Amperometric biosensor for choline based on layer-by-layer assembled functionalized carbon nanotube and polyaniline multilayer film

Conducting polymer film was prepared by electrochemical polymerization of aniline. Multiwalled carbon nanotubes (MWNTs) were treated with a mixture of concentrated sulfuric and nitric acid to introduce carboxylic acid groups to the nanotubes. By using the layer-by-layer method, homogeneous and stable MWNTs and polyaniline (PANI) multilayer films were alternately assembled on glassy carbon (GC) electrodes. Conducting polymer of PANI had three main functions: (i) excellent antiinterference ability, (ii) protection ability in favor of increasing the amount of the MWNTs immobilized on GC electrodes, and (iii) superior transducing ability. The protection effect of PANI film and the electrostatic interaction between positively charged PANI and negatively charged MWNTs both attributed to immobilizing abundant MWNTs stably, thereby enhancing the catalytic activity. The layer-by-layer assembled MWNTs and PANI-modified GC electrodes offered a significant decrease in the overvoltage for the H2O2 and were shown to be excellent amperometric sensors for H2O2 from +0.2V over a wide range of concentrations. As an application example, by linking choline oxidase (CHOD), an amplified biosensor toward choline was prepared. The choline biosensor exhibited a linear response range of 1x10(-6) to 2x10(-3) M with a correlation coefficient of 0.997, and the response time and detection limit (S/N=3) were determined to be 3 s and 0.3 microM, respectively. The antiinterference biosensor displays a rapid response and an expanded linear response range as well as excellent reproducibility and stability.     

Amperometric glucose biosensor based on adsorption of glucose oxidase at platinum nanoparticle-modified carbon nanotube electrode

A new amperometric biosensor, based on adsorption of glucose oxidase (GOD) at the platinum nanoparticle-modified carbon nanotube (CNT) electrode, is presented in this article. CNTs were grown directly on the graphite substrate. The resulting GOD/Pt/CNT electrode was covered by a thin layer of Nafion to avoid the loss of GOD in determination and to improve the anti-interferent ability. The morphologies and electrochemical performance of the CNT, Pt/CNT, and Nafion/GOD/Pt/CNT electrodes have been investigated by scanning electron microscopy, cyclic voltammetry, and amperometric methods. The excellent electrocatalytic activity and special three-dimensional structure of the enzyme electrode result in good characteristics such as a large determination range (0.1-13.5mM), a short response time (within 5s), a large current density (1.176 mA cm(-2)), and high sensitivity (91mA M(-1)cm(-2)) and stability (73.5% remains after 22 days). In addition, effects of pH value, applied potential, electrode construction, and electroactive interferents on the amperometric response of the sensor were investigated and discussed. The reproducibility and applicability to whole blood analysis of the enzyme electrode were also evaluated.     

A novel multi-walled carbon nanotube-based biosensor for glucose detection

The bioelectrochemical characteristics of a novel multi-walled carbon nanotube (MWNT)-based biosensor for glucose detection are studied and compared with those of glassy carbon (GC)-based biosensor. The MWNT-based biosensor exhibits a strong glucose response at applied potentials of 0.65 and 0.45 V versus Ag/AgCl, respectively, while GC-based biosensor shows a weak glucose response at 0.65 V and no response at 0.45 V. Besides, the MWNT-based biosensor shows a high stability of 86.7% of the initial activity to glucose after four-month storage, much higher than 37.2%, the corresponding value for a GC-based biosensor. The detection mechanism of the MWNT-based biosensor is also discussed in detail.     

An amperometric oxalate biosensor based on sorghum oxalate oxidase bound carboxylated multiwalled carbon nanotubes-polyaniline composite film

A highly sensitive, specific and rapid electrochemical oxalate biosensor was constructed by covalently immobilizing sorghum leaf oxalate oxidase on carboxylated multiwalled carbon nanotubes and conducting polymer, polyaniline nanocomposite film electrodeposited over the surface of platinum (Pt) wire using N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy succinimide (NHS) chemistry. The modified electrode was characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectrophotometry. The optimized oxalate biosensor showed linear response range of 8.4-272 μM with correlation coefficient of 0.93 and rapid response within 5 s at a potential of 0.4 V vs Ag/AgCl. The sensitivity was approximately 0.0113 μA/μM with a detection limit of 3.0 μM. Proposed oxalate biosensor was successfully applied to human urine sample.     

Facile preparation of amperometric laccase biosensor with multifunction based on the matrix of carbon nanotubes-chitosan composite

The carbon nanotubes–chitosan (CNTs–CS) composite provides a suitable biosensing matrix due to its good conductivity, high stability, and good biocompatibility. Enzymes can be firmly incorporated into the matrix without the aid of other cross-linking reagents. The composite is easy to form insoluble film in solution above pH 6.3. Based on this, a facilely fabricated amperometric biosensor by entrapping laccase into the CNTs–CS composite film has been developed. At pH 6.0, the fungi laccase incorporated into the composite film remains better catalytic activity than that dissolved in solution. The system is in favor of the accessibility of substrate to the active site of laccase, thus the affinity to substrates is improved greatly, such as 2,2′-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt (ABTS), catechol, and O₂ with K_m values of 19.86 μM, 9.43 μM, and 3.22 mM, respectively. The major advantages of the as-prepared biosensor are: detecting different substrates (ABTS, catechol, and O₂), possessing high affinity and sensitivity, durable long-term stability, and facile preparation procedure. On the other hand, the system can be applied in fabrication of biofuel cells as the cathodic catalysts based on its good electrocatalysis for oxygen reduction. It can be extended to immobilize other enzymes and biomolecules, which will greatly facilitate the development of biosensors, biofuel cells, and other bioelectrochemical devices.     

Amperometric glucose biosensor based on single-walled carbon nanohorns

The biosensing application of single-walled carbon nanohorns (SWCNHs) was demonstrated through fabrication of an amperometric glucose biosensor. The biosensor was constructed by encapsulating glucose oxidase in the Nafion-SWCNHs composite film. The cyclic voltammograms for glucose oxidase immobilized on the composite film displayed a pair of well-defined and nearly symmetric redox peaks with a formal potential of -0.453 V. The biosensor had good electrocatalytic activity toward oxidation of glucose. To decrease detection potential, ferrocene monocarboxylic acid was used as a redox mediator. The mediated glucose biosensor shows a linear range from 0 to 6.0 mM. The biosensor shows high sensitivity (1.06 microA/mM) and stability, and can avoid the commonly coexisted interference. Because of impressive properties of SWCNHs, such as high purity and high surface area, SWCNHs and their composites are expected to be promising material for biomolecular immobilization and biosensing applications.     

An amperometric cholesterol biosensor based on multiwalled carbon nanotubes and organically modified sol-gel/chitosan hybrid composite film

A new type of amperometric cholesterol biosensor based on sol-gel chitosan/silica and multiwalled carbon nanotubes (MWCNTs) organic-inorganic hybrid composite material was developed. The hybrid composite film was used to immobilize cholesterol oxidase on the surface of Prussian blue-modified glass carbon electrode. Effects of some experimental variables such as enzyme loading, concentration of Triton X-100, pH, temperature, and applied potential on the current response of the biosensor were investigated. Analytical characteristics and dynamic parameters of the biosensors with and without MWCNTs in the hybrid film were compared, and the results show that analytical performance of the biosensor can be improved greatly after introduction of the MWCNTs. Response time, sensitivity, linear range, limit of detection (S/N=3), and apparent Michaelis-Menten constant Km are 25s, 0.54 microA mM(-1), 8.0 x 10(-6) to 4.5 x 10(-4) M, 4.0 x 10(-6) M, and 0.41 mM for the biosensor without MWCNTs and 13 s, 1.55 microA mM(-1), 4.0 x 10(-6) to 7.0 x 10(-4) M, 1.0 x 10(-6) M, and 0.24 mM for the biosensor with MWCNTs, respectively. The activation energy of the enzyme-catalyzed reaction was measured to be 42.6 kJ mol(-1). This method has been used to determine the free cholesterol concentration in real human blood samples.     

Cholesterol biosensor based on N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane self-assembled monolayer

Cholesterol oxidase (ChOx) has been covalently immobilized onto two-dimensional self-assembled monolayer (SAM) of N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane (AEAPTS) deposited on the indium-tin oxide (ITO) coated glass plates using N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) chemistry. These ChO x/AEAPTS/ITO bioelectrodes are characterized using contact angle (CA) measurements, UV-visible spectroscopy, atomic force microscopy (AFM), electrochemical impedance technique, and Fourier transform infrared (FT-IR) technique. The covalently immobilized ChOx-modified AEAPTS bioelectrodes are used for the estimation of cholesterol in solution using UV-visible technique. These cholesterol sensing bioelectrodes show linearity as 50 to 500 mg/dl for cholesterol solution, detection limit as 25mg/dl, sensitivity as 4.499 x 10(-5) Abs (mg/dl)(-1), K(m) value as 58.137 mg/dl (1.5mM), apparent enzyme activity as 1.81 x 10(-3) U cm(-2), shelf life of approximately 10 weeks, and electrode reusability as 10 times.     

Glucose biosensor based on glucose oxidase immobilized at gold nanoparticles decorated graphene-carbon nanotubes

Biopolymer pectin stabilized gold nanoparticles were prepared at graphene and multiwalled carbon nanotubes (GR-MWNTs/AuNPs) and employed for the determination of glucose. The formation of GR-MWNTs/AuNPs was confirmed by scanning electron microscopy, X-ray diffraction, UV–vis and FTIR spectroscopy methods. Glucose oxidase (GOx) was successfully immobilized on GR-MWNTs/AuNPs film and direct electron transfer of GOx was investigated. GOx exhibits highly enhanced redox peaks with formal potential of −0.40 V (vs. Ag/AgCl). The amount of electroactive GOx and electron transfer rate constant were found to be 10.5 × 10⁻¹⁰ mol cm⁻² and 3.36 s⁻¹, respectively, which were significantly larger than the previous reports. The fabricated amperometric glucose biosensor sensitively detects glucose and showed two linear ranges: (1) 10 μM – 2 mM with LOD of 4.1 μM, (2) 2 mM – 5.2 mM with LOD of 0.95 mM. The comparison of the biosensor performance with reported sensors reveals the significant improvement in overall sensor performance. Moreover, the biosensor exhibited appreciable stability, repeatability, reproducibility and practicality. The other advantages of the fabricated biosensor are simple and green fabrication approach, roughed and stable electrode surface, fast in sensing and highly reproducible.     

Amperometric cholesterol biosensor based on in situ reconstituted cholesterol oxidase on an immobilized monolayer of flavin adenine dinucleotide cofactor

A new amperometric biosensor for determining cholesterol based on deflavination of the enzyme cholesterol oxidase (ChOx) and subsequent reconstitution of the apo-protein with a complexed flavin adenine dinucleotide (FAD) monolayer is described. The charge transfer mediator pyrroquinoline quinone (PQQ) was covalently bound to a cystamine self-assembled monolayer (SAM) on an Au electrode. Boronic acid (BA) was then bound to PQQ using the carbodiimide procedure, and the BA ligand was complexed to the FAD molecules on which the apo-ChOx was subsequently reconstituted. The effective release of the FAD from the enzyme and the successful reconstitution were verified using molecular fluorescence and cyclic voltammetry. The optimal orientation of FAD toward the PQQ mediator and the distances between FAD and PQQ and between PQQ and electrode enhance the charge transfer, very high sensitivity (about 2,500 nAmM(-1)cm(-2)) being obtained for cholesterol determination. The biosensor is selective toward electroactive interferents (ascorbic acid and uric acid) and was tested in reference serum samples, demonstrating excellent accuracy (relative errors below 3% in all cases). The biosensor activity can be successfully regenerated in a simple process by successive reconstitution with batches of recently prepared apo-ChOx on the same immobilized Au/SAM-PQQ-BA-FAD monolayer (it was tested five times); the lifetime of the biosensor is about 45-60 days.     

Nanoscopic modeling of a carbon nanotube force-measuring biosensor

In this paper we perform a theoretical study of a potential design of a carbon nanotube device able to transduce forces developed at the scale of basic cellular processes into electric current variations. The first part of this study consists of an assessment of the sensitivity of the device with forces in the tens of pico Newtons (pN), developed typically at the cellular scale. In the second stage, we focus on the transduction of the deflection of a cantilever into an electrical signal, employing methods borrowed from non-equilibrium Green's functions. Several issues related to the importance of thermal effects in the proper operation of the sensor are then discussed. Following a simple method we include non-zero temperature through molecular dynamics in quantum conductance calculations that results in the displacement-current characteristic found at the end of this paper.     

Carbon nanotube aqueous sol-gel composites: enzyme-friendly platforms for the development of stable biosensors

A new type of composite material based on carbon nanotubes and an aqueous sol-gel process has been developed. The electrochemical characteristics of these composites were investigated and compared to composites made with an alkoxy silane sol-gel process. The use of carbon nanotubes, as the conductive part of the composite, facilitated fast electron transfer rates. The feasibility of this type of composite for the development of biosensors was demonstrated using l-amino acid oxidase. The stability of the enzyme was increased when it was encapsulated in the aqueous sol-gel, and the sensor retained more that 50% of its response after 1 month of testing.     

Amperometric sensor based on ferrocene-modified multiwalled carbon nanotube nanocomposites as electron mediator for the determination of glucose

A kind of nanocomposite with good dispersion in water was prepared through covalent adsorption of ferrocenecarboxaldehyde on multiwalled carbon nanotubes (MWNTs) for electrical communication between glucose oxidase (GOD) and electrode. The ferrocene-modified multiwalled carbon nanotube nanocomposites (MWNTs-Fc) could be conveniently cast on electrode surfaces. With the aid of chitosan, GOD was then immobilized on the nanostructure film to form a reagentless amperometric sensor for glucose determination. FTIR spectra and cyclic voltammetry were used to characterize the nanocomposites. The presence of both ferrocene as mediator of electron transfer and MWNTs as conductor enhanced greatly the enzymatic response to the oxidation of glucose. The novel biosensor exhibited a fast response toward glucose with a detection limit of 3.0 × 10⁻⁶ mol/L and the linear range extended up to 3.8 × 10⁻³ mol/L.     

Glucose biosensor prepared by glucose oxidase encapsulated sol-gel and carbon-nanotube-modified basal plane pyrolytic graphite electrode

A new glucose biosensor has been fabricated by immobilizing glucose oxidase into a sol-gel composite at the surface of a basal plane pyrolytic graphite (bppg) electrode modified with multiwall carbon nanotube. First, the bppg electrode is subjected to abrasive immobilization of carbon nanotubes by gently rubbing the electrode surface on a filter paper supporting the carbon nanotubes. Second, the electrode surface is covered with a thin film of a sol-gel composite containing encapsulated glucose oxidase. The carbon nanotubes offer excellent electrocatalytic activity toward reduction and oxidation of hydrogen peroxide liberated in the enzymatic reaction between glucose oxidase and glucose, enabling sensitive determination of glucose. The amperometric detection of glucose is carried out at 0.3 V (vs saturated calomel electrode) in 0.05 M phosphate buffer solution (pH 7.4) with linear response range of 0.2-20 mM glucose, sensitivity of 196 nA/mM, and detection limit of 50 microM (S/N=3). The response time of the electrode is < 5s when it is stored dried at 4 degrees C, the sensor showed almost no change in the analytical performance after operation for 3 weeks. The present carbon nanotube sol-gel biocomposite glucose oxidase sensor showed excellent properties for the sensitive determination of glucose with good reproducibility, remarkable stability, and rapid response and in comparison to bulk modified composite biosensors the amounts of enzyme and carbon nanotube needed for electrode fabrication are dramatically decreased.     

Amperometric glucose biosensor based on multilayer films via layer-by-layer self-assembly of multi-wall carbon nanotubes, gold nanoparticles and glucose oxidase on the Pt electrode

A novel amperometric glucose biosensor based on the nine layers of multilayer films composed of multi-wall carbon nanotubes (MWCNTs), gold nanoparticles (GNp) and glucose oxidase (GOD) was developed for the specific detection of glucose. MWCNTs were chemically modified with the H₂SO₄–HNO₃ pretreatment to introduce carboxyl groups which were used to interact with the amino groups of poly(allylamine) (PAA) and cysteamine via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide cross-linking reaction, respectively. A cleaned Pt electrode was immersed in PAA, MWCNTs, cysteamine and GNp, respectively, followed by the adsorption of GOD, assembling the one layer of multilayer films on the surface of Pt electrode (GOD/GNp/MWCNTs/Pt electrode). Repeating the above process could assemble different layers of multilayer films on the Pt electrode. PBS washing was applied at the end of each assembly deposition for dissociating the weak adsorption. Film assembling and characterization were studied by transmission electron microscopy and quartz crystal microbalance, and properties of the resulting glucose biosensors were measured by electrochemical measurements. The marked electrocatalytic activity of Pt electrode based on multilayer films toward H₂O₂ produced during GOD enzymatic reactions with glucose permitted effective low-potential amperometric measurement of glucose. Taking the sensitivity and selectivity into consideration, the applied potential of 0.35 V versus Ag/AgCl was chosen for the oxidation detection of H₂O₂ in this work. Among the resulting glucose biosensors, the biosensor based on nine layers of multilayer films was best. It showed a wide linear range of 0.1–10 mM glucose, with a remarkable sensitivity of 2.527 μA/mM, a detection limit of 6.7 μM estimated at a signal-to-noise ratio of 3 and fast response time (within 7 s). Moreover, it exhibited good reproducibility, long-term stability and the negligible interferences of ascorbic acid, uric acid and acetaminophen. The study can provide a feasible approach on developing new kinds of oxidase-based amperometric biosensors, and can be used as an illustration for constructing various hybrid structures.     

DNA biosensor based on chitosan film doped with carbon nanotubes

A biosensor based on chitosan doped with carbon nanotube (CNT) was fabricated to detect salmon sperm DNA. Methylene blue (MB) was employed as a DNA indicator. It was found that CNTs can enhance the electroactive surface area threefold (0.28 +/- 0.03 and 0.093 +/- 0.06 cm(2) for chitosan-CNT- and chitosan-modified electrodes, respectively) and can accelerate the rate of electron transfer between the redox-active MB and the electrode. A low detection limit of 0.252 nM fish sperm DNA was achieved, and no interference was found in the presence of 5 microg/ml human serum albumin. The differential pulse voltammetry signal of MB was linear over the fish sperm DNA concentration range of 0.5-20 nM.     

Polycrystalline bismuth oxide films for development of amperometric biosensor for phenolic compounds

An attractive biocomposite based on polycrystalline bismuth oxide (BiO_x) film and polyphenol oxidase (PPO) was proposed for the construction of a mediator-free amperometric biosensor for phenolic compounds in environmental water samples. The phenolic biosensor could be easily achieved by casting the biocomposite on the surface of glassy carbon electrode (GCE) via the cross-linking step by glutaraldehyde. The laboratory-prepared bismuth oxide semiconductor was polymorphism. Its hydrophilicity provided a favorable microenvironment for retaining the biological activity of the immobilized protein. The parameters of the fabrication process and the various experimental variables for the enzyme electrode were optimized. The proposed PPO/BiO_x biosensor provided a linear response to catechol over a concentration range of 4 × 10⁻⁹ M to 1.5 × 10⁻⁵ M with a dramatically developed sensitivity of 11.3 A M⁻¹ cm⁻² and a detection limit of 1 × 10⁻⁹ M based on S/N = 3. In addition, the PPO/BiO_x biocomposite was characterized by scanning electron microscope (SEM), Fourier transform infrared spectra (FTIR) and rotating disk electrode voltammetry.     

Carbon nanotubes promote Cr(VI) reduction by alginate-immobilized Shewanella oneidensis MR-1

Bioreduction of hexavalent chromium (Cr(VI)) into trivalent one (Cr(III)) based on microbial immobilization techniques has been recognized as a promising way to remove Cr contaminants from wastewater. However, such a bioreduction process is inefficient due to limited electron transfer through the immobilization matrix. In this study, a modified immobilization process was proposed by impregnating carbon nanotubes (CNTs) into Ca-alginate beads, which were then used to immobilize Shewanella oneidensis MR-1 for enhanced Cr(VI) reduction. Compared with the free cells and the beads without CNTs, the AL/CNT/cell beads showed up to 4 times higher reduction rates, mainly attributed to an enhanced electron transfer by the CNTs. In addition, the dose of CNTs greatly improved the stability of beads, suggesting a high feasibility of the AL/CNT/cell beads for repeated use. The optimized CNT concentration, temperature and pH for Cr(VI) reduction by the AL/CNT/cell beads were 0.5%, 30 °C and 6.0–7.0, respectively.     

Mediator-free electrochemical biosensor based on buckypaper with enhanced stability and sensitivity for glucose detection

Here we report on a novel platform based on buckypaper for the design of high-performance electrochemical biosensors. Using glucose oxidase as a model enzyme, we constructed a biocompatible mediator-free biosensor and studied the potential effect of the buckypaper on the stability of the biosensor with both amperometry and FTIR spectroscopy. The results showed that the biosensor responses sensitively and selectively to glucose with a considerable functional lifetime of over 80 days. The fabricated enzymatic sensor detects glucose with a dynamic linear range of over 9 mM and a detection limit of 0.01 mM. To examine the efficiency of enzyme immobilization, the Michaelis–Menten constant was calculated to be 4.67 mM. In addition, the fabricated electrochemical biosensor shows high selectivity; no amperometric response to the common interference species such as ascorbic acid, uric acid and acetamidophenol was observed. The facile and robust buckypaper-based platform proposed in this study opens the door for the design of high-performance electrochemical biosensors for medical diagnostics and environmental monitoring.     

Heatable carbon nanotube composite membranes for sustainable recovery from biofouling

Membrane filtration is one of the most reliable methods for water treatment. However, wider application is limited due to biofouling caused by accumulation of microorganisms on the membrane surface. This report details a heatable carbon nanotube composite membrane with self-cleaning properties for sustainable recovery from biofouling. Microfiltration polycarbonate/carbon-nanotubes hybrid membranes were fabricated using drawable nanotubes that maintained the porosity and provided electrical conductivity to the membrane. Less than 25 V potential and 2–3 W power increase membrane temperature to 100°C in ~10 s. This temperature is above what most microbial life, bacteria and viruses can handle. When this membrane was employed, filtered Escherichia coli collected on its surface were successfully annihilated within 1 min. Ohmic heating of this membrane could be an effective solution to combat biofouling and complications associated with membrane-based filtration. This is a novel and highly desirable approach to combat biofouling, due to its simplicity and economic advantage.