Novel heteroarene oligomers, consisting of two pyridinium groups, linked by thiophene units of variable length, thienoviologens, are described as promising candidates for molecular wires. Two representative thienoviologens were coated by adsorption from micromolar concentrations in ethanol onto octadecylmercaptan (ODM)-coated gold electrodes and induced a gradual restoration of the electrochemistry with hexacyanoferrate as a function of molecular wire concentration. Glucose oxidase and choline oxidase showed strong adsorption to these conductive layers, but showed striking differences in adsorption to the different thienoviologen layers. The measurements support the hypothesis that the molecules are incorporated in the ODM layer in a different fashion. Also the complex formation of an engineered azurin redox protein with water-soluble pyridyl ligands is presented in relation to a possible application of the thienoviologens as conductive spacers, in which the contact with the redox protein is achieved via complex formation with a free pyridine nitrogen. (C) 1997 Elsevier Science S.A.
The sixth workshop of the European Community Concerted Action on Chemical sensors for in vivo monitoring was held at Snogeholm, Sweden, in October 1991. The meeting reviewed recent in vivo and ex vivo results, and also included a shorter session on ethical and safety problems.
The latest workshop of the European Community (EC) Concerted Action on Chemical sensors for in vivo monitoring was held in Nauplion, Greece, in April this year. This fifth workshop focused on The design and development of new sensors for in vivo monitoring, and was organized into five sessions: design and development of new sensors; operational considerations; performance of analytical systems; novel sensors/tissue heterogeneity; and infra-red spectroscopy.
Considerable progress has been made in the development of practical glucose sensors for in vivo use. In some designs, the sensor can be replaced by the patient, who "injects" a new sensor into a fresh site using a hypodermic needle. An open-loop hypoglycaemia sensor providing an alarm signal could be available soon, and it would be very useful in the management of diabetes. The sensor for this application would need to function for a minimum period of about 12 hours. Further developments in the glucose sensor area are expected to lead to a comfortable implantable sensor/pump device, which would provide closed loop control for a prolonged period. An acceptable system would be a sensor replaced every 2 to 3 days, coupled with a microprocessor-controlled pump replaced after 2 to 3 years. Further engineering input is needed to design the monitoring and control unit containing the alarm and pump systems. The world market in clinical analysis is about US $6 billion, and covers about 10% of the total expenses for health care. It is in the field of continuous microsensors that future competition for the clinical diagnostic market will take place. Funding in vivo projects is less attractive to industry than the more proven and well-established area of in vitro diagnostics. However, the benefits of in vivo monitoring to the health of the community, and the financial rewards to manufacturers are ultimately great. It is clearly of interest for industry to invest in the development and commercialisation of this high added-value technology.
A highly-sensitive glucose biosensor amenable to ultra-miniaturisation was fabricated by immobilisation of glucose oxidase (GOx), onto a poly(2,6-diaminopyridine)/multi-walled carbon nanotube/glassy carbon electrode (poly(2,6-DP)/MWNT/GCE). Cyclic voltammetry was used for both the electrochemical synthesis of poly-(2,6-DP) on the surface of a MWNT-modified GC electrode, and characterisation of the polymers deposited on the GC electrode. The synergistic effect of the high active surface area of both the conducting polymer, i.e., poly-(2,6-DP) and MWNT gave rise to a remarkable improvement in the electrocatalytic properties of the biosensor. The transfer coefficient (alpha), heterogeneous electron transfer rate constant and Michaelis-Menten constant were calculated to be 0.6, 4 s(-1) and 0.20 mM at pH 7.4, respectively. The GOx/poly(2,6-DP)/MWNT/GC bioelectrode exhibited two linear responses to glucose in the concentration ranging from 0.42 mu M to 8.0 mM with a correlation coefficient of 0.95, sensitivity of 52.0 mu AmM-1 cm(-2), repeatability of 1.6% and long-term stability, which could make it a promising bioelectrode for precise detection of glucose in the biological samples. (C) 2013 Elsevier B.V. All rights reserved.
A highly-sensitive glucose biosensor amenable to ultraminiaturisation was fabricated by immobilization of glucose oxidase (wGOX), onto a poly(2,6-diaminopyridine)/multi-walled carbon nanotube/glassy carbon electrode (poly(2,6-DP)/MWCNT/GCE). Cyclic voltammetry was used for both the electrochemical synthesis of poly-(2,6-DP) on the surface of a MWCNT-modified GC electrode, and characterization of the polymers deposited on the GC electrode. The synergistic effect of the high active surface area of both the conducting-polymer, i.e., poly-(2,6-DP) and MWCNT gave rise to a remarkable improvement in the electrocatalytic properties of the biosensor. The transfer coefficient (alpha), heterogeneous electron transfer rate constant and Michaelis-Menten constant were calculated to be 0.6, 4 s-1 and 0.22 mM at pH 7.4, respectively. The GOx/poly(2,6-DP)/MWCNT/GC bioelectrode exhibited two linear responses to glucose in the concentration ranging from 0.42 mu M to 8.0 mM with a correlation coefficient of 0.95, sensitivity of 52.0 mu AmM-1 cm-2, repeatability of 1.6% and long-term stability, which could make it a promising bioelectrode for precise detection of glucose in the biological samples. (C) 2016 Elsevier B.V. All rights reserved.
In this paper, a simple and sensitive approach for human epidermal growth factor receptor 2 (HER2) detection is presented, using antibody-functionalised magnetic beads coupled to screen-printed cells. The immunoassay is based on a sandwich format in which a primary monoclonal antibody anti-HER2 is coupled to protein A modified magnetic beads. The modified beads are then used to capture the protein from the sample solution and a sandwich assay is performed by adding a secondary monoclonal antibody anti-HER2 labelled with biotin. The enzyme alkaline phosphatase (AP) conjugated with streptavidin and its substrate (1-naphthyl-phosphate) are then used for the electrochemical detection by differential pulse voltammetry (DPV). The experimental conditions for the immunoassay were optimised. The performance of the assay in terms of sensitivity, reproducibility and selectivity has been studied in buffer and serum samples from hospital patients.
Regeneration is a key goal in the design of immunosensors. In this study, we report the temperature-regulated interaction of N-isopropylacrylamide (PNIPAAm) functionalised cardiac troponin T (cTnT) with anti-cTnT. Covalently bonded PNIPAAm on an anti-cTnT bioelectrode showed on/off-switchability, regeneration capacity and temperature triggered sensitivity for cTnT. Above the lower critical solution temperature (LCST), PNIPAAm provides a liphophilic microenvironment with specific volume reduction at the bioelectrode surface, making available binding space for cTnT, and facilitating analyte recognition. Computational studies provide details about the structural changes occurring at the electrode above and below the LCST. Furthermore, free energies associated with the binding of cTnT with PNIPAAm at 25 (Delta G(coil)=-6.0 Kcal/mole) and 37 degrees C (Delta G(globular)=-41.0 kcal/mole) were calculated to elucidate the interaction and stability of the antigen-antibody complex. The responsiveness of such assemblies opens the way for miniaturised, smart immuno-technologies with built-in programmable interactions of antigen-antibody upon receiving stimuli.
Regeneration of immunosensors is a longstanding challenge. We have developed a re-usable troponin-T (TnT) immunoassay based on localised surface plasmon resonance (LSPR) at gold nanorods (GNR). Thermosensitive poly(N-isopropylacrylamide) (PNIPAAM) was functionalised with anti-TnT to control the affinity interaction with TnT. The LSPR was extremely sensitive to the dielectric constant of the surrounding medium as modulated by antigen binding after 20 min incubation at 37 degrees C. Computational modelling incorporating molecular docking, molecular dynamics and free energy calculations was used to elucidate the interactions between the various subsystems namely, IgG-antibody (c. f., anti-TnT), PNIPAAM and/or TnT. This study demonstrates a remarkable temperature dependent immuno-interaction due to changes in the PNIPAAM secondary structures, i.e., globular and coil, at above or below the lower critical solution temperature (LCST). A series of concentrations of TnT were measured by correlating the lambda(LSPR) shift with relative changes in extinction intensity at the distinct plasmonic maximum (i. e., 832 nm). The magnitude of the red shift in lambda(LSPR) was nearly linear with increasing concentration of TnT, over the range 7.6 x 10(-15) to 9.1 x 10(-4) g/mL. The LSPR based nano-immunoassay could be simply regenerated by switching the polymer conformation and creating a gradient of microenvironments between the two states with a modest change in temperature.
n/a
Current electrochemical biosensors for multiple miRNAs require tedious immobilization of various nucleic acid probes. Here, we demonstrate an innovative approach using a generic neutravidin biosensor combined with electrochemically encoded responsive nanolabels for facile and simultaneous multiplexed detection of miRNA-21 and miRNA-141. The selectivity of the biosensor arises from the intrinsic properties of the electrochemically encoded responsive nanolabels, comprising biotinylated molecular beacons (biotin-MB) and metal nanoparticles (metal-NPs). The procedure is a simple one-pot assay, where the targeted miRNA causes the opening of biotin-MB followed by capturing of the biotin-MB-metal-NPs by the neutravidin biosensor and simultaneous detection of the captured metal-NPs by stripping square-wave voltammetry (SSWV). The multiplexed detection of miRNA-21 and miRNA-141 is achieved by differentiation of the electrochemical signature (i.e., the peak current) for the different metal-NP labels. The biosensor delivers simultaneous detection of miRNAs with a linear range of 0.5-1000 pM for miRNA-21 and a limit of detection of 0.3 pM (3 sigma/sensitivity, n = 3), and a range of 50-1000 pM for miRNA-141, with a limit of detection of 10 pM. Furthermore, we demonstrate multiplexed detection of miRNA-21 and miRNA-141 in a spiked serum sample.
Alteration in expression of miRNAs has been correlated with different cancer types, tumour stage and response to treatments. In this context, a structurally responsive oligonucleotide-based electrochemical impedimetric biosensor has been developed for the simple and sensitive detection of miRNA-21. A highly specific biotinylated DNA/LNA molecular beacon (MB) probe was conjugated with gold nanoparticles (AuNPs) to create an integrated, dual function bio-label (biotin-MB-AuNPs) for both biorecognition and signal generation. In the presence of target miRNA-21, hybridisation takes place resulting in the "activation" of the biotin-MB; this event makes the biotin group, which was previously "protected" by the steric hindrance of the MB stem-loop structure, accessible. The activated biotin-MB-AuNPs/miRNA complexes become available for capture, via supramolecular interaction, onto a nentravidin-modified electrode for electrochemical transduction. The binding event results in a decrease of the charge transfer resistance at the working electrode/electrolyte interface. The biosensor responded linearly in the range 1-1000 pM of miRNA-21, with a limit of detection of 0.3 pM, good reproducibility (Relative Standard deviation (RSD) =3.3%) and high selectivity over other miRNAs (i.e. miRNA221 and miRNA-205) sequences. Detection of miRNA-21 in spiked serum samples at clinically relevant levels (low pM range) was also demonstrated, thus illustrating the potential of the biosensor for point-of-care clinical applications. The proposed biosensor design, based on the combination of a neutravidin transducing surface and the dual-function biotin-MB-AuNPs bio-label, provides a simple and robust approach for detection of short-length nucleic acid targets, such as miRNAs.
The extremely specialised anatomical function of citrate inside the prostate, make it one of the preferred biomarkers for early stage detection of prostate cancer. However, current detection methods are seriously limited due to the very low citrate concentrations that need to be measured in order to follow disease progression. In the present work, we report a novel citrate-selective-sensor based on iron (III) phthalocyanine chloride-C-monoamido-Poly-n-Butyl Acrylate (Fe(III)MAPcC1 P n BA) modified gold -electrodes for the electrochemical determination and estimation of the pathophysiological range of citrate. The newly synthesised ionophore has been structurally characterised using Fourier transform infrared (FTIR) and UV-vis spectroscopy. Contact angle measurements and atomic force microscopy (AFM) have been used to investigate the adhesion and morphological properties of the membrane. The developed citrate-selective-electrodes had a Nernstian sensitivity of-19.34 +/- 0.83 mV/decade with a detection limit of about 9 x 10-6M and a linear range from 4 x 10(-5)M to 10(-1) M, which covered the pathologically important clinical range. Electrochemical impedance spectroscopy (EIS) showed very high sensitivity with a lower Limit of detection 1.7 x 10(-9) M and linear detection range (10(-8)-10(-1) M), which is very important not only for the early-stage diagnosis and screening procedures, but also in mapping the stage of the cancer too. (C) 2016 Elsevier B.V. All rights reserved.
Fast and accurate detection of microorganisms is of key importance in clinical analysis and in food and water quality monitoring. Salmonella typhimurium is responsible for about a third of all cases of food borne diseases and consequently, its fast detection is of great importance for ensuring the safety of foodstuffs. We report the development of a label-free impedimetric aptamer-based biosensor for S. typhimurium detection. The aptamer biosensor was fabricated by grafting a diazonium-supporting layer onto screen printed carbon electrodes (SPEs), via electrochemical or chemical approaches, followed by chemical immobilisation of aminated-aptamer. FTIR-ATR, contact angle and electrochemical measurements were used to monitor the fabrication process. Results showed that electrochemical immobilisation of the diazonium-grafting layer allowed the formation of a denser aptamer layer, which resulted in higher sensitivity. The developed aptamer-biosensor responded linearly, on a logarithm scale, over the concentration range 1 x 10(1) to 1 x 10(8) CFU mL(-1), with a limit of quantification (LOQ) of 1 x 10(1) CFU mL(-1) and a limit of detection (LOD) of 6 CFU mL(-1). Selectivity studies showed that the aptamer biosensor could discriminate S. typhimurium from 6 other model bacteria strains. Finally, recovery studies demonstrated its suitability for the detection of S. typhimurium in spiked (1 x 10(2), 1 x 10(4) and 1 x 10(6) CFU mL(-1)) apple juice samples. (C) 2016 Elsevier B.V. All rights reserved.
Glucose-6-phosphateplays an important role in carbohydrate metabolism of all living organisms.Compared to the conventional analytical methods available for estimation of glucose-6-phosphate,the biosensors having relative simplicity, specificity, low-cost and fastresponse time are a promising alternative. We have reported a glucose-6-phosphatesensor based on screen-printed electrode utilizing Prussian blue nanoparticlesand enzymes, glucose-6-phosphate dehydrogenase and glutathione reductase. The Prussianblue nanoparticles acted as a mediator enhancing the rate of electrochemical responses.The Fourier transforminfrared spectroscopy and energy-dispersiveX-ray spectroscopy study confirmed the formation of Prussian blue, whereas, the atomic forced microscopy revealed that Prussian bluenanoparticles were about 25-30 nm in diameter. To obtainmaximum amperometric response, optimization studies were conducted for pH,enzyme and cofactor loading. The proposed glucose-6-phosphate biosensor showed goodstability, rapid response time and broad linear response in the range of 0.01-1.25mM and detection limit of 6.3 mM. The biosensor also worked well for serum samples and exhibitedexcellent anti-interference ability.
Mobile diagnostics for healthcare, food safety and environmental monitoring, demand a new generation of inexpensive sensing systems suitable for production in high volume. Herein we report on the development of a new disposable electrochemical instrument exploiting the latest advances in printed electronics and printed biosensors. The current system is manufactured under ambient conditions with all interconnections printed; electrochemical measurements and data elaboration are realized by the integration onto the platform of two chips: a MICROCHIP-PIC24F16KA101 and a Texas Instruments LMP91000. A PEDOT.PSS vertical electrochromic display (VECD) is also incorporated into the system to visualize the data. A printed Enfucell 3V manganese dioxide battery was used to deliver the required power. Finally, in order to demonstrate the utility of the system, screen-printed sensors for the detection of glucose were added and the performance of the overall system was evaluated.
This chapter contains sections titled:
Introduction
Carbon Nanotubes
Conductive Polymer Nanostructures
Nanoparticles
Conclusions
References
In this work we explored a simple, cheap and fast route to grow polyaniline (PANI) nanotubes arranged in an ordered structure directly on an electrode surface by electrochemical polymerisation. The deposited nanostructures were electrochemically and morphologically characterised and then used as a functional substrate for biochemical sensing by combining the intrinsic advantages of nanostructures as optimal transducers and the well known benefits of molecularly imprinted polymers (MIPs) as receptors. The hybrid nanostructured-MIP sensor was applied to the molecular recognition of catechol. Moreover, a gas sensing application was also investigated by exploiting resistance variation of the polymer in presence of different gases (CO, NO2, NH3 and ethanol).
Recent advances in nanotechnology have allowed significant progress in utilising cutting-edge techniques associated with nanomaterials and nano-fabrication to expand the scope and capability of biosensors to a new level of novelty and functionality. The aim of this work was the development and characterisation of conductive polyaniline (PANI) nanostructures for applications in electrochemical biosensing. We explore a simple, inexpensive and fast route to grow PANI nanotubes, arranged in an ordered structure directly on an electrode surface, by electrochemical polymerisation using alumina nanoporous membranes as a nano-mould. The deposited nanostructures have been characterised electrochemically and morphologically prior to grafting with a molecularly imprinted polymer (MIP) receptor in order to create a model sensor for catechol detection. In this way, PANI nanostructures resulted in a conductive nanowire system which allowed direct electrical connection between the electrode and the synthetic receptor (MIP). To our knowledge, this is the first example of integration between molecularly imprinted polymers and PANI nanostructured electrodes. The advantages of using nanostructures in this particular biosensing application have been evaluated by comparing the analytical performance of the sensor with an analogous non-nanostructured MIP-sensor for catechol detection that was previously developed. A significantly lower limit of detection for catechol has been obtained (29 nM, one order of magnitude), thus demonstrating that the nanostructures are capable of improving the analytical performance of the sensor. (C) 2010 Elsevier B.V. All rights reserved.
The antioxidant reduced glutathione tripeptide is conventionally called glutathione (GSH). The oxidized form is a sulfur-sulfur linked compound. known as glutathione disulfide (GSSG). Glutathione is an essential cofactor for antioxidant enzymes. it provides protection also for the mitochondria against endogenous oxygen radicals. The ratio of these two forms can act as a marker for oxidative stress. The majority of the methods available for estimation of both the forms of glutathione are based on colorimetric and electrochemical assays. In this study, electrochemical sensors were developed for the estimation of both GSH and GSSG. Two different types of transducers were used: i) screen-printed three-electrode disposable sensor (SPE) containing carbon working electrode, carbon counter electrode and silver/silver chloride reference electrode; ii) three-electrode disposable system (CDE) consisting of three copper electrodes. 5.5-dithlobis(2-nitrobenzoic acid) (DTNB) was used as detector element for estimation of total reduced thiol content. The enzyme glutathione reductase along with a co-enzyme reduced nicotinamide adenine dinucleotide phosphate was used to estimate GSSG. By combining the two methods GSH can also be estimated. The detector elements were immobilized on the working electrodes of the sensors by bulk polymerization of acrylamide. The responses were observed amperometrically. The detection limit for thiol (GSH) was less than 0.6 ppm when DTNB was used. whereas for GSSG it was less than 0.1 ppm.
Pure enzymes are costly and highly sensitive to change in pH, temperature, ionic strength etc. Hence biomimetic or synthetic enzymes could be useful alternatives to such natural proteins. Although the selectivity of a biomimic is somewhat less than that of enzyme, it can be used as a detector element in inexpensive but stable biosensors. An organic compound, 4-[(1E)-ethanehydrazonoyl]benzoic acid, has been designed and synthesized as biomimic for the enzyme acetylcholine esterase. An acetylcholine chloride two-electrode screen-printed sensor was first developed using the immobilized enzyme acetylcholine esterase. The performance of the mimic in the hydrolysis of acetylcholine chloride was then tested with the same transducer by immobilizing the biomimic in place of the enzyme. The response of the sensor constructed using the mimic was comparable to that of the pure acetylcholine esterase enzyme electrode.
Amperometric biosensors for hydrogen peroxide were produced, based on TTF-modified graphite disc electrodes, incorporating immobilised peroxidase. At the optimum operating potential (-200 mV vs. calomel electrode), a Linear response to hydrogen peroxide was obtained in the range 0-2 mM (Km value = 1.98 mM). Low sensitivity to pH fluctuations and good stability at room temperature were exhibited.
The possibility of introducing a computationally assisted method to study aptamer-protein interaction was evaluated with the aim of streamlining the screening and selection of new aptamers. Starting from information on the 15-mer (5-GGTTGGTGTGGTTGG-3 thrombin binding aptamer (TBA), a library of mutated DNA sequences (994 elements) was generated and screened using shapegauss a shape-based scoring function from openeye software to generate computationally derived binding scores. The TBA and three other mutated oligonucleotides, selected on the basis of their binding score (best, medium, worst), were incorporated into surface plasmon resonance (SPR) biosensors. By reducing the ionic strength (binding buffer, 50 mMTrisHC1pH 7.4, 140 mM NaCl, 1 mM MgCl(2), diluted 1:50) in order to match the simulated condition, the analytical performances of the four oligonucleotide sequences were compared using signal amplitude, sensitivity (slope), linearity (R(2)) and reproducibility (CVav %). The experimental results were in agreement with the simulation findings.
The synthesis of poly-aminophenylboronic acid (ABPA) imprinted beads for the recognition of the protein human serum albumin (HSA) is reported. In order to create homogeneous recognition sites, covalent immobilisation of the template HSA was exploited. The resulting imprinted beads were selective for HSA. The indirect imprinting factor (IF) calculated from supernatant was 1.6 and the direct IF, evaluated from the protein recovered from the beads, was 1.9. The binding capacity was 1.4 mg/g, which is comparable to commercially available affinity materials. The specificity of the HSA recognition was evaluated with competitive experiments, indicating a molar ratio 4.5/1 of competitor was necessary to displace half of the bound HSA. The recognition and binding of the imprinted beads was also tested with a complex sample, human serum and targeted removal of HSA without a loss of the other protein components was demonstrated. The easy preparation protocol of derivatised beads and a good protein recognition properties make the approach an attractive solution to analytical and bio-analytical problems in the field of biotechnology. (c) 2007 Elsevier B.V. All rights reserved.
Molecular imprinting has proved to be an effective technique for the creation of recognition sites on a polymer scaffold. Protein imprinting has been a focus for many chemists working in the area of molecular recognition, since the creation of synthetic polymers that can specifically recognise proteins is a very challenging but potentially extremely rewarding objective. It is expected that molecularly imprinted polymers (MIPs) with specificity for proteins will find application in medicine, diagnostics, proteomics, environmental analysis, sensors and drug delivery. In this review, the authors provide an overview of the progress achieved in the decade between 1994 and 2005, with respect to the challenging area of MIPs for protein recognition. The discussion furnishes a comparative analysis of different approaches developed, underlining their relative advantages and disadvantages and highlighting trends and possible future directions. (c) 2006 Elsevier B.V. All rights reserved.
Extending the previous preliminary study on the construction of a capillary electrophoresis (CE)/sensor for the detection of reducing analytes, we focus the interest on the simultaneous detection of redox active species, which are important indicators of the oxidative damage in tissues, of food preservation, and of pollution. The CE/sensor was built by modifying the detector-portion of the capillary with the redox-sensitive polymer polyaniline (PANI). The analyte is detected by monitoring the changes in optical absorption of the PANI film. The CE/sensor was tested, with good results, with ascorbic acid, glutathione (GSH), as well as with compounds with very close similarity (ascorbic and isoascorbic acid). The kinetics of oxidation and reduction of PANI were evaluated. Further a PANI/CE-biological sensor was developed by coupling an enzyme, glucose oxidase (GOD), to the PANI-modified portion of the capillary. The stability of the immobilized GOD and the sensitivity of the CE/biosensor were studied, by using glucose as test analyte in concentrations within the physiological range. The results indicate that the CE/biosensor had good stability (more than 75% of original activity retained after 30 operational days), manufacturing reproducibility and a sensing range convenient for monitoring physiological glucose (1-24 mm).
A technique for modification of the microtiter reader plates well with a polyaniline (PANI) film sensitive for ascorbic acid is presented. The principle of the analyte detection is based on monitoring the changes in optical absorption of the PANI film resulting from the reduction process initiated by ascorbic acid. The detection limit for ascorbic acid is 1 mg/L. Testing with real samples (soft drinks, fruit juices) gave good correlation of the method with iodimetric titration. High sensitivity, stability, and good reproducibility of the measurements make the proposed system an attractive alternative to traditional assays, used in medicine, ecology and biotechnology.
A technique for coating microplate wells with molecularly imprinted polymers (MIPs) specific for proteins is presented. 3-Aminophenylboronic acid was polymerized in the presence of the following templates: microperoxidase, horseradish peroxidase, lactoperoxidase, and hemoglobin, via oxidation of the monomer by ammonium persulfate. This process resulted in the grafting of a thin polymer layer to the polystyrene surface of the microplates. Imprinting resulted in an increased affinity of the polymer toward the corresponding templates. The influence of the washing procedure, template concentration, and buffer pH on the polymer affinity was analyzed. It was shown that the stabilizing function of the support and spatial orientation of the polymer chains and template functional groups are the major factors affecting the imprint formation and template recognition. Easy preparation of the MIPs, their high stability, and their ability to recognize small and large proteins, as well as to discriminate molecules with small variations in charge, make this approach attractive and broadly applicable in biotechnology, assays and sensors.
The present review highlights some modern aspects of biosensor revelation, a detection method which has already found a large number of applications in healthcare, food industry and environmental analysis. First, the concept of bio-recognition, which is at the heart of biosensor technology, is discussed, with emphasis on host-guest-like recognition mechanisms. This detection device has been successfully coupled, in its first applications, to chromatographic columns, which allow a high resolution of complex mixtures of analytes prior to interaction with the biosensing unit. The properties of the transducing elements, which should generate a signal (e.g., electrochemical, thermal, acoustic, optical) of proper intensity and of relative fast rise, are additionally evaluated and discussed. The review then focuses on potential applications of biosensing units in capillary electrophoresis (CE) devices. CE appears to be an excellent separation methodology to be coupled to biosensor detection, since it is based on miniaturized electrophoretic chambers, fast analysis times, complete automation in sample handling and data treatment and requires extremely small sample volumes. Although only a few applications of CE-based biosensors have been described up to the present, it is anticipated that this hyphenated technique could have a considerable expansion in the coming years, (C) 2000 Elsevier Science B.V. All rights reserved.
The capillary walls of fused-silica capillary electrophoresis (CE) columns were modified with a thin film of polyaniline (PANI), providing open-tubular columns with a stable coating containing aromatic groups and amine functionalities. Fast and efficient separations were observed for small bioactive peptides under acidic conditions on PANI-coated columns. The mechanism of separation is based on hydrophobic interactions between the analytes and the polymeric matrix. Good reproducibility was observed from run-to-run. Due to the simple derivatization procedure, method flexibility, the uniformity of the coating and its stability, conjugated polymers could find practical application in capillary zone electrophoresis (CZE) separations.
Microfluidic devices find many applications in biotechnologies. Here, we introduce a flexible and biocompatible microfluidic ice-based platform with tunable parameters and configuration of microfluidic patterns that can be changed multiple times during experiments. Freezing and melting of cavities, channels and complex relief structures created and maintained in the bulk of ice by continuous scanning of an infrared laser beam are used as a valve action in microfluidic systems. We demonstrate that pre-concentration of samples and transport of ions and dyes through the open channels created can be achieved in ice microfluidic patterns by IR laser-assisted zone melting. The proposed approach can be useful for performing separation and sensing processes in flexible reconfigurable microfluidic devices.
The polymerisation of 3-aminophenylboronic acid (APBA) in aqueous environment has been used for the open tubular modification of capillary electrophoresis (CE) capillaries. Being poly-APBA endowed with boronic acid, aromatic rings and secondary amines groups, it posses a variety of functional groups affecting selectivity. Diastereoisomers (e.g. ascorbic and isoascorbic acid) and proteins (e.g. haemoglobins) were successfully separated onto poly-APBA column, by means of a combination of electrophoresis and open tubular electrochromatography. The mechanism of selection was investigated: results indicate an interplay between enhancing or silencing the contribution of the protonable functionahties (amino groups, boronic acid). The properties of APBA polymer coating make it attractive for CE separation and for further application in affinity separations and chip technologies.
An entirely new means of printing molecular information on a planar film, involving casting nanoscale impressions of the template protein molecules in molten gallium, is presented here for the first time. The metallic imprints not only replicate the shape and size of the proteins used as template. They also show specific binding for the template species. Such a simple approach to the creation of antibody-like properties in metallic mirrors can lead to applications in separations, microfluidic devices, and the development of new optical and electronic sensors, and will be of interest to chemists, materials scientists, analytical specialists, and electronic engineers.
This contribution considers the origins, principles and recent literature published on enzymatic biofuel cells, with a focus on performance and stability. Modified or new biofuel cell components, such as modified electrodes, new enzymes and the use of new mediators to improve power output and stability are reviewed. The development of biofuel cells to date leaves huge potential for further improvement and practical application. Cooperation between different fields of science is essential to realise important potential applications in human health and power generation; future research needs to achieve this are discussed.
Biosensors have been extensively studied during the last 20 years, and a myriad of laboratory biosensors have been developed. Improvements are required in biosensor design and performance before they become widely accepted in industrial process monitoring. However, as the biotechnology industry expands, biosensors may become more acceptable because, despite their limitations, they are the only devices capable of delivering the information required.
A disposable choline biosensor based on ruthenized carbon screen-printed electrodes has been prepared and its use for monitoring organophosphorus pesticides and carbamates is described. The presence of 0.5% ruthenium on activated carbon mixed to form a simple graphite-based ink for the working electrode surface increased the sensitivity towards hydrogen peroxide. The choline biosensor is based on such an electrode coupled with choline oxidase immobilized by adsorption. The inhibition effect of carbamates and organophosphorus pesticides on acetylcholinesterase was detected using this biosensor. With the optimized procedure described (pH, buffer composition, incubation time, substrate concentration) we are able to detect concentrations of pesticides (carbofuran) as low as 1 nM.
Plastic disposable choline biosensors based on ruthenized-carbon screen-printed electrodes were prepared and their use for monitoring organophosphorus pesticides and carbamates is described. The presence of 0.5% ruthenium on activated carbon mixed to form a simple graphite-based ink for the working electrode surface increased the sensitivity towards hydrogen peroxide. The choline biosensor is based on such an electrode coupled with choline oxidase immobilized by adsorption and was used to detect the inhibition effect of carbamates and organophosphorus pesticides on acetylcholinesterase. With the optimized procedure described (pH, buffer composition, incubation time, substrate concentration), concentrations of pesticides (Carbofuran) as low as 1 nM could be detected.
In this work, the first example of a hierarchically structured hollow silica system is reported without any chemical modification to the enzyme involved in the process. The leaching of the physically adsorbed enzyme is substantially restrained in comparison to pure hollow silica supports. The hierarchical architecture is composed of the ordered hollow silica spheres with a shell-in-shell structure. This rationally integrated architecture, which serves as the host for glucose oxidase immobilization, displays many significant advantages, including increases in mechanical stability, enzyme loading, and bioactivity, and a decrease in enzyme leaching compared to existing pure hollow silica matrices. This facilitates further multifarious applications for enhanced enzyme immobilization, biosensors, and biocatalysis.
A highly efficient nanoreactor that contains silver nanoparticles in hollow silica spheres and an interpolymer network as a gate-keeper has been developed following a facile procedure. The fast "signal-triggered switch of the smart network results in a high reactivity and a high response rate, yielding improved potential for many practical applications.
A CE/biosensor for measuring ascorbic acid was developed by coupling a polyaniline optical sensor and capillary electrophoresis (CE). The capillary column was partially modified with a thin film of polyaniline redox sensitive material. Ascorbic acid was detected by monitoring the changes in optical absorbance occurring to the polyaniline film upon the reduction reaction. The sensor response (change in optical absorbance at 650 nm) is proportional to the concentration of ascorbic acid over a range of 2.5-250 mg/L and the response range has shown a clear dependence on the characteristics of the polymerized film. High specificity and sensitivity of the present method, low sample consumption, short times of response (ca. 2 min) and the reproducibility of the results demonstrate that the CE/polyaniline-sensor could be further employed in the study of the relation between the content of L-ascorbic acid in body fluids and clinical parameters, e.g., cell ageing.