A biosensing platform based on horseradish peroxidase immobilized onto chitosan-wrapped single-walled carbon nanotubes

One-step, diameter-selective dispersion of single-walled carbon nanotubes (SWCNTs) has been accomplished through noncovalent complexation of the nanotubes with a water-soluble, biocompatible polymer chitosan at room temperature. Such chitosan-wrapped individual SWCNTs can be used for the immobilization of horseradish peroxidase (HRP) and be used to construct an electrode for direct bioelectrochemical sensing without an electron mediator. The direct electron transfer between HRP and the electrode surface was observed with a formal potential of approximately −0.35 V (vs. saturated calomel electrode) in phosphate buffer solution and the calculated heterogeneous electron transfer rate constant is approximately 23.5 s⁻¹. Experimental results indicate that the immobilized HRP retains its catalytic activity for the reduction of nitric oxide. Such an HRP–SWCNT–chitosan-based biosensor exhibited a rapid response time of less than 6 s and a good linear detection range for nitrite concentration, from 25 to 300 μM with a detection limit of 3 μM. The apparent Michaelis–Menten constant (K _m) and the maximum electrode sensitivity (i_max/K _m) are found to be 7.0 mM and 0.16 μA mM⁻¹, respectively. Both the unique electrical properties of SWCNTs and biocompatibility of chitosan enable the construction of an excellent biosensing platform for improved electrocatalysis of HRP, allowing, specifically, the detection of trace levels of nitric oxide.     

Cu2+-doped polypyrrole nanotubes with promoted efficiency for peroxidase mimicking and electrochemical biosensing

A facile template together with doping strategy is presented to fabricate Cu²⁺-doped polypyrrole (Cu²⁺/PPy) nanotubes (NTs) as efficient mimicking peroxidase and electrocatalyst. PPy NTs were first prepared using electrospun polyacrylonitrile nanofibers as templates and subsequently doped with Cu²⁺ via a simple immersion strategy. The as-prepared Cu²⁺/PPy NTs not only exhibit an outstanding peroxidase-like catalytic efficiency but also possess an excellent electrocatalytic activity, which is used for the detection of glucose, displaying a great promise for biosensing applications. The good enzyme-like and electrochemical performances of Cu²⁺/PPy NTs are owing to the reduced Fermi level compared with bare PPy NTs, which is beneficial for promoting the electron transfer to the substrate. The novel Cu²⁺/PPy NTs are a novel type of enzyme mimics and electrocatalysts for potential bright applications in biotechnology and environment science.     

Direct electron transfer of horseradish peroxidase and its electrocatalysis based on carbon nanotube/thionine/gold composites

Horseradish peroxidase (HRP) was incorporated into multiwalled carbon nanotube/thionine/Au (MTAu) composite film by electrostatic interactions between positively charged HRP and negatively charged MTAu composite. The results of electrochemical impedance spectroscopy (EIS) confirmed adsorption of HRP on the surface of MTAu modified GC electrode. Moreover, the electrochemical results showed that HRP retained its bioactivity and bioelectrocatalytical activity, and also showed good direct electron transfer behavior on such a composite film.     

Carbon nanotubes for electrochemical biosensing

The aim of this review is to summarize the most relevant contributions in the development of electrochemical (bio)sensors based on carbon nanotubes in the last years.Since the first application of carbon nanotubes in the preparation of an electrochemical sensor, an increasing number of publications involving carbon nanotubes-based sensors have been reported, demonstrating that the particular structure of carbon nanotubes and their unique properties make them a very attractive material for the design of electrochemical biosensors.The advantages of carbon nanotubes to promote different electron transfer reactions, in special those related to biomolecules; the different strategies for constructing carbon nanotubes-based electrochemical sensors, their analytical performance and future prospects are discussed in this article.     

Shape-coded silica nanotubes for biosensing

Shape-coded silica nanotubes (SNTs) were fabricated on the basis of template synthesis as a new dispersible microarray system. The template synthesis of shape-coded SNTs begins with the fabrication of a porous alumina film that has well-defined cylindrical pores with two or more different diameter segments by multistep anodization of an aluminum substrate. Then, SNTs were fabricated with a surface sol-gel method that can control the wall thickness of SNTs on the single-nanometer level. Attractively, the difference in optical reflectance between the segmented parts of individual silica nanotube makes it very convenient to identify each nanotube and enables these shape-coded SNTs to work as coding materials for biosensing.     

Coadsorption of n monomer species on terraces and nanotubes

The knowledge of the partition function, Z, of a system of particles adsorbed on a surface is all that is required to determine the occupational characteristics of the adsorbates and the thermodynamic properties of the system. The surface considered is a terrace or a nanotube of arbitrary periodic lattice geometry, L atomic sites in length, and M' sites in the width of the terrace or in the normal cross section of the nanotube. The matrix method introduced in 2007 to obtain Z for the adsorption study of one species of monomers is now generalized to the study of the coadsorption of any number, n, of monomer species. We provide proof that Z can be related to the eigenvalues of a real and non-negative matrix (T matrix) of rank (n + 1)(M), where M is an integer multiple of M'. In the infinite-L limit, we also prove that Z is the largest eigenvalue of the T matrix, raised to the power of (1)/(M). Because the rank of this matrix increases exponentially with M, we develop a technique for its recursive construction applicable to any lattice geometry, which is easily programmed and efficiently adaptable for supercomputing and multiparallel processing. As examples, we consider the coadsorption on square, equilateral triangular, and honeycomb surfaces. This general formulation can now be applied to model a whole new set of experiments involving the coadsorption of two or more monomer species, on terrace or nanotube surfaces with various periodic lattice structures.     

Amperometric biosensor for hydrogen peroxide based on coimmobilized horseradish peroxidase and methylene green in ormosils matrix with multiwalled carbon nanotubes

A novel amperometric biosensor for the analytical determination of hydrogen peroxide was developed. The fabrication of the biosensor was based on the coimmobilization of horseradish peroxidase (HRP), methylene green (MG) and multiwalled carbon nanotubes within ormosils; 3-aminopropyltrimethoxysilane (APTMOS), 2-(3,4-epoxycyclohexyl)ethyltrimethoxysilane (ETMOS) and phenyltrimethoxysilane (PHTMOS). APTMOS determined the hydrophilicity/hydrophobicity of the ormosils and PHTMOS and ETMOS increased the physical and mechanical strength of the ormosil matrix. The ormosil modified electrodes were characterized with SEM, UV-vis spectroscopy and electrochemical methods. Cyclic voltammetry and amperometric measurements demonstrated the MG coimmobilized with HRP in this way, displayed good stability and could efficiently shuttle electrons between immobilized enzyme and electrode, and MWCNTs facilitated the electrocatalytic reduction of H₂O₂ at reduced over potential. The Micheaelis constant of the immobilized HRP was 1.8 mM, indicating a high affinity of the HRP to H₂O₂ without loss of enzymatic activity in ormosil matrix. The prepared biosensor had a fast response of H₂O₂, less than 10 s, and excellent linear range of concentration from 5 × 10⁻⁷ to 2 × 10⁻⁵ M with the detection limit of 0.5 μM (S/N = 3) under the optimum conditions. At the same time, the influence of solution pH, effect of enzyme amount, steady-state applied potential and temperature on the biosensor were investigated. The enzyme electrode retained about 90% of its initial activity after 30 days of storage in a dry state at 4 °C. The preparation of the developed biosensor was convenient and showed high sensitivity with good stability.     

Electric field directed layer-by-layer assembly of horseradish peroxidase nanotubes via anodic aluminum oxide template

A novel method based on electric field directed layer-by-layer assembly (EFDLA) and template synthesis is successfully developed to fabricate horseradish peroxidase (HRP) nanotubes. It provides a rapid and general strategy for functional protein nanoarrays. The alternative deposition of poly(diallyldimethylammonium chloride) (PDDA) and HRP are characterized by SEM, TEM, UV–vis and electrochemical impedance spectroscopy (EIS). Moreover, the UV–vis spectrometry and electrochemistry of HRP indicate that immobilization of PDDA/HRP via EFDLA coupled with template synthesis could retain the conformations and bioactivities of HRP.     

Electrochemical biosensors based on horseradish peroxidase

The principles used for the development of electrochemical biosensors based on horseradish peroxidase are described. Peroxidase is the enzyme which catalyses the oxidation of a variety of organic molecules in the presence of hydrogen peroxide. The features of this enzyme are high catalytic activity and low specificity towards second substrate as well. Horseradish peroxidase may be used as a component of active part of biosensors for the detection of hydrogen peroxide and other compounds when peroxidase is co-immobilized together with other oxidases. Also horseradish peroxidase may be used as a component of detecting system for the biosensors based on biological recognition using specific antibodies, receptors, nucleic acids. The examples of the bio-, immuno-, DNA-sensors developed for the determination of various biologically active compounds are given.     

Activity and stability of horseradish peroxidase in hydrophilic room temperature ionic liquid and its application in non-aqueous biosensing

The activity and stability of horseradish peroxidase (HRP) were investigated in a hydrophilic room temperature ionic liquid 1-butyl-3-methylimidazolium tetrafluroborate ([bmim][BF₄]) by electrochemical methods. Although no detectable activity exhibited in anhydrous [bmim][BF₄], HRP was active in the presence of a small amount of water (4.53%, v/v). And its activity can be improved by immobilization in agarose hydrogel. The immobilized HRP possesses excellent activity at 65 °C. It remained 80.2% of its initial activity after being immersed for 10.5 h in an aqueous mixture of [bmim][BF₄] with some hydrogen peroxide (H₂O₂) under room temperature, implying extremely high stability. Moreover, the immobilized HRP was found to be very sensitive and stable in H₂O-containing [bmim][BF₄] for the detection of H₂O₂, with a wide linear range of 6.10 × 10⁻⁷ to 1.32 × 10⁻⁴ mol l⁻¹ and low detection limit of 1.0 × 10⁻⁷ mol l⁻¹.     

Amperometric sensor for hydrogen peroxide based on electric wire composed of horseradish peroxidase and toluidine blue-multiwalled carbon nanotubes nanocomposite

A kind of nanocomposites with good dispersion in water was prepared through noncovalent adsorption of toluidine blue (Tb) on multiwalled carbon nanotubes (MWCNT) for electric communication between horseradish peroxidase (HRP) and electrode. The nanocomposites could be conveniently cast on electrode surface. With the aid of chitosan, HRP was then immobilized on the nanostructure to form a reagentless amperometric sensor for hydrogen peroxide. UV-vis spectroscopy and electrochemical impedance spectroscopy were used to characterize the adsorption of Tb on MWCNT. The presence of both Tb as mediator of electron transfer and MWCNT as conductor enhanced greatly the enzymatic response to the reduction of hydrogen peroxide. The novel biosensor exhibited fast response towards hydrogen peroxide with a detection limit of 1.7x10(-6)M and the linear range extended up to 4x10(-4)M without the interference of ascorbic acid and uric acid. The Michaelis-Menten constant (K'(m)) of the immobilized HRP was evaluated to be 0.16mM.     

Small diameter TiO2 nanotubes with enhanced photoresponsivity

We report ultraviolet (UV) light detection of thin wall TiO₂ nanotubes (TNTs) with open diameter ~ 20 nm obtained by a two anodization procedure. This small diameter nanotubular geometry shows significant enhancement of the photoresponsivity and results in a large increase of photocurrent. The photocurrent is one order higher than that of classical nanotubes with diameter of 140 nm at − 1.0 V bias. We attribute this improvement to the modulation of hole carrier density as a result of field effects from the diameter-dependent population of the surface-trapped electrons. This finding demonstrates inherent size effects of internal gain in semiconductor nanotubes.     

Functionalized carbon nanotubes and nanofibers for biosensing applications

This review summarizes recent advances in electrochemical biosensors based on carbon nanotubes (CNTs) and carbon nanofibers (CNFs) with an emphasis on applications of CNTs. CNTs and CNFs have unique electric, electrocatalytic and mechanical properties, which make them efficient materials for developing electrochemical biosensors.We discuss functionalizing CNTs for biosensors. We review electrochemical biosensors based on CNTs and their various applications (e.g., measurement of small biological molecules and environmental pollutants, detection of DNA, and immunosensing of disease biomarkers). Moreover, we outline the development of electrochemical biosensors based on CNFs and their applications. Finally, we discuss some future applications of CNTs.     

A direct electrochemical biosensing platform constructed by incorporating carbon nanotubes and gold nanoparticles onto redox poly(thionine) film.

A direct electrochemical biosensing platform has been fabricated by covalent incorporation of carbon nanotubes (CNT) and gold nanoparticles (GNP) onto the poly(thionine) (PTH) film deposited by electropolymerization. With the synergic effects of the composite nanomaterials together with the excellent mediating redox polymer, the proposed platform could allow for faster electron transfer and higher enzyme immobilization efficiency than the platforms designed by using CNT or GNP alone. Comparison studies indicated that the as-developed H(2)O(2) sensor could show greatly improved performances of amperometric responses.     

Some critical factors for photocatalysis on self-organized TiO2 nanotubes

In the present work, different intrinsic and extrinsic parameters are investigated that affect the photocatalytic activity of self-organized TiO₂ nanotube layers. Particularly, the influence of annealing temperature and annealing atmosphere, the influence of different gas purging in the electrolyte, and the effect of applied voltage on the photocatalytic degradation rates of acid orange (AO7) are discussed. We find that the effect of the reducing gas atmosphere dominates over the anatase/rutile ratio in activating the nanotube layers. Moreover, we show that the effect of different gas purging (Ar and O₂) of the electrolyte affects the reaction rate twofold: (1) by providing electron acceptor states and also by (2) a different change in the red–ox potential, i.e., the band bending in TiO₂. By an external anodic voltage, the reaction rates can be increased drastically due to increased band bending. Nevertheless, the magnitude of the effect is also affected by the presence or absence of O₂ in the electrolyte.     

TiO2 nanotube arrays fabricated by anodization in different electrolytes for biosensing

Titania nanotube arrays were fabricated by anodic oxidation of titanium foil in different electrolytes. The morphology, crystallinity and composition of the as-prepared nanotube arrays were studied by XRD, SEM and EDX. Electrochemical impedance spectroscopy (EIS) was employed to investigate their electrical conductivity and capacitance. Titania nanotube arrays co-adsorbed with horseradish peroxidase (HRP) and thionine chloride (Th) were studied for their sensitivity to hydrogen peroxide by means of cyclic voltammetric and galvanostatic measurements. The experiments showed that TiO₂ nanotube arrays possessed appreciably different sensitivities to H₂O₂ due to their different conductivity. Further experiments revealed that TiO₂ nanotubes have noticeably different ability of adsorbing HRP and Th, and the best sensitivity was achieved when the density of HRP is the highest. The TiO₂ nanotube arrays fabricated in potassium fluoride solution demonstrated the best sensitivity on hydrogen peroxide in the range of 10⁻⁵–3 × 10⁻³ M at pH 6.7 and at a potential of −600 mV (vs. Ag/AgCl).     

Titania nanotubes decorated with gold nanoparticles for electrochemiluminescent biosensing of glycosylated hemoglobin

A glycated hemoglobin (HbA1c) biosensor with high performance has been constructed in this work. Here the fructosyl amino acid oxidase was immobilized onto a pre-functionalized indium tin oxide glass with titania nanotubes decorated with gold nanoparticles. The property of nanocomposite was characterized by transmission electromicroscopy, scanning electron microscopy, electrochemistry and spectroscopy. Under the optimum conditions, fructosyl valine was detected by this biosensor. It exhibited a linear detection range from 4.0 × 10⁻⁹ M to 7.2 × 10⁻⁷ M, and a limit of detection for 3.8 × 10⁻⁹ M at the signal-to-noise ratio of 3. Thus the HbA1c level in whole blood samples of healthy individuals or diabetic patients were evaluated with designed biosensor after pre-treatment of hydrolysis. The results of our detection were closely consistent with that of the standard method. At the same time, our biosensor has some advantages including high sensitivity, disposable usage and low cost, which implies its great promising application in point-of-care testing of HbA1c.