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PubMed2006

Characterization of an organic phase peroxide biosensor based on horseradish peroxidase immobilized in Eastman AQ.

Due to their frequent occurrence in food, cosmetics and pharmaceutical products, and their poor solubility in water, the detection of peroxides in organic solvents has aroused significant interest. For diagnostics or on-site testing, a fast and specific experimental approach is required. Although aqueous peroxide biosensors are well known, they are usually not suitable for nonaqueous applications due to their instability. Here we describe an organic phase biosensor for hydrogen peroxide based on horseradish peroxidase immobilized in an Eastman AQ 55 polymer matrix. Rotating disc amperometry was used to examine the effect of the solvent properties, the amount and pH of added buffer, the concentration of peroxide and ferrocene dimethanol, and the amount of Eastman AQ 55 and of enzyme on the response of the biosensor to hydrogen peroxide. The response of the biosensor was limited by diffusion. Linear responses (with detection limits to hydrogen peroxide given in parentheses) were obtained in methanol (1.2 microM), ethanol (0.6 microM), 1-propanol (2.8 microM), acetone (1.4 microM), acetonitrile (2.6 microM), and ethylene glycol (13.6 microM). The rate of diffusion of ferrocene dimethanol was more constrained than the rate of diffusion of hydrogen peroxide, resulting in a comparatively narrow linear range. The main advantages of the sensor are its ease of use and a high degree of reproducibility, together with good operational and storage stability.

Biosensing Techniques↗

PubMed2006

Glutamate optical biosensor based on the immobilization of glutamate dehydrogenase in titanium dioxide sol-gel matrix.

A simple and novel titania sol-gel derived optical biosensor coupled with carboxy seminaphthorhodamine-1-dextran (SNARF-1-dextran) as the fluorescent dye was fabricated for the determination of glutamate in water and biological samples. The NADH-dependent glutamate dehydrogenase (GLDH) was trapped in titania sol-gel derived matrix prepared by vapor deposition method. In addition, scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to characterize the surface morphology of the spots. SEM and AFM images showed that the deposition of titania precursor at 27 degrees C for 6.5h was found to be suitable to form transparent titania sol-gel matrix to encapsulate GLDH and fluorescent probe. AFM images showed that the roughness of TiO(2) surface increased from 2.16 nm in the absence of GLDH and SNARF to 37.8 nm after the immobilization. The developed titania biosensor has good analytical performance with water samples. A dynamic range between 0.04 and 10mM with the detection limit of 5.5 microM were observed. The responses to glutamate in biological samples also showed good performances, and the dynamic range and detection limit were 0.02-10mM and 6.7 microM, respectively. High precision with relative standard deviations of 4.2 and 10.7% in water and biological samples, respectively, were also demonstrated. In addition, the biosensor showed a relatively high storage stability over more than 1 month. Results obtained in this study clearly demonstrate that this simple vapor deposition method can be successfully used to form transparent titania sol-gel film for the fabrication of glutamate biosensors that are suitable for optical detection of glutamate in water and biological samples.

Biosensing Techniques↗

PubMed2006

Cr(VI) quantification using an amperometric enzyme-based sensor: interference and physical and chemical factors controlling the biosensor response in ground waters.

The development of an amperometric enzyme-based sensor for chromate (CrO(4)(2-)) quantification in ground waters was investigated. Crucial physical and chemical factors characterising ground waters were tested for their influence or interference on chromate quantification: pH (7.6-8.5), temperature (9-25 degrees C), ionic strength (0-0.2M), oxygen, metals, bicarbonate and sulphate. The biosensor's response was dependent on temperature and pH as sensitivity increased with temperature and was higher at pH 7.6 than at pH 8.5. Sensitivity decreased with ionic strength until 0.1M, and was stable for higher values. Dissolved oxygen did not allow chromate quantification when it was present, but O(2) could be eliminated by adding Na(2)SO(3) or bubbling nitrogen gas into the solution. Bicarbonate did not interfere with chromate quantification by the biosensor. Sulphate was detected with a detection threshold 80 times higher than that of chromate and a lower sensitivity. Several metals (V(V), W(VI), Mn(VII), Mo(VI)) similar to chromate due to their oxidative properties and structure (oxyanions) were tested as possible interfering compounds. The sensitivity of the biosensor for these metals was low and the detection level was 30 times higher than that of chromate. These metal concentrations are usually weaker than chromate concentration in polluted ground waters so that dilution of the sample should allow chromate quantification by the biosensor. This study shows that the cytochrome c(3)-based sensor can detect compounds other than chromate but with a lower sensitivity. Although non-specific for the detection of chromate, it can however be adapted and used for the quantification of chromate in ground waters containing low sulphate concentration.

Anions↗

PubMed2006

Integrated-optic biosensor by electro-optically modulated surface plasmon resonance.

We present a new integrated-optic surface plasmon resonance (SPR) biosensor based on electro-optical modulation. The SPR characteristics for the analyte concentration detection can be electro-optically modulated by applying the voltage on the electrodes of the biosensor fabricated on lithium niobate, which is an excellent electro-optic material. Two measurement methods, electro-optically modulated SPR spectral measurement and electro-optically modulated SPR intensity measurement, are demonstrated and their measurands are the SPR wavelength and the output optical intensity, respectively. Human serum albumin is coated on the gold film surface of the proposed biosensor to detect the concentration of beta-blocker, which is a remedy for heart disease. As the applied voltage increases such that the effective index of guided mode rises, the SPR wavelength shifts toward the long wavelength side and the output optical intensity at the wavelength of 632.8 nm diminishes. The linear regression slope of the relation between the measurand and the applied voltage is dependent on the analyte concentration and can be used to determine the concentration variation. Experimental results measured by the electro-optically modulated SPR methods are compared with those measured by the conventional spectral and intensity methods, and the effects of waveguide width on the biosensor performance are discussed.

Electrodes↗

PubMed2006

An amperometric uric acid biosensor based on modified Ir-C electrode.

The level of uric acid (UA) has a high relationship with gout, hyperuricemia and Lesch-Nyan syndrome. The determination of UA is an important indicator for clinics and diagnoses of kidney failure. An amperometric UA biosensor based on an Ir-modified carbon (Ir-C) working electrode with immobilizing uricase (EC 1.7.3.3) was developed by thick film screen printing technique. This is the first time to report the utilization of an uricase/Ir-C electrode for the determination of UA by using chronoamperometric (CA) method. The high selectivity of UA biosensor was achieved due to the reduction of H(2)O(2) oxidation potential based on Ir-C electrode. Using uricase/Ir-C as the sensing electrode, the interference from the electroactive biological species, such as ascorbic acid (AA) and UA (might be directly oxidized on the sensing electrode) was slight at the sensing potential of 0.25 V (versus Ag/AgCl). UA was detected amperometrically based on uricase/Ir-C electrode with a sensitivity of 16.60 microAmM(-1) over the concentration range of 0.1-0.8 mMUA, which was within the normal range in blood. The detection limit of UA biosensor was 0.01 mM (S/N=6.18) in pH 7 phosphate buffer solution (PBS) at 37 degrees C. The effects of pH, temperature, and enzymatic loading on the sensing characteristics of the UA biosensor were also investigated in this study.

Biosensing Techniques↗

PubMed2006

Development of amperometric biosensor for glucose based on a novel attractive enzyme immobilization matrix: calcium carbonate nanoparticles.

Calcium carbonate nanoparticles (nano-CaCO3) may be a promising material for enzyme immobilization owing to their high biocompatibility, large specific surface area and their aggregation properties. This attractive material was exploited for the mild immobilization of glucose oxidase (GOD) in order to develop glucose amperometric biosensor. The GOD/nano-CaCO3-based sensor exhibited a marked improvement in thermal stability compared to other glucose biosensors based on inorganic host matrixes. Amperometric detection of glucose was evaluated by holding the modified electrode at 0.60 V (versus SCE) in order to oxidize the hydrogen peroxide generated by the enzymatic reaction. The biosensor exhibited a rapid response (6s), a low detection limit (0.1 microM), a wide linear range of 0.001-12 mM, a high sensitivity (58.1 mAcm-2M-1), as well as a good operational and storage stability. In addition, optimization of the biosensor construction, the effects of the applied potential as well as common interfering compounds on the amperometric response of the sensor were investigated and discussed herein.

Biosensing Techniques↗

PubMed2006

Interdigitated array microelectrode based impedance biosensor coupled with magnetic nanoparticle-antibody conjugates for detection of Escherichia coli O157:H7 in food samples.

An impedance biosensor based on interdigitated array microelectrode (IDAM) coupled with magnetic nanoparticle-antibody conjugates (MNAC) was developed and evaluated for rapid and specific detection of E. coli O157:H7 in ground beef samples. MNAC were prepared by immobilizing biotin-labeled polyclonal goat anti-E. coli antibodies onto streptavidin-coated magnetic nanoparticles, which were used to separate and concentrate E. coli O157:H7 from ground beef samples. Magnitude of impedance and phase angle were measured in a frequency range of 10 Hz to 1 MHz in the presence of 0.1M mannitol solution. The lowest detection limits of this biosensor for detection of E. coli O157:H7 in pure culture and ground beef samples were 7.4 x 10(4) and 8.0 x 10(5)CFU ml(-1), respectively. The regression equation for the normalized impedance change (NIC) versus E. coli O157:H7 concentration (N) in ground beef samples was NIC=15.55 N-71.04 with R(2)=0.95. Sensitivity of the impedance biosensor was improved by 35% by concentrating bacterial cells attached to MNAC in the active layer of IDAM above the surface of electrodes with the help of a magnetic field. Based on equivalent circuit analysis, it was observed that bulk resistance and double layer capacitance were responsible for the impedance change caused by the presence of E. coli O157:H7 on the surface of IDAM. Surface immobilization techniques, redox probes, or sample incubation were not used in this impedance biosensor. The total detection time from sampling to measurement was 35 min.

Antibodies↗

PubMed2006

Application of a unique server-based oligonucleotide probe selection tool toward a novel biosensor for the detection of Streptococcus pyogenes.

We developed a software program for the rapid selection of detection probes to be used in nucleic acid-based assays. In comparison to commercially available software packages, our program allows the addition of oligotags as required by nucleic acid sequence-based amplification (NASBA) as well as automatic BLAST searches for all probe/primer pairs. We then demonstrated the usefulness of the program by designing a novel lateral flow biosensor for Streptococcus pyogenes that does not rely on amplification methods such as the polymerase chain reaction (PCR) or NASBA to obtain low limits of detection, but instead uses multiple reporter and capture probes per target sequence and an instantaneous amplification via dye-encapsulating liposomes. These assays will decrease the detection time to just a 20 min hybridization reaction and avoid costly enzymatic gene amplification reactions. The lateral flow assay was developed quantifying the 16S rRNA from S. pyogenes by designing reporter and capture probes that specifically hybridize with the RNA and form a sandwich. DNA reporter probes were tagged with dye-encapsulating liposomes, biotinylated DNA oligonucleotides were used as capture probes. From the initial number of capture and reporter probes chosen, a combination of two capture and three reporter probes were found to provide optimal signal generation and significant enhancement over single capture/reporter probe combinations. The selectivity of the biosensor was proven by analyzing organisms closely related to S. pyogenes, such as other Streptococcus and Enterococcus species. All probes had been selected by the software program within minutes and no iterative optimization and re-design of the oligonucleotides was required which enabled a very rapid biosensor prototyping. While the sensitivity obtained with the biosensor was only 135 ng, future experiments will decrease this significantly by the addition of more reporter and capture probes for either the same rRNA or a different nucleic acid target molecule. This will lead to the possibility of detecting S. pyogenes with a rugged assay that does not require a cell culturing or gene amplification step and will therefore enable rapid, specific and sensitive onsite testing.

Biosensing Techniques↗

PubMed2006

Development of a carbon nanotube paste electrode osmium polymer-mediated biosensor for determination of glucose in alcoholic beverages.

This paper describes a new amperometric biosensor for glucose monitoring. The biosensor is based on the activity of glucose dehydrogenase (GDH) and diaphorase (DI) co-immobilized with NAD(+) into a carbon nanotube paste (CNTP) electrode modified with an osmium functionalized polymer. This mediator was demonstrated to shuttle the electron transfer between the immobilized diaphorase and the CNTP electrode, thus, showing a good electrocatalytic activity towards NADH oxidation at potentials around +0.2V versus Ag|AgCl, where interfering reactions are less prone to occur. The biosensor exhibits a detection limit of 10 micromol L(-1), linearity up to 8 x 10(-4) mol L(-1), a sensitivity of 13.4 microA cm(-2)mmol(-1)L(-1), a good reproducibility (R.S.D. 2.1%, n=6) and a stability of about 1 week when stored dry at 4 degrees C. Finally, the proposed biosensor was applied for the determination of glucose in different samples of sweet wine and validated with a commercial spectrophotometric enzymatic kit.

Biosensing Techniques↗

PubMed2006

Biosensor based on polyaniline-Prussian Blue/multi-walled carbon nanotubes hybrid composites.

In this work, a novel route for fabrication polyaniline (PANI)-Prussian Blue (PB) hybrid composites is proposed by the spontaneous redox reaction in the FeCl(3)-K(3)[Fe(CN)(6)] and the aniline solution. With the introduction of multi-walled carbon nanotubes (MWNTs), the PANI-PB/MWNTs system shows synergy between the PANI-PB and MWNTs which amplified the H(2)O(2) sensitivity greatly. A linear range from 8 x1 0(-8) to 1 x 10(-5)M and a high sensitivity 508.1 8 microA microM cm(2) for H(2)O(2) detection are obtained. The composites also show good stability in neutral solution. A glucose biosensor was further constructed by immobilizing glucose oxidase (GOD) with Nafion and glutaraldehyde on the electrode surface. The performance factors influencing the resulted biosensor were studied in detail. The biosensor exhibits excellent response performance to glucose with the linear range from 1 to 11 mM and a detection limit of 0.01 mM. Furthermore, the biosensor shows rapid response, high sensitivity, good reproducibility, long-term stability and freedom of interference from other co-existing electroactive species.

Aniline Compounds↗

PubMed2006

Immobilization of enzymes into nanocavities for the improvement of biosensor stability.

Nanoporous materials with different pore sizes are evaluated as immobilization and stabilization matrices of proteins for the development of highly stable biosensors. It has been proven experimentally that confinement of proteins in cages with a diameter that is 2-6 times larger than their size increases considerably the stability of the biomolecules, as has been shown earlier by theoretical calculations. Porous silica beads with pore sizes of 10nm were utilized for the immobilization of the enzymes HRP and GOx with diameters in the order of 5 and 7 nm, respectively. The sensitivity of the corresponding biosensor systems was monitored for 70 h under continuous operation conditions (+600 mV) and it was found that the stabilization factor of GOx is 1.7 times higher compared to HRP. Also the stabilization efficiency of enzymes against leaching and inactivation in porous polymer beads with pore diameters of 10 and 30 nm was examined. The leaching rate of the enzyme AChE from the 30 nm polymer beads was found to be 1.1 times higher than that from the 10nm beads. At the same time the remaining activity of GOx biosensors after 5 days of continuous operation conditions (+600 mV) was found to be 2.1 times higher when the enzyme had been immobilized in the 10nm beads compared to the 30 nm beads. It is thus evident that the matching between the pore size of nanoporous materials and the molecular size of enzymes is essential for the development of biosensors with extended shelf and operational lifetimes.

Biosensing Techniques↗

PubMed2006

Amperometric and impedimetric characterization of a glutamate biosensor based on Nafion and a methyl viologen modified glassy carbon electrode.

An electrochemical biosensor based on a glassy carbon (GC) electrode chemically modified with the perfluorinated cation-exchange polymer Nafion and methyl viologen (MV) is described. The enzyme was immobilized by cross-linking with glutaraldehyde in the presence of bovine serum albumin (BSA), methyl viologen and Nafion. Operating variables such as the enzyme/BSA ratio, cross-linking time in glutaraldehyde vapor, methyl viologen and Nafion percentages were investigated with regard to their influence on the biosensor sensitivity by using glucose oxidase as the enzyme model due to its high stability and low cost. The glutamate biosensor was elaborated by using optimized parameters and its electrochemical properties were investigated by cyclic voltammetry, amperometry and by electrochemical impedance spectroscopy. The glutamate biosensor shows a detection limit of 20 microM and a linear range extended to 0.75 mM. Its selectivity was tested with 15 different amino acids, each with a concentration of 20 microM, 25 microM acetaminophen, 20 microM uric acid and 200 microM ascorbic acid. No amperometric response was observed for the interfering species. This good selectivity allows glutamate detection in biological media without previous separation of the analyte.

Biosensing Techniques↗

PubMed2003

On-line and in situ biosensors for monitoring environmental pollution.

Efficient tools for on-line and in situ monitoring of environmental pollutants are required to provide early warning systems. In addition, such tools can contribute important information on the progress of various remediation treatments. One of the recently developed monitoring technologies involves the use of whole-cell biosensors. Such biosensors could be constructed to detect general toxicity or specific toxicity caused by one or more pollutants. Currently, a large spectrum of microbial biosensors have been developed that enable the monitoring of pollutants by measuring light, fluorescence, color or electric current. Electrochemical monitoring is of special interest for in situ measurements as it can be performed using simple, compact and mobile equipment and is easily adaptable for on-line measurements. Here we survey the potential application of electrochemical biosensors in monitoring of general toxicity as well as hydrocarbons and heavy metals.

Bacteria↗

PubMed2005

A fractal analysis of pathogen detection by biosensors.

A fractal analysis is presented for the detection of pathogens such as Franscisela tularensis, Yersinia pestis (the bacterium that causes plague), Bacillus anthracis, Venezuelan equine encephalitis (VEE) virus, Vavcinia virus, and Escherichia coli using a cellular analysis and notification of antigens risks and yields (CANARY) biosensor [T.H. Rider, M.S. Petrovic, F.E. Nargi, J.D Harper, E.D. Schwoebel, R.H. Mathews, D.J. Blanchard, L.T Bortolin, A.M. Young, J. Chen, M.A. Hollis, A cell-based sensor for rapid identification of pathogens, Science 301 (2003, 11 July) 213-215, T.H. Rider, M.S. Petrovic, F.E. Nargi, J.D. Harper, E.D. Schwoebel, R.H. Mathews, D.J. Blanchard, L.T. Bortolin, A.M. Young, J. Chen, M.A. Hollis, A cell-based sensor for rapid identification of pathogens, Science 301 (2003, 11 July) 213-215. Science Online, www.sciencemag.org/cgi/content/full/031/5630/213/DC1]. In general, the binding and dissociation rate coefficients may be adequately described by either a single- or a dual-fractal analysis. An attempt is made to relate the binding rate coefficient to the degree of heterogeneity (fractal dimension value) present on the biosensor surface. Binding and dissociation rate coefficient values obtained are presented. Due to the dilute nature of the analyte(s) present, in some cases, a triple-fractal analysis is required to adequately describe the binding kinetics. It should be noted, and this is not entirely unexpected, that there is a lot of variation in the original experimental data when dilute concentrations of the analyte were analyzed by the CANARY biosensor [T.H. Rider, M.S. Petrovic, F.E. Nargi, J.D Harper, E.D. Schwoebel, R.H. Mathews, D.J. Blanchard, L.T Bortolin, A.M. Young, J. Chen, M.A. Hollis, A cell-based sensor for rapid identification of pathogens, Science 301 (2003, 11 July) 213-215, Science Online, www.sciencemag.org/cgi/content/full/031/5630/213/DC1]. The data analyzed in this manuscript appears smoother since only discrete points at different time intervals were analyzed. The kinetics aspects along with the affinity values presented are of interest and should along with the rate coefficients presented for the binding and the dissociation phase be of significant interest in help designing better biosensors for an application area that is bound to gain increasing importance in the future.

B-Lymphocytes↗

PubMed2006

Biosensors and bioelectrochemistry.

This review describes recent developments in the field of biosensors and bioelectrochemistry. Nanoparticles have been used to improve sensor performance and to develop biosensors based on new detection principles. Their use has extended into all areas of biosensor and bioelectrochemistry research. Other active areas of biosensor development include DNA sensing, immunosensing, direct electron transfer between an electrode and a redox protein or enzyme, and in vivo sensors.

Biosensing Techniques↗

PubMed2004

Biosensors for detection of mercury in contaminated soils.

Biosensors based on whole bacterial cells and on bacterial heavy metal binding protein were used to determine the mercury concentration in soil. The soil samples were collected in a vegetable garden accidentally contaminated with elemental mercury 25 years earlier. Bioavailable mercury was measured using different sensors: a protein-based biosensor, a whole bacterial cell based biosensor, and a plant sensor, i.e. morphological and biochemical responses in primary leaves and roots of bean seedlings grown in the mercury-contaminated soil. For comparison the total mercury concentration of the soil samples was determined by AAS. Whole bacterial cell and protein-based biosensors gave accurate responses proportional to the total amount of mercury in the soil samples. On the contrary, plant sensors were found to be less useful indicators of soil mercury contamination, as determined by plant biomass, mercury content of primary leaves and enzyme activities.

Biosensing Techniques↗

PubMed2005

Biosensor-based diagnostics of contaminated groundwater: assessment and remediation strategy.

Shallow groundwater beneath a former airfield site in southern England has been heavily contaminated with a wide range of chlorinated solvents. The feasibility of using bacterial biosensors to complement chemical analysis and enable cost-effective, and focussed sampling has been assessed as part of a site evaluation programme. Five different biosensors, three metabolic (Vibrio fischeri, Pseudomonas fluorescens 10568 and Escherichia coli HB101) and two catabolic (Pseudomonas putida TVA8 and E. coli DH5alpha), were employed to identify areas where the availability and toxicity of pollutants is of most immediate environmental concern. The biosensors used showed different sensitivities to each other and to the groundwater samples tested. There was generally a good agreement with chemical analyses. The potential efficacy of remediation strategies was explored by coupling sample manipulation to biosensor tests. Manipulation involved sparging and charcoal treatment procedures to simulate remediative engineering solutions. Sparging was sufficient at most locations.

Aliivibrio fischeri↗

PubMed2005

Assessment of heavy metal bioavailability in contaminated sediments and soils using green fluorescent protein-based bacterial biosensors.

A green fluorescent protein (GFP)-based bacterial biosensor Escherichia coli DH5alpha (pVLCD1) was developed based on the expression of gfp under the control of the cad promoter and the cadC gene of Staphylococcus aureus plasmid pI258. DH5alpha (pVLCD1) mainly responded to Cd(II), Pb(II), and Sb(III), the lowest detectable concentrations being 0.1 nmol L(-1), 10 nmol L(-1), and 0.1 nmol L(-1), respectively, with 2h exposure. The biosensor was field-tested to measure the relative bioavailability of the heavy metals in contaminated sediments and soil samples. The results showed that the majority of heavy metals remained adsorbed to soil particles: Cd(II)/Pb(II) was only partially available to the biosensor in soil-water extracts. Our results demonstrate that the GFP-based bacterial biosensor is useful and applicable in determining the bioavailability of heavy metals with high sensitivity in contaminated sediment and soil samples and suggests a potential for its inexpensive application in environmentally relevant sample tests.

Biological Availability↗