Facile preparation of a DNA sensor for rapid herpes virus detection

In this paper, a simple DNA sensor platform was developed for rapid herpes virus detection in real samples. The deoxyribonucleic acid (DNA) sequences of the herpes simplex virus (DNA probe) were directly immobilized on the surface of interdigitated electrodes by electrochemical polymerization along with pyrrole monomers. The potential was scanned from ? 0.7 to + 0.6 V, and the scanning rate was 100 mV/s. Fourier transform infrared spectroscopy was employed to verify specific DNA sequence binding and the conducting polymer. The morphology of the conducting polymer doped with DNA strands was characterized using a field emission scanning electron microscope. As-obtained DNA sensor was used to detect the herpes virus DNA in the real samples. The results show that the current DNA sensors detected the lowest DNA concentration of 2 nM. This sensitivity appears to be better than that of the DNA sensors prepared by immobilization of the DNA probe on the 3-aminopropyl-triethoxy-silance (APTS) membrane.     

Organische Polymere als funktionelle Werkstoffe für chemische Sensoren

Organic polymers as functional materials for chemical sensors The function of many chemical sensors for measurements in liquids and in gases with ambient temperature is based on the combination of a transducer with organic membranes. These membrans determine essential sensor properties as selectivity, sensitivity and response characteristics. In addition they protect the detection system against external influences. Therefore the selection and synthesis of polymer membranes are an essential constituent of the sensor investigation and sensor development. Electrical, optical and biological properties of the polymers are important in this case. A survey of the materials used in the remote sensing is given. Of special interest to the sensor investigation are in last time intrinsic conducting polymers (ILP) whose properties opened new possibilities of the sensor development. With the help of an electrochemical pH glass electrode with inner solid contact it is shown that polypyrrole can be used as a material for a long‐lived inner solid contact and as substitute for inner secondary reference electrode. Practice tests confirm the suitability of this polymer material. Aspects of the transport mechanism of electrical charges through the boundary surface conducting polymer | glass are discussed.     

Hydrazine-catalyzed ultrasensitive detection of DNA and proteins

A sensitive electrochemical assay of DNA and proteins employing electrocatalytic reduction of hydrogen peroxide by labeled hydrazine on the probe immobilized surfaces was developed. The method utilizes a conducting polymer, poly-5, 2':5', 2'-terthiophene-3'-carboxylic acid (pTTCA), covalently linked to the dendrimer (DEN) and hydrazine. The detection signal was amplified by the pTTCA/DEN assembly loaded with Au nanoparticles (particle size, approximately 3.5 nm) onto which huge target DNA- or proteins-linked hydrazine labels (avidin-hydrazine) were adsorbed. The linear dynamic ranges for the electrocatalytic detection of DNA and proteins, extending from 1.0 fM to 10 microM and 10 fg/mL to 10 ng/mL, were observed, along with the detection limits of 450 aM (2700 DNA molecules in a 10-microL sample) and 4.0 fg/mL, respectively. The method eliminates the use of enzymes for DNA and protein detection and opens a way for DNA-free detection of proteins. The simplicity, good reproducibility (RSD, <4.3% for n = 10), and low detection limit of the method offer a good promise for practical DNA and protein analyses.     

Experimental and theoretical investigation of sensing parameters in carbon nanotube‐based DNA sensor

Nowadays, sensitive biosensors with high selectivity, lower costs and short response time are required for detection of DNA. The most preferred materials in DNA sensor designing are nanomaterials such as carbon and Au nanoparticles, because of their very high surface area and biocompatibility which lead to performance and sensitivity improvements in DNA sensors. Carbon nanomaterials such as carbon nanotubes (CNTs) can be considered as a suitable DNA sensor platform due to their high surface‐to‐volume ratio, favourable electronic properties and fast electron transfer rate. Therefore, in this study, the CNTs which are synthesised by pulsed AC arc discharge method on a high‐density polyethylene substrate are used as conducting channels in a chemiresistor for the electrochemical detection of double stranded DNA. Moreover, the response of the proposed sensor is investigated experimentally and analytically in different temperatures, which confirm good agreement between the presented model and experimental data.Inspec keywords: electrochemical sensors, polymers, arcs (electric), biological techniques, nanosensors, carbon nanotubes, DNAOther keywords: C, chemiresistor, double stranded DNA detection, CNT, electronic properties, surface‐to‐volume ratio, nanoparticles, biosensors, electrochemical detection, high‐density polyethylene substrate, pulsed AC arc discharge method, electron transfer rate, carbon nanomaterials, carbon nanotube‐based DNA sensor     

Label-free detection of DNA hybridization at a liquid|liquid interface

A novel electrochemical approach for label-free detection of DNA primary sequence has been proposed. The flow of nonelectroactive ions across a liquid|liquid interface was used as an electrochemical probe for detection of DNA hybridization. Disposable graphite screen-printed electrodes shielded with a thin layer of inert polymer plasticized with water-immiscible polar organic solvent were modified by probe oligonucleotide and used as a DNA sensor. The specific DNA coupling has been detected with impedance spectroscopy by decrease of ion-transfer resistance. The detection limit was of 10-8 M of target oligonucleotide. The reported sensor was suitable for discrimination of a single mismatch oligonucleotide from the full complementary one. The reported DNA sensor was advantageous over known physicochemical approaches, providing the most significant changes in the measured parameters.     

Enzyme-amplified electrochemical detection of DNA using electrocatalysis of ferrocenyl-tethered dendrimer

We have developed a sandwich-type enzyme-linked DNA sensor as a new electrochemical method to detect DNA hybridization. A partially ferrocenyl-tethered poly(amidoamine) dendrimer (Fc-D) was used as an electrocatalyst to enhance the electronic signals of DNA detection as well as a building block to immobilize capture probes. Fc-D was immobilized on a carboxylic acid-terminated self-assembled monolayer (SAM) by covalent coupling of unreacted amine in Fc-D to the acid. Thiolated capture probe was attached to the remaining amine groups of Fc-D on the SAM via a bifunctional linker. The target DNA was hybridized with the capture probe, and an extension in the DNA of the target was then hybridized with a biotinylated detection probe. Avidin-conjugated alkaline phosphatase was bound to the detection probe and allowed to generate the electroactive label, p-aminophenol, from p-aminophenyl phosphate enzymatically. p-Aminophenol diffuses into the Fc-D layer and is then electrocatalytically oxidized by the electronic mediation of the immobilized Fc-D, which leads to a great enhancement in signal. Consequently, the amount of hybridized target can be estimated using the intensity of electrocatalytic current. This DNA sensor exhibits a detection limit of 20 fmol. Our method was also successfully applied to the sequence-selective discrimination between perfectly matched and single-base mismatched target oligonucleotides.     

Flow injection based renewable electrochemical sensor system

A novel class of electrochemical sensors is proposed utilizing electrically conducting beads to form a disposable electrode as well as nonconducting beads to form renewable layers of immobilized enzymes. The concept, aimed to prevent fouling, is tested on an amperometric sensor coupled to nonconducting beads with different immobilized oxidases: galactose, lactate, alcohol, or glucose oxidase, the latter two being used to determine alcohol and gluocse, respectively, in samples of beer and wine. Glucose oxidase was also immobilized on conducting glassy carbon particles to explore the performance of a biosensor where both enzyme and electrode can be automatically renewed in less than 1 min. The results confirm that the concept of a flow injection renewable electrochemical sensor (FI-RES) is practical. It provides a novel approach to biosensing, to comparing enzyme activity, to studying enzyme immobilization on different supports, and to voltammetry in general.     

Pulsed galvanostatic control of solid-state polymeric ion-selective electrodes

We report on galvanostatically controlled solid-state reversible ion-selective sensors for cationic analytes utilizing a conducting polymer as a transduction layer between the polymeric membrane and electron-conductive substrate. The instrumental control of polymeric membrane ion-selective electrodes based on electrochemically induced periodic ion extraction in alternating galvanostatic/potentiostatic mode was introduced recently creating exciting possibilities to detect clinically relevant polyions such as heparin and protamine and drastically improve the sensitivity of ion-selective sensors limited by the Nernst equation. The present study forms the basis for development of reliable, robust, and possibly maintenance-free sensors that can be fabricated using screen-printing technology. Various aspects of the development of solid-contact galvanostatically controlled ion-selective electrodes with a conducting polymer as a transduction layer are considered in the present work on the example of a model system based on a sodium-selective membrane. The protamine-selective solid-contact sensor was fabricated and characterized, which represents the next step toward commercially viable polyion sensing technology. A substantial improvement of a low detection limit (0.03 mg L-1) was achieved. A simplified diffusion-based theoretical model is discussed predicting the polarization at the interface of the conducting polymer and the membrane, which can cause the disruption of the sensor response function at relatively small current densities.     

Elaboration of a new hydrogen peroxide biosensor using microperoxidase 8 (MP8) immobilized on a polypyrrole coated electrode

A novel strategy for elaborating a new biosensor for hydrogen peroxide has been developed by combining the known properties of microperoxidase 8 (MP8) as an oxidation catalyst, and the interesting properties of conducting polypyrrole as a supporting matrix to allow a good bioelectrochemical interface and a large dispersion of MP8 on the modified glassy carbon electrode.

MP8 was immobilized into the conducting polypyrrole by entrapment during the electrochemical polymerization, and the modified electrode was characterized both by electrochemical and FT-IR measurements. We demonstrated that MP8 could be immobilized into polypyrrole and could undergo an efficient electron transfer.

The obtained modified electrode showed a high catalytic activity toward H₂O₂ without the need for an electron mediator. A linear calibration curve was obtained by amperometric measurement at a potential of − 0.1 V/ECS for concentrations of H₂O₂ ranging from 1 to 10 μM. The detection limit obtained was 1 nM which constituted a real improvement, by about three orders of magnitude, when compared to the values reported for other systems using an electrochemical detection.     

Impedimetric biosensor based on magnetic nanoparticles for electrochemical detection of dopamine

One of the difficulties which limit the use of electrochemical sensors for detection of dopamine is the interference from ascorbic acid. We have sought to address this problem through the synthesis and characterization of a suitable electrode material based on magnetic nanoparticles. The interference from the ascorbic acid was overcome by fabricating a negatively charged electrode surface using PEGylated arginine functionalized magnetic nanoparticles (PA-MNPs). The nanoparticles were characterized by various techniques viz., X-ray diffraction, FT-Infrared spectroscopy, transmission electron microscopy and vibrating sample magnetometer. The electrochemical behavior of the proposed sensor was investigated by cyclic voltammetry and the sensor showed high sensitivity and selectivity for dopamine. The response mechanism of the modified electrode is based on the interaction between the negatively charged electrode and the positively charged dopamine. Under optimized conditions, linear calibration plots were obtained for amperometric detection of dopamine (DA) over the concentration range of 1–9 mM dopamine, with a linear correlation coefficient of 0.9836, sensitivity of 121 μA/mM and a detection limit of 7.25 μM. Electrochemical impedance spectroscopy (EIS) has been used to study the interface properties of modified electrodes. The value of the polarization resistance (R_p) increases linearly with dopamine concentration in the range of 10 μM to 1 mM and the limit of detection (LOD) was calculated to be 14.1 μM. High sensitivity and selectivity, micromolar detection limit, high reproducibility, along with ease of preparation of the electrode surface make this system suitable for the determination of DA in pharmaceutical and clinical preparations.     

Inkjet printing of nanoporous gold electrode arrays on cellulose membranes for high-sensitive paper-like electrochemical oxygen sensors using ionic liquid electrolytes

A simple approach to the mass production of nanoporous gold electrode arrays on cellulose membranes for electrochemical sensing of oxygen using ionic liquid (IL) electrolytes was established. The approach, combining the inkjet printing of gold nanoparticle (GNP) patterns with the self-catalytic growth of these patterns into conducting layers, can fabricate hundreds of self-designed gold arrays on cellulose membranes within several hours using an inexpensive inkjet printer. The resulting paper-based gold electrode arrays (PGEAs) had several unique properties as thin-film sensor platforms, including good conductivity, excellent flexibility, high integration, and low cost. The porous nature of PGEAs also allowed the addition of electrolytes from the back cellulose membrane side and controllably produced large three-phase electrolyte/electrode/gas interfaces at the front electrode side. A novel paper-based solid-state electrochemical oxygen (O(2)) sensor was therefore developed using an IL electrolyte, 1-butyl-3-methylimidazolium hexafluorophosphate (BMIMPF(6)). The sensor looked like a piece of paper but possessed high sensitivity for O(2) in a linear range from 0.054 to 0.177 v/v %, along with a low detection limit of 0.0075% and a short response time of less than 10 s, foreseeing its promising applications in developing cost-effective and environment-friendly paper-based electrochemical gas sensors.     

Conducting polymer 1-dimensional nanostructures for FET sensors

In this review we have highlighted advantages of 1-dimensional nanostructures for field effect transistor (FET)/chemiresistor based sensors and advantages of conducting polymer as material of construction over other nanomaterials. Here we have ensembled different techniques used for the fabrication, assembly/alignment, functionalization and sensing applications of conducting polymer nanowire/tube/junction based FET sensors for gas and biomolecule detection. The advantages and disadvantages of various fabrication, functionalization, and assembling techniques are discussed. We evaluate how such devices have enabled the achievement of improved sensor performance in terms of high sensitivity, selectivity and low detection limits. Finally, we conclude by highlighting overall merits of different techniques and challenges researchers face in the field of conducting polymer 1-dimensional nanostructures-based sensors and also predict the future direction in which research efforts are likely to flourish.     

Highly Sensitive and Selective Field‐Effect‐Transistor NonEnzyme Dopamine Sensors Based on Pt/Conducting Polymer Hybrid Nanoparticles

Dopamine (DA), as one of catecholamine family of neurotransmitters, is crucially important in humans owing to various critical effects on biometric system such as brine circuitry, neuronal plasticity, organization of stress responses, and control of cardiovascular and renal organizations. Abnormal level of dopamine in the central nervous system causes several neurological diseases, e.g., schizophrenia, Parkinson's disease, and attention deficit hybperactivity disorder (ADHD)/attention deficit disorder (ADD). In this report, we suggest the fabrication of nonenzyme field effect transistor (FET) sensor composed of immobilized Pt particle decorated conducting‐polymer (3‐carboxylate polypyrrole) nanoparticles (Pt_CPPy) to detect dopamine. The hybrid nanoparticles (NPs) are produced by means of facile chemical reduction of pristine CPPyNP‐contained Pt precursor (PtCl₄) solution. The Pt_CPPys are then immobilized on an amine‐functionalized (–NH₂) interdigitated‐array electrode substrate, through the formation of covalent bonds with amine groups (–CONH). The resulting Pt_CPPy‐based FET sensors exhibit high sensitivity and selectivity toward DA at unprecedentedly low concentrations (100 × 10^?15m ) and among interfering biomolecules, respectively. Additionally, due to the covalent bonding involved in the immobilization process, a longer lifetime is expected for the FET sensor.     

Bulk acoustic wave affinity biosensor for genetically modified organisms detection

Bulk acoustic waves have been applied as affinity sensors. In particular, a nucleic acid sensor for hybridization studies has been developed and applied for detecting DNA target sequences in solution. A DNA probe is immobilized on the sensor surface while the target sequence is free in solution; the interaction between the two complementary strands (hybridization) is followed in real-time, without the use of any label. The system has been applied to analytical problems, i.e., genetically modified organisms (GMOs) detection. The probe was complementary to characteristic DNA sequences present in GMOs. The probe sequences were internal to the sequence of 35S promoter and Nos terminator that are inserted sequences in the genome of the GMO regulating the transgene expression. Two different probe immobilization procedures were characterized to improve the performances of a piezoelectric crystal DNA sensor for GMOs detection: 1) thiol-dextran-streptavidin-biotin procedure and 2) thiol-derivatized probe and blocking thiol procedure. The system has been optimized using synthetic oligonucleotides. The probe immobilization step was monitored by a surface plasmon resonance system.     

Hybrid surface platform for the simultaneous detection of proteins and DNAs using a surface plasmon resonance imaging sensor

In this work, we present a novel surface and assay for the simultaneous detection of DNA and protein analytes on a surface plasmon resonance (SPR) imaging sensor. A mixed DNA/oligo (ethylene glycol) (OEG) self-assembled monolayer (SAM) is created using a microarrayer. Thiol-modified single-stranded DNA sequences are spotted onto a gold-coated glass substrate. Backfilling with an OEG-modified alkanethiol creates a protein-resistant surface background. Antibodies conjugated to complementary single-stranded DNA sequences are immobilized on the surface through DNA hybridization. By converting only part of the DNA array into a protein array, simultaneous detections of DNA and protein analytes are possible. A model system of two cDNA sequences and two human pregnancy hormones are used to demonstrate the assay. No cross-reactivity was observed between DNA or protein analytes and nontargeted immobilized cDNA sequence or antibodies. A response from a detection of a single analyte in a mixture of protein and DNA analytes corresponds well with that of a single-analyte solution.     

DNA nanostructure-decorated surfaces for enhanced aptamer-target binding and electrochemical cocaine sensors

The sensitivity of aptamer-based electrochemical sensors is often limited by restricted target accessibility and surface-induced perturbation of the aptamer structure, which arise from imperfect packing of probes on the heterogeneous and locally crowded surface. In this study, we have developed an ultrasensitive and highly selective electrochemical aptamer-based cocaine sensor (EACS), based on a DNA nanotechnology-based sensing platform. We have found that the electrode surface decorated with an aptamer probe-pendant tetrahedral DNA nanostructure greatly facilitates cocaine-induced fusion of the split anticocaine aptamer. This novel design leads to a sensitive cocaine sensor with a remarkably low detection limit of 33 nM. It is also important that the tetrahedra-decorated surface is protein-resistant, which not only suits the enzyme-based signal amplification scheme employed in this work, but ensures high selectivity of this sensor when deployed in sera or other adulterated samples.     

Electrical Graphene Aptasensor for Ultra‐Sensitive Detection of Anthrax Toxin with Amplified Signal Transduction

Detection of the anthrax toxin, the protective antigen (PA), at the attomolar (aM) level is demonstrated by an electrical aptamer sensor based on a chemically derived graphene field‐effect transistor (FET) platform. Higher affinity of the aptamer probes to PA in the aptamer‐immobilized FET enables significant improvements in the limit of detection (LOD), dynamic range, and sensitivity compared to the antibody‐immobilized FET. Transduction signal enhancement in the aptamer FET due to an increase in captured PA molecules results in a larger 30 mV/decade shift in the charge neutrality point (V_g,min) as a sensitivity parameter, with the dynamic range of the PA concentration between 12 aM (LOD) and 120 fM. An additional signal enhancement is obtained by the secondary aptamer‐conjugated gold nanoparticles (AuNPs‐aptamer), which have a sandwich structure of aptamer/PA/aptamer‐AuNPs, induce an increase in charge‐doping in the graphene channel, resulting in a reduction of the LOD to 1.2 aM with a three‐fold increase in the V_g,min shift.     

Electrochemical DNAzyme sensor for lead based on amplification of DNA-Au bio-bar codes

An electrochemical DNAzyme sensor for sensitive and selective detection of lead ion (Pb(2+)) has been developed, taking advantage of catalytic reactions of a DNAzyme upon its binding to Pb(2+) and the use of DNA-Au bio-bar codes to achieve signal enhancement. A specific DNAzyme for Pb(2+) is immobilized onto an Au electrode surface via a thiol-Au interaction. The DNAzyme hybridizes to a specially designed complementary substrate strand that has an overhang, which in turn hybridizes to the DNA-Au bio-bar code (short oligonucleotides attached to 13 nm gold nanoparticles). A redox mediator, Ru(NH3)6(3+), which can bind to the anionic phosphate of DNA through electrostatic interactions, serves as the electrochemical signal transducer. Upon binding of Pb(2+) to the DNAzyme, the DNAzyme catalyzes the hydrolytic cleavage of the substrate, resulting in the removal of the substrate strand along with the DNA bio-bar code and the bound Ru(NH3)6(3+) from the Au electrode surface. The release of Ru(NH3)6(3+) results in lower electrochemical signal of Ru(NH3)6(3+) confined on the electrode surface. Differential pulse voltammetry (DPV) signals of Ru(NH3)6(3+) provides quantitative measures of the concentrations of Pb(2+), with a linear calibration ranging from 5 nM to 0.1 microM. Because each nanoparticle carries a large number of DNA strands that bind to the signal transducer molecule Ru(NH3)6(3+), the use of DNA-Au bio-bar codes enhances the detection sensitivity by five times, enabling the detection of Pb(2+) at a very low level (1 nM). The DPV signal response of the DNAzyme sensor is negligible for other divalent metal ions, indicating that the sensor is highly selective for Pb(2+). Although this DNAzyme sensor is demonstrated for the detection of Pb(2+), it has the potential to serve as a general platform for design sensors for other small molecules and heavy metal ions.     

Solid-contact electrochemical polyion sensors for monitoring peptidase activities

We report here on improved solid-contact electrochemical polyion sensors for the detection of polyion protamine. The polymeric membrane sensors were fabricated with a conducting polymer as an ion-electron transduction layer. We observed that decreasing the magnitude of the applied current pulse caused a significant improvement of the sensor sensitivity to low protamine levels. The protamine sensors exhibited a stable and reversible response to protamine concentrations ranging from 0.05 to 30 mg L-1. The sensors were used for monitoring peptidase activities utilizing galvanostatically controlled solid-contact membrane sensors. The polyion protamine was used as a substrate to detect the activity of the protease trypsin. The enzyme activity was continuously monitored by measuring the protamine concentration as it is cleaved by enzyme into smaller fragments to which the sensor is less sensitive. In the presence of a given level of protamine the initial rate of reaction can be linearly related to the trypsin activity within a 0-5 U mL-1 concentration range. The interference with the enzymatic reaction product arginine was specifically examined.     

Electrochemical and optical sugar sensors based on phenylboronic acid and its derivatives

Recent progress in electrochemical and optical sugar sensors based on phenylboronic acid (PBA) and its derivatives as recognition components is reviewed. PBAs are known to bind diol compounds including sugars to form cyclic boronate esters that are negatively charged as a result of the addition of OH^? ions from solution. Based on the formation of PBA charged species, sugars and their derivatives can be detected by means of electrochemical and optical techniques. For the development of PBA-based electrochemical sensing systems or sensors, PBA is modified with a redox-active marker, because PBA itself is electrochemically inactive, and ferrocene derivatives are often employed for this purpose. Ferrocene-modified PBAs have been used as redox-active additives in solution for the electrochemical detection of sugars and derivatives. PBA-modified electrodes have also been constructed as reagentless electrochemical sensors, where PBAs are immobilized on the surface of metal and carbon electrodes through mainly two routes: as a self-assembled monolayer film and as a polymer thin film. PBA-modified electrodes can be successfully used to detect sugars and derivatives through potentiometric and voltammetric responses. In addition, PBA-modified electrodes can be used for the immobilization of glycoenzymes on an electrode surface by the formation of boronate esters with carbohydrate chains in the glycoenzymes, thus resulting in enzyme biosensors. For the development of PBA-based optical sensors, a variety of chromophores and fluorophores have been coupled with PBA. Azobenzene dyes have been most frequently used for the preparation of colorimetric sugar sensors, in which the absorption wavelength and intensity of the dye are dependent on the type and concentration of added sugars. The sensitivity of the sensors is significantly improved based on multi-component systems in which alizalin red S, pyrocatechol violet, starch–iodine complex, and cyclodextrin are employed as indicators. Anthracene, pyranine, fluorescein, and rhodamine dyes have been used as fluorophores for fluorescence sensors. These dyes have been used in solution or immobilized in films, hydrogels, nanospheres, and quantum dots (QDs) to enhance the sensitivity. QDs-based sensors have been successfully applied for continuous monitoring of glucose in cells. Holographic glucose sensors have also been developed by combining PBA-immobilized hydrogels and photonic crystal colloidal arrays.