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At least 127 records · Page 7Linked to original sources
PubMed1996

Optimization of biosensing using grating couplers: immobilization on tantalum oxide waveguides.

This paper presents a comparative study of immobilization strategies for integrated optical grating couplers using tantalum oxide waveguides. As a model system the affinity reaction between protein G and human IgG was investigated. The receptors were coupled to the waveguide by adsorption, covalent attachment and avidin-biotin bridges after modifying the sensor surface by silanization introducing epoxy and amino groups. In addition, the results obtained by a monolayer coverage of the sensors with the receptors were compared with those using a carboxymethyl-dextran matrix allowing the immobilization of an increased amount of protein. The different coupling procedures were assessed by the system response due to the specific binding of human IgG to the sensing layer and are discussed with respect to their effectiveness, stability and the absence of non-specific binding.

Adsorption↗
PubMed2005

A new method of biosensing with 1 microl of Escherichia coli suspension using atomic force microscopy.

We developed a new method for detecting bacterial cells from 1-mul samples with atomic force microscopy (AFM). The use of a parafilm surface as a sample palette was effective for reacting small amounts of samples with an AFM probe. This was due to the parafilm's hydrophobic, semitransparent, and nonadhesive surface. In this way, all processes, such as the surface functionalization of a cantilever and the adhesion of Escherichia coli cells to a cantilever, were easily completed. In addition, we succeeded in detecting cell adsorption on the same AFM cantilever by both the drive mode and the thermal mode. The resonance frequency shift caused by cell adhesion was clearly detected by the two modes for the first time. Our data indicated the potential of applying AFM nanobiosensing to extremely small amounts of samples.

Bacterial Adhesion↗
PubMed2005

Multilayer membranes for glucose biosensing via layer-by-layer assembly of multiwall carbon nanotubes and glucose oxidase.

A bilayer of the polyelectrolytes poly(dimethyldiallylammonium chloride) (PDDA) and poly(sodium 4-styrenesulfonate) (PSS) was formed on a 3-mercapto-1-propanesulfonic-acid-modified Au electrode. Subsequently, multiwall carbon nanotubes (MWCNTs) wrapped by positively charged PDDA were assembled layer-by-layer with negatively charged glucose oxidase (GOx) onto the PSS-terminated bilayer. Electrochemical impedance spectroscopy and atomic force microscopy were adopted to monitor the regular growth of the PDDA-MWCNTs/GOx bilayers. Using GOx as a model enzyme, the assembled multilayer membranes showed some striking features such as the adsorbed form of GOx on individual MWCNT, uniformity, good stability, and electrocatalytic activity toward oxygen reduction. Based on the consumption of dissolved oxygen during the oxidation process of glucose catalyzed by the immobilized GOx, a sensitive amperometric biosensor was developed for the detection of glucose up to 5.0 mM with a detection limit of 58 microM. The sensitivity increased with increasing sensing layers up to five bilayers. Ascorbic acid and uric acid did not cause any interference due to the use of a low operating potential. The present method showed high reproducibility for the fabrication of carbon-nanotubes-based amperometric biosensors.

Biosensing Techniques↗
PubMed2005

Design, preparation and testing of suitable probe-receptors for RNA biosensing.

Absolute measurements of a given RNA in a cheap, easy, rapid and reproducible manner using biosensors technology could overcome many of the operative and analytical limits of conventional molecular biology methods. To this end, an integrated approach for the design, synthesis, and connection of RNA probes to the transducing surface of a microgravimetric biosensor has been developed. Suitable probes to be used as the bioreceptors in RNA biosensor were successfully designed by using a purposely developed computational method whose selection criteria are based on the accessibility of target region to probe, on pairing stability of probe-target duplex and on the uniqueness of selected targets over all known expressed sequences from a genome data base. Automated chemical synthesis of selected probes was performed and the oligonucleotides produced were covalently conjugated to the sensing surface of a quartz microbalance. The microgravimetric sensor was tested in a flow chamber by measuring the variation of resonance frequency due to the binding of synthetic target substrates. Specific dose dependent binding was observed. Furthermore, the binding of a transcribed full-length mRNA substrate was successfully monitored under similar conditions.

Base Sequence↗
PubMed2005

Characterization of natural chitosan membranes from the carapace of the soldier crab Mictyris brevidactylus and its application to immobilize glucose oxidase in amperometric flow-injection biosensing system.

This study investigated characteristics of a chitosan membrane from the carapace of the soldier crab Mictyris brevidactylus intended to construct an amperometric biosensor. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used in this study to characterize these chitosan membranes intended for constructing enzymatic biosensors. Chitosan membranes suffering various durations (>10 min) of deacetylation had small charge-transfer resistances (<7.88 kohms) but large double-layer capacitances (>0.55 microF). They were found in EIS where both the solution resistance and Warburg impedance upon electrode interface were almost independent of the durations and degree of deacetylation. The degree of deacetylation and the thickness of chitosan membranes were also determined. Membrane thickness was slightly dependent with the duration but degree of deacetylation was slightly dependent on the duration. Chitosan membranes with various thicknesses suffered various durations of deacetylation, but this did not influence their electrochemical characteristics. The chitinous membrane was covalently immobilized with glucose oxidase (EC 1.3.4.3) and then attached onto the platinum electrode of a homemade amperometric flow cell. Sensor signal was linearly related to glucose concentration (r=0.999 for glucose up to 1.0 mM). The system was sensitive (S/N>5 for 10 microM glucose) and reproducible (CV<1.3% for 50 microM glucose, n=5).

Acetylation↗
PubMed2006

Electrocatalytical properties of polymethylferrocenyl dendrimers and their applications in biosensing.

The electrochemical characterization of polymethylferrocenyl dendrimers deposited onto a platinum electrode and their applications as hydrogen peroxide and glucose sensor are described. The redox dendrimers consist of flexible poly(propileneimine) dendrimer cores functionalised with octamethylferrocenyl units. Amperometric biosensors for glucose were prepared by immobilization of glucose oxidase onto these modified electrodes. The influence of the dendrimer generation and the thickness of the dendrimer layer, the effect of the substrate concentration, and the interferences and reproducibility on the response of the sensors were investigated.

Biosensing Techniques↗
PubMed2005

A conductive ormosil encapsulated with ferrocene conjugate and multiwall carbon nanotubes for biosensing application.

Highly non-toxic and conductive ormosil composite film was prepared using (3-aminopropyl)triethoxysilane and 2-(3,4-epoxycyclohexyl)-ethyltrimethoxysilane by doping with ferrocenemonocarboxylic acid-bovine serum albumin (FMC-BSA) conjugate and multiwall carbon nanotubes (MWNTs). With glucose oxidase (GOD) as a model enzyme this film could be used to design an amperometric biosensor for glucose determination. The entrapped FMC-BSA conjugate performed excellent redox electrochemistry and the immobilized GOD was highly stable. Under optimal conditions this biosensor was able to detect glucose with a detection limit of 20 microm (S/N=3) in the linear range of 0.05-20.0 mm in flow system, which was wider than the batch amperometric mode, with an analysis time of 25 s for each sample. The value of K(M)(app) was 6.6 mm. The proximity of these three components FMC-BSA, MWNTs and GOD enhanced the electron transfer between the film and electrode. This film could be used efficiently for the entrapment of other redox bioactive compounds and biosensing/bioelectrochemical applications.

Biosensing Techniques↗
PubMed2006

Biomolecular immobilization on conducting polymers for biosensing applications.

A detail study on different aspects of biomolecule immobilization techniques on conducting polymers (CP) for applications in biosensors is described. Comparative studies are conducted in between the different mode of biomolecule immobilization techniques, viz. physical, covalent and electrochemical immobilization onto the conducting polymer films for the fabrication of electrochemical biosensors for clinical, food and environmental monitoring applications. This review focuses on the current status of biomolecule immobilization techniques on CP and their applications in the development of amperometric biosensors.

Adsorption↗
PubMed2004

Characterization of 3-D collagen hydrogels for functional cell-based biosensing.

To address the growing demand for functional cell-based assay technologies with accelerated drug discovery applications, we have proposed the use of human neuroblastoma cells (IMR-32) immobilized in three-dimensional (3-D) collagen hydrogel matrices. The gel protects weakly adherent cells from fluid mechanical forces while providing a more physiologically relevant 3-D environment. Hydrogels made up of collagen, between 0.5 and 1.0mg/ml, exhibited mechanical stability adequate to withstand fluid mechanical forces (<0.11 mN) typical of automated commercial fluid transfer equipment. Collagen-entrapped cells visualized with the aid of confocal microscopy and a potentiometric-sensitive dye, TMRM, exhibited round morphology in comparison to flat morphology typical of cells in two-dimensional (2-D) monolayer cultures. Morphological differentiation characterized by neurite extension and cell aggregation was observed for both 2-D and 3-D cultures. Differentiated IMR-32 cells failed to develop a resting membrane potential typical of excitable cells. Free intracellular calcium was monitored with Calcium Green-1. Depolarization-induced Ca 2+influx was only observed with differentiated 3-D cells unlike 2-D cells, where calcium flux was observed in both differentiated and undifferentiated cells. Taken together, the results revealed that collagen hydrogels (0.5 mg/ml collagen) were suitable structural supports for weakly adherent cells. However, for voltage-dependent calcium channel function applications, further investigations are needed to explain the difference between 2-D monolayer and 3-D collagen-entrapped cells.

Biocompatible Materials↗
PubMed2005

In vitro cortical neuronal networks as a new high-sensitive system for biosensing applications.

By taking advantages of the main features of the microelectrode array (MEA) technology (i.e. multisite recordings, stable and long-term coupling with the biological preparation), we analyzed the changes in activity patterns induced by applying specific substances to dissociated cortical neurons from rat-embryos (E18). Data were recorded simultaneously from 60 electrodes, and the electrophysiological behavior was investigated during the third week in vitro, both at the spike and burst level. The analysis of the electrophysiological activity modulation, by applying agonists of the ionotropic glutamate receptors at low (i.e. 0.2-1-5 microM) and high (i.e. 50-100 microM) concentrations, is presented. Preliminary results show that the dynamics of the in vitro cortical neurons is very sensitive to pharmacological manipulation of the glutamatergic transmission and the effects on the network behavior are strictly dependent from the drug concentration. In particular, the addition of a high-dose of agonist determined a global and irreversible depression of the network activity, while, in the low-concentration case, the electrophysiological behavior showed different results, depending on the type of receptor involved. From these observations, we are encouraged to think of a more engineered system, based on in vitro cortical neurons, as a novel sensitive system for drug (pre)-screening and neuropharmacological evaluations.

Action Potentials↗
PubMed2005

Electrochemical biosensing based on universal affinity biocomposite platforms.

Rigid conducting biocomposites are versatile and effective transducing materials for the construction of a wide range of amperometric biosensors such as immunosensors, genosensors and enzymosensors, particularly if the transducer is bulk-modified with universal affinity biomolecules. The strept(avidin)-graphite-epoxy biocomposite could be considered as an universal immobilization platform whereon biotinylated DNAs, oligonucleotides, enzymes or antibodies can be captured by means of the highly affinity (strept)avidin-biotin reaction. Universal affinity biocomposite-based biosensors offer many potential advantages compared to more traditional electrochemical biosensors commonly based on a biologically surface-modified transducer. The integration of many materials into one matrix is their main advantage. As biological bulk-modified materials, the conducting biocomposites act not only as transducers, but also as reservoir for the biomaterial. After its use, the electrode surface can be renewed by a simple polishing procedure, establishing a clear advantage of these approaches relative to classical biosensors and other common biological assays. Moreover, the same material is useful for the analysis of many molecules whose determinations are based on genetic, enzymatic or immunological reactions. The different strategies for electrochemical genosensing, immunosensing and enzymosensing, all of them being dependent on the presence of a redox enzyme marker for the generation of the electrochemical signal, based on this universal affinity biocomposite platform are all presented and discussed.

Biosensing Techniques↗
PubMed2005

Catechol biosensing using a nanostructured layer-by-layer film containing Cl-catechol 1,2-dioxygenase.

The detection of aromatic compounds from pesticides and industrial wastewater has become of great interest, since these compounds withstand chemical oxidation and biological degradation, accumulating in the environment. In this work, a highly sensitive biosensor for detecting catechol was obtained with the immobilization of Cl-catechol 1,2-dioxygenase (CCD) in nanostructured films. CCD layers were alternated with poly(amidoamine) generation 4 (PAMAM G4) dendrimer using the electrostatic layer-by-layer (LbL) technique. Circular dichroism (CD) measurements indicated that the immobilized CCD preserved the same conformation as in solution. The thickness of the very first CCD layers in the LbL films was estimated at ca. 3.6 nm, as revealed by surface plasmon resonance (SPR). PAMAM/CCD 10-bilayer films were employed in detecting diluted catechol solutions using either an optical or electrical approach. Due to the mild immobilization conditions employed, especially regarding the pH and ionic strength of the dipping solutions, CCD remained active in the films for periods longer than 3 weeks. The optical detection comprised absorption experiments in which the formation of cis-cis muconic acid, resulting from the reaction between CCD and catechol, was monitored by measuring the absorbance at 260 nm after film immersion in catechol solutions. The electrical detection was carried out using LbL films deposited onto gold-interdigitated electrodes immersed in aqueous solutions at different catechol concentrations. Using impedance spectroscopy in a broad frequency range (1Hz-1kHz), we could detect catechol in solutions at concentrations as low as 10(-10) M.

Adsorption↗
PubMed2005

Human neuroblastoma (SH-SY5Y) cell culture and differentiation in 3-D collagen hydrogels for cell-based biosensing.

Cell-based three-dimensional systems are desirable in the field of high throughput screening assays due to their potential similarity to in vivo environment. We have used SH-SY5Y human neuroblastoma cells cultured in 3-D collagen hydrogel, confocal microscopy and immunofluorescence staining, to assess the merit of the system as a functional, cell-based biosensor. Our results show differences between 2-D and 3-D resting membrane potential development profile upon differentiation. There was no statistically significant difference in SH-SY5Y proliferation rate between 2-D monolayer and 3-D collagen culture formats. A large percentage of cells (2-D, 91.30% and 3-D, 84.93%) did not develop resting membrane potential value equal to or lower than -40 mV; instead cells exhibited a heterogeneous resting membrane potential distribution. In response to high K(+) (50 mM) depolarization, 3-D cells were less responsive in terms of increase in intracellular Ca(2+), in comparison to 2-D cells, supporting the hypothesis that 2-D cell calcium dynamics may be exaggerated. L-Type Ca(2+) expression levels based on staining results was inconsistent with Bay K 8644 channel activation results, strongly suggesting that either the majority of the channels were non-functional or could not be activated by Bay K 8644. In general, the results in this study confirm the depolarization-induced differences in intracellular calcium release when cultured using a 2-D versus a 3-D matrix.

Biological Assay↗
PubMed2005

Biosensing approach for alcohol determination using immobilized alcohol oxidase.

Alcohol oxidase (AOD) was immobilized in polypyrrole (PPy) and a random copolymer containing 3-methylthienyl methacrylate and p-vinylbenzyloxy poly(ethyleneoxide) matrices. Immobilization of enzyme was performed via entrapment in conducting polymers during electrochemical polymerization of pyrrole through the thiophene moiety of the copolymer. Three different alcohols, namely methanol, ethanol and n-propanol, were used as substrates. Maximum reaction rates, Michaelis-Menten constants, optimum temperature and pH values, operational stabilities and shelf life of the enzyme electrodes were investigated.

Alcohol Oxidoreductases↗
PubMed2006

Nanocrystalline diamond modified gold electrode for glucose biosensing.

Boron-doped diamond has drawn much attention in electrochemical sensors. However there are few reports on non-doped diamond because of its weak conductivity. Here, we reported a glucose biosensor based on electrochemical pretreatment of non-doped nanocrystalline diamond (N-NCD) modified gold electrode for the selective detection of glucose. N-NCD was coated on gold electrode and glucose oxidase (GOx) was immobilized onto the surfaces of N-NCD by forming amide linkages between enzyme amine residues and carboxylic acid groups on N-NCD. The anodic pretreatment of N-NCD modified electrode not only promoted the electron transfer rate in the N-NCD thin film, but also resulted in a dramatic improvement in the reduction of the dissolved oxygen. This performance could be used to detect glucose at negative potential through monitoring the current change of oxygen reduction. The biosensor effectively performs a selective electrochemical analysis of glucose in the presence of common interferents, such as ascorbic acid (AA), acetaminophen (AP) and uric acid (UA). A wide linear calibration range from 10 microM to 15 mM and a low detection limit of 5 microM were achieved for the detection of glucose.

Biosensing Techniques↗
PubMed2006

Electrochemical biosensing utilizing synergic action of carbon nanotubes and platinum nanowires prepared by template synthesis.

Platinum nanowires (PtNWs) prepared by electrodeposition method with the help of porous anodic aluminum oxide (AAO) templates have been solubilized in chitosan (CHIT) together with carbon nantubes (CNTs) to form a PtNW-CNT-CHIT organic-inorganic system. The resulting PtNW-CNT-CHIT material brings capabilities for utilizing synergic action of PtNWs and CNTs to facilitate electron-transfer process in electrochemical sensor design. The PtNW-CNT-CHIT film modified electrode offered a significant decrease in the overvoltage for the hydrogen peroxide and showed to be excellent amperometric sensors for hydrogen peroxide at -0.1 V over a wide range of concentrations, and the sensitivity is 260 microAmM-1cm-2. As an application example, by linking glucose oxidase (GOx), an amplified biosensor toward glucose was prepared. The glucose biosensor exhibits a selective determination of glucose at -0.1 V with a linear response range of 5 x 10(-6) to 1.5 x 10(-2)M with a correlation coefficient of 0.997, and response time <10s. The high sensitivity of the glucose biosensor is up to 30 microAmM-1cm-2 and the detection limit was 3 microM. The biosensor displays rapid response and expanded linear response range, and excellent repeatability and stability.

Biosensing Techniques↗
PubMed2006

Integrated nanoparticle-biomolecule systems for biosensing and bioelectronics.

The similar dimensions of biomolecules such as enzymes, antibodies or DNA, and metallic or semiconductor nanoparticles (NPs) enable the synthesis of biomolecule-NP hybrid systems where the unique electronic, photonic and catalytic properties of NPs are combined with the specific recognition and biocatalytic properties of biomolecules. The unique functions of biomolecule-NP hybrid systems are discussed with several examples: (i) the electrical contacting of redox enzymes with electrodes is the basis for the development of enzymatic electrodes for amperometric biosensors or biofuel cell elements. The reconstitution of the apo-glucose oxidase or apo-glucose dehydrogenase on flavin adenine dinucleotide (FAD)-functionalized Au NPs (1.4 nm) associated with electrodes, or on pyrroloquinoline quinone (PQQ)-functionalized Au NPs (1.4 nm) associated with electrodes, respectively, yields electrically contacted enzyme electrodes. The aligned, reconstituted enzymes on the electrode surfaces reveal effective electrical contacting, and the glucose oxidase and glucose dehydrogenase reveal turnover rates of 5000 and 11,800 s(-1), respectively. (ii) The photoexcitation of semiconductor nanoparticles yields fluorescence with a wavelength controlled by the size of the NPs. The fluorescence functions of semiconductor NPs are used to develop a fluorescence resonance energy transfer (FRET) assay for nucleic acids, and specifically, for analyzing telomerase activity in cancer cells. CdSe-ZnS NPs are functionalized by a primer recognized by telomerase, and this is elongated by telomerase extracted from HeLa cancer cells in the presence of dNTPs and Texas-red-functionalized dUTP. The dye integrated into the telomers allows the FRET process that is intensified as telomerization proceeds. Also, the photoexcited electron-hole pair generated in semiconductor NPs is used to generate photocurrents in a CdS-DNA hybrid system associated with an electrode. A redox-active intercalator, methylene blue, was incorporated into a CdS-duplex DNA monolayer associated with a Au electrode, and this facilitated the electron transfer between the electrode and the CdS NPs. The direction of the photocurrent was controlled by the oxidation state of the intercalator. (iii) Biocatalysts grow metallic NPs, and the absorbance of the NPs provides a means to assay the biocatalytic transformations. This is exemplified with the glucose oxidase-induced growth of Au NPs and with the tyrosinase-stimulated growth of Au NPs, in the presence of glucose or tyrosine, respectively. The biocatalytic growth of the metallic NPs is used to grow nanowires on surfaces. Glucose oxidase or alkaline phosphatase functionalized with Au NPs (1.4 nm) acted as 'biocatalytic inks' for the synthesis of metallic nanowires. The deposition of the Au NP-modified glucose oxidase, or the Au NP-modified alkaline phosphatase on Si surfaces by dip-pen nanolithography led to biocatalytic templates, that after interaction with glucose/AuCl4- or p-aminophenolphosphate/Ag+, allowed the synthesis of Au nanowires or Ag nanowires, respectively.

Biosensing Techniques↗
PubMed2006

Biosensing nitrite using the system nitrite redutase/Nafion/methyl viologen--a voltammetric study.

This work describes the construction and voltammetric characterization of a nitrite biosensor based on a cytochrome c-type nitrite reductase (ccNiR) and the Nafion ionomeric matrix loaded with methyl viologen as redox mediator. Despite the potential electrostatic repulsions between the anionic substrate and the Nafion sulfonate groups, the resulting bioelectrode exhibited electrocatalytic activity toward nitrite. This phenomenon must be due to the nonuniformity of the enzyme/Nafion membrane, which allows the direct interaction between the substrate and numerous enzyme molecules. Nevertheless, the anionic nature of Nafion exerted a certain diffusion barrier to nitrite, as revealed by the unusually elevated limits of the linear dynamic range and k(m)(app). The irregularity of the composite membrane also contributed to slow down the rate of charge transfer throughout the Nafion polymer. The level of viologens incorporated within the Nafion membrane had a strong influence in the analytical parameters: as much mediator was present, lower was the sensitivity and wider was the linear range. For an optimized ratio enzyme/mediator the sensitivity was 445+/-8 mA M(-1)cm(-2), within the linear range 75-800 microM; the lowest detected nitrite concentration was 60 microM. The operational stability of the biosensor and the influence of some possible interferences were evaluated.

Biosensing Techniques↗