Telemetric electrochemical sensor

A telemetric system was designed and constructed to sense pH and ethanol variation in aqueous solutions. The measured signals were transferred by software digitally and transmitted wirelessly by the telemeter, personal digital assistant (PDA), through the General Packet Radio Service (GPRS) protocol. The pH sensing electrode was designed to measure a chemical potential induced by a proton concentration gradient on the electrode's surface which exhibits internal Donnon diffusion behavior, and a linear relationship between the electrical potential and pH was found. The result shows that the wireless sensing system allowed not only long-term usage and long-distance transmission but also with high accuracy (e.g. S.D. less than +/-2%). The telemetric system can also be modified to measure ethanol concentration in aqueous solution amperometrically. It was found that the sensitivity of that ex situ measurements matched those of in field measurements with negligible deviation, less than 4%.     

A urea electrochemical sensor based on molecularly imprinted chitosan film doping with CdS quantum dots

An improved imprinted film-based electrochemical sensor for urea recognition was developed using CdS quantum dots (QDs) doped chitosan as the functional matrix. The microstructure and composition of the imprinted films depicted by scanning electron microscopy (SEM), attenuated total reflection infrared (ATR-IR), X-ray diffraction (XRD), and electrochemical impedance spectroscopy (EIS) indicated the fabricated feasibility of the nanoparticle doped films via in situ electrodeposition. Differential pulse voltammetric responses under the optimal fabrication conditions showed that the sensitivity of CdS QDs-MIP (molecularly imprinted polymer) electrochemical sensor was enhanced from the favorable electron transfer and magnified surface area of CdS QDs with a short adsorption equilibrium time (7 min), wide linear range (5.0 × 10(-12) to 4.0 × 10(-10) M and 5.0 × 10(-10) to 7.0 × 10(-8) M), and low detection limit (1.0 × 10(-12) M). Meanwhile, the fabricated sensor showed excellent specific recognition to template molecule among the structural similarities and coexistence substances. Furthermore, the proposed sensor was applied to determine the urea in human blood serum samples based on its good reproducibility and stability, and the acceptable recovery implied its feasibility for practical application.     

DNA hybridization electrochemical sensor using conducting polymer

We report the use of poly(thiophen-3-yl-acetic acid 1,3-dioxo-1,3-dihydro-isoindol-2-yl ester (PTAE) for application to electrochemical hybridization sensor. A synthetic route for the thiophen-3-yl-acetic acid 1,3-dioxo-1,3-dihydro-isoindol-2-yl ester (TAE) is described, which is used as a monomer of conducting polymer sensor. A direct chemical substitution of probe oligonucleotide to good leaving group site in the PTAE is carried out on the conducting polymer film. A biological recognition can be monitored by comparison with the electrochemical signal (cyclic voltammogram) of single and double strand state oligonucleotide. The sensitivity of the electrochemical sensor is 0.62 microA/nmole and the detection limit is 1 nmole. The oxidation current of double strand state oligonucleotide is a half of that of single strand, that is corresponding to the decrease of electrochemical activity of conducting polymer with increase of stiffness of side group of the polymer. The oxidation current decreasing ratios of perfect matched and single nucleotide mismatched samples are 52 and 25-30%, respectively. The more decreasing ratio is attributable to the more steric hindrance of single nucleotide mismatched sample.     

An electrochemical sensor for determination of calcium dobesilate based on PoPD/MWNTs composite film modified glassy carbon electrode

A poly-o-phenylenediamine and multi-wall carbon nanotubes composite (PoPD/MWNTs) modified glassy carbon electrode (GCE) was prepared by in situ electropolymerization using an ionic surfactant as the supporting electrolyte. The morphology of the resulting PoPD/MWNTs composite was characterized by TEM and the electrochemical properties of the modified electrode were characterized by cyclic voltammetry. The electrochemical behavior of calcium dobesilate on PoPD/MWNTs modified electrode was also investigated. The large current response of calcium dobesilate on PoPD/MWNTs modified electrode is probably caused by the synergistic effect of the electrocatalytic property of PoPD and MWNTs. The reductive peak current increased linearly with the concentration of calcium dobesilate in the range of 0.1–1.0 μmol/L and 4.0–400 μmol/L by square wave adsorptive stripping voltammetry, respectively. The detection limit (three times the signal blank/slope) was 0.035 μmol/L. The modified electrode could eliminate the interference of dopamine, norepinephrine and epinephrine at 100-, 90- and 70-fold concentration of 1.0 μmol/L calcium dobesilate, respectively. The proposed modified electrode provides a new promising and alternative way to detect calcium dobesilate.     

Disposable DNA electrochemical sensor for hybridization detection

A disposable electrochemical sensor for the detection of short DNA sequences is described. Synthetic single-stranded oligonucleotides have been immobilized onto graphite screen printed electrodes with two procedures, the first involving the binding of avidinbiotinylated oligonucleotide and the second adsorption at a controlled potential. The probes were hybridized with different concentrations of complementary sequences. The formed hybrids on the electrode surface were evaluated by differential pulse voltammetry and chronopotentiometric stripping analysis using daunomycin hydrochloride as indicator of hybridization reaction. The probe immobilization step, the hybridization event and the indicator detection, have been optimized. The DNA sensor obtained by adsorption at a controlled potential was able to detect 1 microgram/ml of target sequence in the buffer solution using chronopotentiometric stripping analysis.     

新型青霉素生物传感器的研究

以青霉素为模板分子,采用溶胶-凝胶法合成分子印迹膜,以浸泡的方法移除印迹分子,制备青霉素分子印迹膜电极。本印迹电极能有效地避免类似物对其测定的干扰。通过循环伏安法研究传感器对青霉素的响应特性,结果表明:富集时间为200 s,在0.1~1.8μg/L质量浓度范围内,青霉素在磷酸缓冲液(PBS)中的电流强度与其浓度呈良好的线性关系。吸附后的膜电极用甲醇洗脱后再生,可以重复利用3次,可以应用到实际检测中。     

Selective electrochemical sensor for folic acid at physiological pH using ultrathin electropolymerized film of functionalized thiadiazole modified glassy carbon electrode

This paper demonstrated the selective determination of folic acid (FA) in the presence of important physiological interferents, ascorbic acid (AA) and uric acid (UA) at physiological pH using electropolymerized film of 5-amino-2-mercapto-1,3,4-thiadiazole (p-AMT) modified glassy carbon (GC) electrode. Bare GC electrode fails to determine the concentration of FA in the presence of AA and UA due to the surface fouling caused by the oxidized products of AA and FA. However, the p-AMT film modified electrode not only separates the voltammetric signals of AA, UA and FA with potential differences of 170 and 410 mV between AA–UA and UA–FA, respectively but also shows higher oxidation current for these analytes. The p-AMT film modified electrode displays an excellent selectivity towards the determination of FA even in the presence of 200-fold AA and 100-fold UA. Using amperometric method, we achieved the lowest detection of 75 nM UA and 100 nM each AA and FA. The amperometric current response was increased linearly with increasing FA concentration in the range of 1.0 × 10⁻⁷–8.0 × 10⁻⁴ M and the detection limit was found to be 2.3 × 10⁻¹⁰ M (S/N = 3). The practical application of the present modified electrode was successfully demonstrated by determining the concentration of FA in human blood serum samples.     

Direct electrochemical sensor for fast reagent-free DNA detection

A novel reagentless direct electrochemical DNA sensor has been developed using ultrathin films of the conducting polymer polypyrrole doped with an oligonucleotide probe. Our goal was to develop a prototype electrochemical DNA sensor for detection of a biowarfare pathogen, variola major virus. The sensor has been optimized for higher specificity and sensitivity. It was possible to detect 1.6 fmol of complementary oligonucleotide target in 0.1 ml in seconds by using chronoamperometry. The sensitivity of the developed sensor is comparable to indirect electrochemical DNA sensors, which use electrochemical labels and reagent-intensive amplification. The developed sensing electrode is reusable, highly stable and suitable for storage in solution or in dry state.     

Hollow microneedle-based sensor for multiplexed transdermal electrochemical sensing

The development of a minimally invasive multiplexed monitoring system for rapid analysis of biologically-relevant molecules could offer individuals suffering from chronic medical conditions facile assessment of their immediate physiological state. Furthermore, it could serve as a research tool for analysis of complex, multifactorial medical conditions. In order for such a multianalyte sensor to be realized, it must be minimally invasive, sampling of interstitial fluid must occur without pain or harm to the user, and analysis must be rapid as well as selective. Initially developed for pain-free drug delivery, microneedles have been used to deliver vaccines and pharmacologic agents (e.g., insulin) through the skin. Since these devices access the interstitial space, microneedles that are integrated with microelectrodes can be used as transdermal electrochemical sensors. Selective detection of glucose, glutamate, lactate, hydrogen peroxide, and ascorbic acid has been demonstrated using integrated microneedle-electrode devices with carbon fibers, modified carbon pastes, and platinum-coated polymer microneedles serving as transducing elements. This microneedle sensor technology has enabled a novel and sophisticated analytical approach for in situ and simultaneous detection of multiple analytes. Multiplexing offers the possibility of monitoring complex microenvironments, which are otherwise difficult to characterize in a rapid and minimally invasive manner. For example, this technology could be utilized for simultaneous monitoring of extracellular levels of, glucose, lactate and pH, which are important metabolic indicators of disease states (e.g., cancer proliferation) and exercise-induced acidosis.     

Electrically addressable diazonium-functionalized antibodies for multianalyte electrochemical sensor applications

The direct electrically addressable deposition of diazonium-modified antibodies is examined for electrochemical immunosensing applications. The immobilized antibodies can be detected by the use of electroactive enzyme tags and nanoparticle-gold labeling. Control over antibody functionalization density and minimal spontaneous grafting of diazonium-antibody adducts is shown. The utility of the technique for a sandwich immunoassay as well as the ability to individually and selectively address closely spaced microelectrodes for multi-target protein detection in an array format is demonstrated.     

Measurement of protein using an electrochemical bi-enzyme sensor.

A screen-printed three-electrode amperometric biosensor for the rapid and quantitative measurement of single protein solutions is described. A membrane immobilised protease preparation of broad specificity was used to digest sample protein liberating free amino acids that were subsequently oxidised at a working electrode by immobilised L-amino acid oxidase (L-AAO). The enzymatically generated hydrogen peroxide was determined amperometrically. The fully optimised device required 30 mU L-AAO and 3.94 U protease and had a limit of detection of 170 microg ml(-1) and linearity of response up to 1 mg ml(-1) for Casilan 90 protein. The analytical performance of the device was comparable to that of a commercially available standard photometric protein test kit and required only a 10 microl volume of sample and a single dilution step. Unlike with photometry, the sensor is able to determine the protein content of turbid samples and hence should find widespread applications. The device was simple to use, low-cost and could be mass-produced, yielding results within 4 min of sample addition with acceptable assay repeatability.     

Label-free electrochemical DNA sensor based on functionalised conducting copolymer

A simple and label-free electrochemical sensor for recognition of the DNA hybridization event was prepared based on a new functionalised conducting copolymer, poly[pyrrole-co-4-(3-pyrrolyl) butanoic acid]. This precursor copolymer can be easily electrodeposited on the electrode surface and shows high electroactivity in an aqueous medium. An amino-substituted oligonucleotide (ODN) probe was covalently grafted onto the surface of the copolymer in a one step procedure and tested on hybridization with complementary ODN segments. The cyclic voltammogram of ODN probe-modified copolymer showed very little change when incubated in presence of non-complementary ODN, while a significant, and reproducible, modification of the voltammogram was observed after addition of complementary ODN. The AC impedance spectrum showed an increased charge transfer resistance (Rct) and double layer capacitance of the sensor film after hybridisation. Sensors with thinner films showed higher sensitivity than thicker films, suggesting that hybridisation at or near the surface of the film produces a larger change in electrical properties than that within the body of the film.     

An electrochemical sensor based on single-stranded DNA-poly(sulfosalicylic acid) composite film for simultaneous determination of adenine, guanine, and thymine

Poly(sulfosalicylic acid) and single-stranded DNA composite (PSSA–ssDNA)-modified glassy carbon electrode (GCE) was prepared by electropolymerization and then successfully used to simultaneously determine adenine (A), guanine (G), and thymine (T). The characterization of electrochemically synthesized PSSA–ssDNA film was investigated by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). The modified electrode exhibited enhanced electrocatalytic behavior and good stability for the simultaneous determination of A, G, and T in 0.1 M phosphate buffer solution (PBS, pH 7.0). Well-separated voltammetric peaks were obtained among A, G, and T presented in the analyte mixture. Under the optimal conditions, the peak currents for A, G, and T increased linearly with the increase of analyte mixture concentration in the ranges of 6.5 × 10⁻⁸ to 1.1 × 10⁻⁶, 6.5 × 10⁻⁸ to 1.1 × 10⁻⁶, and 4.1 × 10⁻⁶ to 2.7 × 10⁻⁵ M, respectively. The detection limits (signal/noise = 3) for A, G, and T were 2.2 × 10^–8, 2.2 × 10^–8, and 1.4 × 10^–6 M, respectively.     

Label-free DNA sensor based on organic thin film transistors

Organic thin film transistors (OTFTs) are excellent candidates for the application on disposable sensors due to their potentially low-cost fabrication process. A novel DNA sensor based on OTFTs with semiconducting polymer poly(3-hexylthiophene) has been fabricated by solution process. Both single- and double-strand DNA molecules are immobilized on the surface of the Au source/drain electrodes of different OTFT devices, producing a dramatic change in the performance of the devices, which is attributed to the increase of the contact resistances at the source/drain electrodes. Single-strand DNA and double-strand DNA are differentiated successfully in the experiments indicating that this is a promising technique for sensing DNA hybridization without labelling.     

DNA hybridization sensor based on pentacene thin film transistor

A DNA hybridization sensor using pentacene thin film transistors (TFTs) is an excellent candidate for disposable sensor applications due to their low-cost fabrication process and fast detection. We fabricated pentacene TFTs on glass substrate for the sensing of DNA hybridization. The ss-DNA (polyA/polyT) or ds-DNA (polyA/polyT hybrid) were immobilized directly on the surface of the pentacene, producing a dramatic change in the electrical properties of the devices. The electrical characteristics of devices were studied as a function of DNA immobilization, single-stranded vs. double-stranded DNA, DNA length and concentration. The TFT device was further tested for detection of λ-phage genomic DNA using probe hybridization. Based on these results, we propose that a "label-free" detection technique for DNA hybridization is possible through direct measurement of electrical properties of DNA-immobilized pentacene TFTs.     

DNA entrapped polypyrrole-polyvinyl sulfonate film for application to electrochemical biosensor

Double-stranded calf thymus (dsCT)-DNA was electrochemically entrapped into polypyrrole-polyvinyl sulfonate (PPy-PVS) films deposited onto indium tin oxide (ITO) coated glass plates. These dsCT-DNA entrapped PPy-PVS/ITO films were characterized using cyclic voltammetry, UV-visible, Fourier transform infrared (FT-IR), scanning tunneling microscopy (STM), and electrochemical impedance measurements. Attempts made to use these dsCT-DNA entrapped PPy-PVS/ITO films for detection of 2-aminoanthracene (0.001-6.0 ppm) and 3-chlorophenol (0.01-55.0 ppm) revealed a response time of 30s and a shelf life of approximately 25 weeks when stored under desiccated conditions at 25 degrees C. The addition of salts such as Ca(2+) (250 ppm), Mg(2+) (200 ppm), Cl(-) (1560 ppm), and Na(+) (150 ppm) ions contained in water does not affect the observed amperometric response of the disposable dsCT-DNA entrapped PPy-PVS film-based electrochemical biosensor.     

Ligase-based multiple DNA analysis by using an electrochemical sensor array

We herein report an electrochemical biosensor for the sequence-specific detection of DNA with high discrimination ability for single-nucleotide polymorphisms (SNPs). This DNA sensor was constructed by a pair of flanking probes that "sandwiched" the target. A 16-electrode electrochemical sensor array was employed, each having one individual DNA capture probe immobilized at gold electrodes via gold-thiol chemistry. By coupling with a biotin-tagged detection probe, we were able to detect multiple DNA targets with a single array. In order to realize SNP detection, a ligase-based approach was employed. In this method, both the capture probe and the detection probe were in tandem upon being hybridized with the target. Importantly, we employed a ligase that specifically could ligate tandem sequences only in the absence of mismatches. As a result, when both probes were complementary to the target, they were ligated in the presence of the ligase, thus being retained at the surface during the subsequent stringent washing steps. In contrast, if there existed 1-base mismatch, which could be efficiently recognized by the ligase, the detection probe was not ligated and subsequently washed away. A conjugate of avidin-horseradish peroxidase was then attached to the biotin label at the end of the detection probe via the biotin-avidin bridge. We then electrochemically interrogated the electrical current for the peroxidase-catalyzed reduction of hydrogen peroxide. We demonstrated that the electrochemical signal for the wild-type DNA was significantly larger than that for the sequence harboring the SNP.     

Aerobic batch cultivation in micro bioreactor with integrated electrochemical sensor array

Aerobic batch cultivations of Candida utilis were carried out in two micro bioreactors with a working volume of 100 μL operated in parallel. The dimensions of the micro bioreactors were similar as the wells in a 96‐well microtiter plate, to preserve compatibility with the current high‐throughput cultivation systems. Each micro bioreactor was equipped with an electrochemical sensor array for the online measurement of temperature, pH, dissolved oxygen, and viable biomass concentration. Furthermore, the CO₂ production rate was obtained from the online measurement of cumulative CO₂ production during the cultivation. The online data obtained by the sensor array and the CO₂ production measurements appeared to be very reproducible for all batch cultivations performed and were highly comparable to measurement results obtained during a similar aerobic batch cultivation carried out in a conventional 4L bench‐scale bioreactor. Although the sensor chip certainly needs further improvement on some points, this work clearly shows the applicability of electrochemical sensor arrays for the monitoring of parallel micro‐scale fermentations, e.g. using the 96‐well microtiterplate format. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

An ultrathin platinum film sensor to measure biomolecular binding.

A sensitive conductimetric immunosensor has been demonstrated based on an ultrathin platinum film on an oxidized silicon base. The film is about 25 A thick and is seen to consist of a discontinuous layer with channels 20-30 A wide. Monoclonal antibodies were bound to the sensor surface using conventional biosensor chemistry. Impedance at fixed frequencies across the film was used to track modification and binding at the surface. Impedance increased 55% at 20 Hz during the activation of the surface with anti-alkaline phosphatase (anti-AP). Binding of alkaline phosphatase (AP) to the prepared surface results in a further increase of 12%. p-Nitrophenyl phosphate hydrolysis confirmed binding and activity of the AP. About 40 amol AP were bound on the 0.5 cm(2) electrode. Non-specific binding of horseradish peroxidase caused an impedance change <6%. Control experiments showed small impedance changes and trace enzyme activity. Since the mechanism of electrical conduction of the thin film was not established, modeling of thin-film response was used to distinguish between redox processes, capacitance and tunneling mechanisms. The data fit well with the diffusion distributed elements (DE) model as well as a transmission line distribution element (DX) model. The first model, DE, is distributed elements for diffusion. The second DX model represents a transmission line. The sensors behave in a distributed network or like a transmission line.