nonenzymatic glucose electrodes with different thicknesses were fabricated by coelectrospinning and compared with 3D mixed ZnO/CuO nanowires (NWs) and pure CuO NWs electrodes. The structural characterization revealed that the ZnO-CuO HNCs were composed of the ZnO and CuO mixed NWs trunk (~200 nm), whose outer surface was attached with small CuO nanoparticles (NPs). Moreover, a good synergetic effect between CuO and ZnO was confirmed. The nonenzymatic biosensing properties of as prepared 3D porous electrodes based on fluorine doped tin oxide (FTO) were studied and the results indicated that the sensing properties of 3D porous ZnO-CuO HNCs electrodes were significantly improved and depended strongly on the thickness of the HNCs. At an applied potential of + 0.7 V, the optimum ZnO-CuO HNCs electrode presented a high sensitivity of 3066.4 µAmM(-1)cm(-2), the linear range up to 1.6 mM, and low practical detection limit of 0.21 µM. It also showed outstanding long term stability, good reproducibility, excellent selectivity and accurate measurement in real serum sample. The formation of special hierarchical heterojunction and the well-constructed 3D structure were the main reasons for the enhanced nonenzymatic biosensing behavior.

Synthesis of Au/graphene oxide composites for selective and sensitive electrochemical detection of ascorbic acid.

In this work, we present a novel ascorbic acid (AA) sensor applied to the detection of AA in human sera and pharmaceuticals. A series of Au nanoparticles (NPs) and graphene oxide sheets (Au NP/GO) composites were successfully synthesized by reduction of gold (III) using sodium citrate. Then the Au NP/GO composites were used to construct nonenzymatic electrodes in practical AA measurement. The electrode that has the best performance presents attractive analytical features, such as a low working potential of +0.15 V, a high sensitivity of 101.86 µA mM(-1) cm(-2) to AA, a low detection limit of 100 nM, good reproducibility and excellent selectivity. And more,it was also employed to accurately and practically detect AA in human serum and clinical vitamin C tablet with the existence of some food additive. The enhanced AA electrochemical properties of the Au NP/GO modified electrode in our work can be attributed to the improvement of electroactive surface area of Au NPs and the synergistic effect from the combination of Au NPs and GO sheets. This work shows that the Au NP/GO/GCEs hold the prospect for sensitive and selective determination of AA in practical clinical application.

Synthesis of graphene oxide based CuO nanoparticles composite electrode for highly enhanced nonenzymatic glucose detection.

CuO nanoparticles (NPs) based graphene oxide (CuO/GO) composites with different CuO NPs loading amount as well as pure CuO NPs with different hydrothermal temperatures were synthesized using a hydrothermal method. Transmission electron microscopy (TEM), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and Raman spectroscopy were employed to characterize the morphology and structures of our samples. The influence of hydrothermal temperature, GO sheet, and loading amount of CuO on particle size and structure of CuO was systemically investigated. The nonenzymatic biosensing properties of CuO/GO composites and CuO NPs toward glucose were studied based on glassy carbon electrode (GCE). The sensing properties of CuO NPs were improved after loading on GO sheets. The CuO/GO composites with saturated loading of the CuO NPs exhibited the best nonenzymatic biosensing behavior. It exhibited a sensitivity of 262.52 µA mM(-1) cm(-2) to glucose with a 0.69 µM detection limit (S/N = 3) and a linear range from 2.79 µM to 2.03 mM under a working potential of +0.7 V. It also showed outstanding long term stability, good reproducibility, excellent selectivity, and accurate measurement in real serum sample. It is believed that CuO/GO composites show good promise for further application on nonenzymatic glucose biosensors.