Chitosan/Nitrogen Doped Reduced Graphene Oxide Modified Biosensor for Impedimetric Detection of microRNA

The development of a low‐cost and disposable biosensor platform for the sensitive and rapid detection of microRNAs (miRNAs) is of great interest for healthcare, pharmaceuticals, and medical science. We designed an impedimetric biosensing platform using Chitosan (CHIT)/nitrogen doped reduced graphene oxide (NRGO) conductive composite to modify the surface of pencil graphite electrodes (PGE) for the sensitive detection of miRNAs. An initial optimisation protocol involved investigation of the effect of NRGO concentration and miR 660 DNA probe concentration on the response of the modified electrode. After the optimization protocol, the sequence‐selective hybridization between miR 660 DNA probe and its RNA target was evaluated by measuring changes on charge transfer resistance, R_ct values. Moreover, the selectivity of impedimetric biosensor was tested in the presence of non‐complementary miRNA (NC) sequences, such as miR 34a and miR 16. The hybridization process was examined both in phosphate buffer (PBS) and in PBS diluted fetal bovine serum (FBS:PBS) solutions. The biosensor demonstrated a detection limit of 1.72 μg/mL in PBS and 1.65 μg/mL in FBS:PBS diluted solution. Given the easy, quick and disposable attributes, the proposed conductive nanocomposite biosensor platform shows great promise as a low‐cost sensor kit for healthcare monitoring, clinical diagnostics, and biomedical devices.     

Highly Sensitive Impedimetric Biosensor Based on Thermolysin Immobilized on a GCE Modified with AuNP-decorated Graphene for the Detection of Ochratoxin A

The fabrication of a thermolysin-based biosensor capable of detecting ochratoxin A (OTA) from food samples is described. The electrochemical deposition of calcium cross-linked cellulose film (CCLC) and gold nanoparticles (AuNPs) on graphene (GR) for modification of a glassy carbon electrode (GCE) is the first step. Then the thermolysin (TLN) enzyme in a polyvinyl alcohol (PVA)/polyethylenimine (PEI) matrix is immobilized. The impedimetric biosensor response is linear from 0.2 nM to 100 nM with a detection limit of 0.2 nM. The obtained stable and reproducible biosensor is then applied for the detection of OTA in spiked extracts from coffee beans.     

Impedimetric Biosensor for Bovine Herpesvirus Type 1‐Antigen Detection

The bovine herpesvirus type 1 (BHV‐1) is a pathogen of great economic impact for livestock, which is related multi‐systemic infections that leads to mortality or morbidity of cattles. Thus, the search for cheap and practical methodologies that allow the selective detection of BHV‐1 antigen (BHV‐1 AG) is of utmost relevance. Therefore, an impedimetric label‐free immunosensor was herein, developed and its performance evaluated in biological samples enriched with BHV‐1 AG. Briefly, the biosensor construction was based on the immobilization of BHV‐1 antibody (BHV‐1 AB) and casein on the activated glassy carbon electrode surface. The BHV‐1 AB was isolated from egg yolk of immunized chickens, which is a less stressful protocol. The bio sensing principle was based on Electrochemical Impedance Spectroscopy by using Fe(CN)₆^4?/3? probe, which were also used to check variation of charge transfer resistance (?R_ct), when the electrode surface was increasingly blocked by immune complex. A linear relationship between ?R_ct and BHV‐1 AG concentration was verified in the range from 10 to 50 TCID₅₀ mL^?1, with LOQ of 2.00 TCID₅₀ mL^?1 and LOD of 0.66 TCID₅₀ mL^?1. Besides the suitable sensitivity, the immunosensor displayed accuracy, stability, and specificity to detect BHV‐1 AG in biological samples of serum, nasal secretions, semen and urine. Moreover, to the best of our knowledge this is the first immunosensor applied to BHV‐1 diagnostic.     

Impedimetric DNA‐Based Biosensor for Silver Ions Detection with Hemin/G‐Quadruplex Nanowire as Enhancer

A DNA‐based biosensor was reported for detection of silver ions (Ag⁺) by electrochemical impedance spectroscopy (EIS) with [Fe(CN)₆]^4?/3? as redox probe and hybridization chain reaction (HCR) induced hemin/G‐quadruplex nanowire as enhanced label. In the present of target Ag⁺, Ag⁺ interacted with cytosine‐cytosine (C? C) mismatch to form the stable C? Ag⁺? C complex with the aim of immobilizing the primer DNA on electrode, which thus triggered the HCR to form inert hemin/G‐quadruplex nanowire with an amplified EIS signal. As a result, the DNA biosensor showed a high sensitivity with the concentration range spanning from 0.1 nM to 100 µM and a detection limit of 0.05 nM.     

Surface‐Enhanced Raman Scattering Based on Controllable‐Layer Graphene Shells Directly Synthesized on Cu Nanoparticles for Molecular Detection

Graphene shells with a controllable number of layers were directly synthesized on Cu nanoparticles (CuNPs) by chemical vapor deposition (CVD) to fabricate a graphene‐encapsulated CuNPs (G/CuNPs) hybrid system for surface‐enhanced Raman scattering (SERS). The enhanced Raman spectra of adenosine and rhodamine 6G (R6G) showed that the G/CuNPs hybrid system can strongly suppress background fluorescence and increase signal‐to‐noise ratio. In four different types of SERS systems, the G/CuNPs hybrid system exhibits more efficient SERS than a transferred graphene/CuNPs hybrid system and pure CuNPs and graphene substrates. The minimum detectable concentrations of adenosine and R6G by the G/CuNPs hybrid system can be as low as 10^?8 and 10^?10 M , respectively. The excellent linear relationship between Raman intensity and analyte concentration can be used for molecular detection. The graphene shell can also effectively prevent surface oxidation of Cu nanoparticles after exposure to ambient air and thus endow the hybrid system with a long lifetime. This work provides a basis for the fabrication of novel SERS substrates.     

Capacitance Polypyrrole‐based Impedimetric Immunosensor for Interleukin‐10 Cytokine Detection

In the present study, we developed a novel label‐free capacitance impedimetric immunosensor based on the immobilization of the human monoclonal antibody anti‐interleukin‐10 (anti‐IL‐10 mAb) onto polypyrrole (PPy)‐modified silicon nitride (Si₃N₄) substrates. The immunosensor was used for the detection of the recombinant interleukin‐10 antigen (rh IL‐10) that may be secreted in patients at the early stage of inflammation. The immunosensor was created by chemical deposition of PPy conducting layer on pyrrole?silane (SPy)‐treated Si/SiO₂/Si₃N₄ substrates (Si/SiO₂/Si₃N₄?SPy), followed by anti‐IL‐10 mAb immobilization through carboxyl‐functionalized diazonium (CMA) protocol and carbodiimide chemistry. The surface characterization and the biofunctionalization steps were characterized by SEM, FTIR and cyclic voltammetry (CV) while the detection process was carried out by using electrochemical impedance spectroscopy (EIS) analyses. The created immunosensor showed two linear fittings (R²=0.999) for the detection of rh IL‐10 within the concentration range from 1–50 pg/mL. It exhibited high sensitivity (0.1128 (pg/mL)^?1) with a very low limit of detection (LOD)=0.347 pg/mL, more particularly, at the low concentration range (1–10 pg/mL). Thus, this developed polypyrrole‐based immunosensor represents a promising strategy for creation of miniaturized label‐free, fast and highly sensitive biosensors for diagnosis of inflammation biomarkers at very low concentrations with reduced cost.     

基于纳米ZnO/壳聚糖复合膜DNA电化学传感器用于HIV基因的免标记检测

以室温固相合成法制备纳米ZnO,通过壳聚糖(CHIT)的成膜效应将纳米ZnO固定在玻碳电极(GCE)表面,制得的ZnO/CHIT/GCE电极成为DNA固定和杂交的良好平台。DNA的固定和杂交通过电化学交流阻抗进行表征。以电化学交流阻抗免标记法检测目标DNA,固定于电极表面的DNA探针与目标DNA杂交后使电极表面的电子传递电阻增大,以此作为检测信号可以高灵敏度地测定目标DNA。电化学阻抗谱检测人类免疫缺陷病毒(HIV)基因片段的线性范围为2.0×10-11~2.0×10-6mol/L,检出限为2.0×10-12mol/L。     

Synthesis,Characterization, Electronic and Gas‐Sensing Properties towards H2 and CO of Transparent,Large‐Area,Low‐Layer Graphene

Low‐layered, transparent graphene is accessible by a chemical vapor deposition (CVD) technique on a Ni‐catalyst layer, which is deposited on a <100> silicon substrate. The number of graphene layers on the substrate is controlled by the grain boundaries in the Ni‐catalyst layer and can be studied by micro Raman analysis. Electrical studies showed a sheet resistance (R_sheet) of approximately 1435 Ω per □, a contact resistance (R_c) of about 127 Ω, and a specific contact resistance (R_sc) of approximately 2.8×10^?4 Ω cm² for the CVD graphene samples. Transistor output characteristics for the graphene sample demonstrated linear current/voltage behavior. A current versus voltage (I_ds–V_ds) plot clearly indicates a p‐conducting characteristic of the synthesized graphene. Gas‐sensor measurements revealed a high sensor activity of the low‐layer graphene material towards H₂ and CO. At 300 °C, a sensor response of approximately 29 towards low H₂ concentrations (1 vol %) was observed, which is by a factor of four higher than recently reported.     

Employing Label‐free Electrochemical Biosensor Based on 3D‐Reduced Graphene Oxide and Polyaniline Nanofibers for Ultrasensitive Detection of Breast Cancer BRCA1 Biomarker

A label‐free DNA biosensor based on three‐dimensional reduced graphene oxide (3D‐rGO) and polyaniline (PANI) nanofibers modified glassy carbon electrode (GCE) was successfully developed for supersensitive detection of breast cancer BRCA1. The results demonstrated that 3D‐rGO and PANI nanofibers had synergic effects for reducing the charge transfer resistance (R_ct), meaning a huge enhancement in electrochemical activity of 3D‐rGO‐PANI/GCE. Probe DNA could be immobilized on 3D‐rGO‐PANI/GCE for special and sensitive recognition of target DNA (1.0×10^?15–1.0×10^?7 M) with a theoretical LOD of 3.01×10^?16 M (3S/m). Furthermore, this proposed nano‐biosensor could directly detect BRCA1 in real blood samples.     

High‐Quality Three‐Dimensional Nanoporous Graphene

We report three‐dimensional (3D) nanoporous graphene with preserved 2D electronic properties, tunable pore sizes, and high electron mobility for electronic applications. The complex 3D network comprised of interconnected graphene retains a 2D coherent electron system of massless Dirac fermions. The transport properties of the nanoporous graphene show a semiconducting behavior and strong pore‐size dependence, together with unique angular independence. The free‐standing, large‐scale nanoporous graphene with 2D electronic properties and high electron mobility holds great promise for practical applications in 3D electronic devices.     

Single‐Walled Carbon Nanotubes Modified Gold Electrodes as an Impedimetric DNA Sensor

A gold surface modified with a self‐assembled monolayer of 11‐amino‐1‐undecanethiol (AUT) was used for the covalent immobilization of oxidized single‐walled carbon nanotubes (SWNTs). The as‐described SWNTs‐modified substrate was subsequently used to attach single‐stranded deoxyribonucleic acid (ssDNA) used as a substrate for DNA hybridization. Electrochemical impedance spectroscopy measurements were performed to follow the DNA hybridization process by using the redox couple [Fe(CN)₆]^3−/4− as a marker ion. Specifically, changes in charge transfer resistance obtained from the Nyquist plots were used as the sensing parameter of DNA hybridization. The substrate sensitivity towards changes in target DNA concentration, its selectivity toward different DNA sequences and its reusability are successfully demonstrated in this report.     

Electrochemical Signal Enhancer Fabricated Using Lysine‐rich Peptide for Ultrasensitive Electrochemical DNA Biosensor Analysis

Here we present a novel design of electrochemical signal enhancer to increase the detection sensitivity of electrochemical DNA biosensors. The key element of this enhancer is a lysine‐rich peptide (LRP). Its C‐terminal is conjugated with a planer molecule, being able to intercalate into the base pairs of probe‐target duplexes. The lysine residues of LRP are covalently linked with electrochemical signal indicators, acting as an assembly of electrochemical signal indicators. Experimental results proved the feasibility of the novel design. We have examined the effects of the numbers of lysine residues and the hybridization conditions on the detection sensitivity. The optimization procedures have led to significant sensitivity enhancement, and the LOD (limit of detection) has been determined to be 1.4 amol. This enhancer demonstrates advantages of easy operation, simple instrumentation, and high exemption from environmental influence.     

Reduced Graphene Oxide/Pt Nanoparticles/Zn‐MOF‐74 Nanomaterial for a Glucose Biosensor Construction

In this study, a new glucose biosensor was fabricated by immobilizing glucose oxidase (GOx) on platinum nanoparticles (Pt NPs) decorated reduced graphene oxide (rGO)/Zn‐MOF‐74 hybrid nanomaterial. Herein, the biosensor fused the advantages of rGO with those of porous Zn‐MOF and conductive Pt NPs. This has not only enlarged the surface area and porosity for the efficient GOx immobilization and faster mass transport, but also provided favorable electrochemical features such as high current density, remarkable electron mobility through metal nanoparticles, and improved electron transfer between the components. The GOx‐rGO/Pt NPs@Zn‐MOF‐74 coated electrode displayed a linear measurement range for glucose from 0.006 to 6 mM, with a detection limit of 1.8 μM (S/N: 3) and sensitivity of 64.51 μA mM^?1 cm^?2. The amperometric response of the enzyme biosensor demonstrated the typical behavior of Michaelis‐Menten kinetics. The obtained satisfying sensitivity and measurement range enabled fast and accurate glucose measurement in cherry juice using the fabricated biosensor. The water‐stable Zn‐MOF‐74 demonstrated higher enzyme loading capacity and can be potent supporting material for biosensor construction.     

Impedimetric DNA Sensor Based on Poly(proflavine) for Determination of Anthracycline Drugs

Electropolymerization is often used as a tool for immobilization of charged biopolymers and their electric wiring in the biosensor assembly. In this work, electropolymerization of proflavine has been for the first time used for the physical adsorption of DNA and measurement of anthracycline drugs (doxorubicin and daunorubicin) able to intercalate DNA. Redox properties of the proflavine polymers on the glassy carbon electrode and DNA deposition were characterized with cyclic voltammetry, scanning electron microscopy and electrochemical impedance spectroscopy. It was shown that DNA decreased the peak currents on voltammogram but increased the charge transfer resistance. The latter effect is pronounced with the following drug application. The impedimetric response regularly increased with the drugs concentration in the range 1 nM to 0.1 μM for doxorubicin and 1 pM–10 nM for daunorubicin (limit of detections 0.3 and 0.001 nM, respectively). The biosensor was tested on pharmaceutical preparations and spiked solution simulating the plasma electrolytes and possible interference of serum proteins.     

Preparation of GOD/Sol‐Gel Silica Film on Prussian Blue Modified Electrode for Glucose Biosensor Application

A novel amperometric glucose biosensor was fabricated by in situ incorporating glucose oxidase (GOD) within the sol‐gel silica film on a Prussian blue (PB) modified electrode. The method is simple and controllable, which combined the merits of in situ immobilizing biomolecules in sol‐gel silica film by electrochemical method and the synergic catalysis effects of PB and GOD molecules. Scanning electron microscopy (SEM) showed that the GOD/sol‐gel silica film was homogeneous with a large number of three‐dimensional nanopores, which not only enhanced mass transport, but also maintained the active configuration of the enzyme molecule and prevented the leakage of enzyme, therefore improved the stability and sensitivity of the biosensor. The fabricated biosensor showed fast response time (10 s), high sensitivity (26.6 mA cm^?2 M^?1), long‐term stability, good suppression of interference, and linear range of 0.01 mM–5.8 mM with a low detection limit of 0.94 μM for the detection of glucose. In addition, the biosensor was successfully applied to determine glucose in human serum samples.     

Superclean Growth of Graphene Using a Cold‐Wall Chemical Vapor Deposition Approach

Chemical vapor deposition (CVD) has become a promising approach for the industrial production of graphene films with appealing controllability and uniformity. However, in the conventional hot‐wall CVD system, CVD‐derived graphene films suffer from surface contamination originating from the gas‐phase reaction during the high‐temperature growth. Shown here is that the cold‐wall CVD system is capable of suppressing the gas‐phase reaction, and achieves the superclean growth of graphene films in a controllable manner. The as‐received superclean graphene film, exhibiting improved optical and electrical properties, was proven to be an ideal candidate material used as transparent electrodes and substrate for epitaxial growth. This study provides a new promising choice for industrial production of high‐quality graphene films, and the finding about the engineering of the gas‐phase reaction, which is usually overlooked, will be instructive for future research on CVD growth of graphene.     

Homogeneous Electrochemical Assay for Real‐time Monitoring of Exonuclease III Activity Based on a Graphene Monolayer

A homogeneous electrochemical assay based on a graphene monolayer electrode was developed for simple, sensitive, rapid and quantitative analysis of the exonuclease III (Exo III) activity. The method utilized a methylene blue (MB) tagged DNA substrate with hairpin structure, and a graphene monolayer attached on the working electrode. Before digestion, the hairpin structure prevents the adsorption of the DNA substrate to the graphene surface. Degradation of the substrate by the 3′–5′ Exo III, however, yields single‐stranded DNA (ssDNA), resulting in its subsequent binding to the graphene surface through π‐π stacking, which produces the voltammetric current from electrochemical reduction of the MB tag anchoring at the end of ssDNA. A direct quantification of the Exo III activity can be achieved by measuring the reductive peak current of the MB tag under easily attainable potential (∼ −0.1 V vs Ag/AgCl) range comparably sensitive to the conventional methods such as a gel‐based or fluorescence‐based assays. Our approach can be applied to measure various exonucleases activity by adjusting the structure of DNA substrate suggesting a new assay method in drug screening and basic research related to the enzymes.     

Electrochemical Impedance Characterization of Nafion‐Coated Carbon Film Resistor Electrodes for Electroanalysis

Carbon film disk electrodes with Nafion coatings have been characterized by electrochemical impedance spectroscopy (EIS) with a view to a better understanding of their advantages and limitations in electroanalysis, particularly in anodic stripping voltammetry of metal ions. After initial examination by cyclic voltammetry, spectra were recorded over the full potential range in acetate buffer solution at the bare electrodes, electrodes electrochemically pretreated in acid solution, and Nafion‐coated pretreated electrodes in the presence and absence of dissolved oxygen. EIS equivalent circuit analysis clearly demonstrated the changes between these electrode assemblies. In order to simulate anodic stripping voltammetry conditions, spectra were also obtained in the presence of cadmium and lead ions in solution at Nafion‐coated electrodes, both after metal ion deposition and following re‐oxidation. Permanent changes to the structure of the Nafion film occurred, which has implications for use of these electrode assemblies in anodic stripping voltammetry at relatively high trace metal ion concentrations.