Detection of glucose using immobilized bienzyme on cyclic bisureas–gold nanoparticle conjugate

A highly sensitive electrochemical glucose sensor has been developed by the co-immobilization of glucose oxidase (GOx) and horseradish peroxidase (HRP) onto a gold electrode modified with biocompatible cyclic bisureas–gold nanoparticle conjugate (CBU–AuNP). A self-assembled monolayer of mercaptopropionic acid (MPA) and CBU–AuNP was formed on the gold electrode through a layer-by-layer assembly. This modified electrode was used for immobilization of the enzymes GOx and HRP. Both the HRP and GOx retained their catalytic activity for an extended time, as indicated by the low value of Michaelis–Menten constant. Analytical performance of the sensor was examined in terms of sensitivity, selectivity, reproducibility, lower detection limit, and stability. The developed sensor surface exhibited a limit of detection of 100 nM with a linear range of 100 nM to 1 mM. A high sensitivity of 217.5 μA mM⁻¹ cm⁻² at a low potential of −0.3 V was obtained in this sensor design. Various kinetic parameters were calculated. The sensor was examined for its practical clinical application by estimating glucose in human blood sample.     

Attachment of gold nanoparticles to glassy carbon electrode and its application for the direct electrochemistry and electrocatalytic behavior of hemoglobin

Gold nanoparticles have been attached onto glassy carbon electrode surface through sulfhydryl-terminated monolayer and characterized by X-ray photoelectron spectroscopy, atomic force microscopy, electrochemical impedance spectroscopy and cyclic voltammetry. The gold nanoparticles-attached glassy carbon electrodes have been applied to the immobilization/adsorption of hemoglobin, with a monolayer surface coverage of about 2.1 x 10(-10) mol cm(-2), and consequently obtained the direct electrochemistry of hemoglobin. Gold nanoparticles, acting as a bridge of electron transfer, can greatly promote the direct electron transfer between hemoglobin and the modified glassy carbon electrode without the aid of any electron mediator. In phosphate buffer solution with pH 6.8, hemoglobin shows a pair of well-defined redox waves with formal potential (E0') of about -0.085 V (versus Ag/AgCl/saturated KCl). The immobilized hemoglobin maintained its biological activity, showing a surface controlled electrode process with the apparent heterogeneous electron transfer rate constant (ks) of 1.05 s(-1) and charge-transfer coefficient (a) of 0.46, and displays the features of a peroxidase in the electrocatalytic reduction of hydrogen peroxide. A potential application of the hemoglobin-immobilized gold nanoparticles modified glassy carbon electrode as a biosensor to monitor hydrogen peroxide has been investigated. The steady-state current response increases linearly with hydrogen peroxide concentration from 2.0 x 10(-6) to 2.4 x 10(-4) M. The detection limit (3sigma) for hydrogen peroxide is 9.1 x 10(-7) M.     

A novel glucose biosensor using bi-enzyme incorporated with peptide nanotubes

A novel amperometric glucose biosensor was developed using the bio-inspired peptide nanotube (PNT) as an encapsulation template for enzymes. Horseradish peroxidase (HRP) was encapsulated by the PNT and glucose oxidase (GO(x)) was co-immobilized with the PNT on a gold nanoparticle (AuNP)-modified electrode. A binary SAM of 3-mercaptopropionic acid (MPA) and 1-tetradecanethiol (TDT) was formed on the surface of the electrode to immobilize the PNT and GO(x). The resulting electrode appeared to provide the enzymes with a biocompatible nanoenvironment as it sustained the enhanced enzyme activity for an extended time and promoted possible direct electron transfer through the PNT to the electrode. Performance of the biosensor was evaluated in terms of its detection limit, sensitivity, pH, response time, selectivity, reproducibility, and stability in a lab setting. In addition the sensor was tested for real samples. The composite of AuNP-SAM-PNT/HRP-GO(x) to fabricate a sensor electrode in this study exhibited a linear response with glucose in the concentration range of 0.5-2.4mM with a R(2)-value of 0.994. A maximum sensitivity of 0.3mAM(-1)and reproducibility (RSD) of 1.95% were demonstrated. The PNT-encapsulated enzyme showed its retention of >85% of the initial current response after one month of storage.     

A novel method to construct a third-generation biosensor: self-assembling gold nanoparticles on thiol-functionalized poly(styrene-co-acrylic acid) nanospheres

A novel method for fabrication of horseradish peroxidase (HRP) biosensor has been developed by self-assembling gold nanoparticles on thiol-functionalized poly(styrene-co-acrylic acid) (St-co-AA) nanospheres. At first, a cleaned gold electrode was immersed in thiol-functionalized poly(St-co-AA) nanosphere latex prepared by emulsifier-free emulsion polymerization of St with AA and function with dithioglycol to assemble the nanospheres, then gold nanoparticles were chemisorbed onto the thiol groups. Finally, horseradish peroxidase was immobilized on the surface of the gold nanoparticles. The sensor displayed an excellent electrocatalytical response to reduction of H2O2 without the aid of an electron mediator. The sensor was highly sensitive to hydrogen peroxide with a detection limit of 4.0 micromoll(-1), and the linear range was from 10.0 micromoll(-1) to 7.0 mmoll(-1). The biosensor retained more than 97.8% of its original activity after 60 days of use. Moreover, the studied biosensor exhibited good current repeatability and good fabrication reproducibility.     

Oligonucleotide probes applied for sensitive enzyme-amplified electrochemical assay of mercury(II) ions

We developed a novel electrochemical sensor for Hg(2+) detection using two mercury-specific oligonucleotide probes and streptavidin-horseradish peroxidase (HRP) enzymatic signal amplification. The two mercury-specific oligonucleotide probes comprised a thiolated capture probe and a biotinated signal probe. The thiolated capture probe was immobilized on a gold electrode. In the presence of Hg(2+), the thymine-Hg(2+)-thymine (T-Hg(2+)-T) interaction between the mismatched T-T base pairs directed the biotinated signal probe hybridizing to the capture probe and yielded a biotin-functioned electrode surface. HRP was then immobilized on the biotin-modified substrate via biotin-streptavidin interaction. The immobilized HRP catalyzed the oxidation of hydroquinone (H(2)Q) to benzoquinone (BQ) by hydrogen peroxide (H(2)O(2)) and the generated BQ was further electrochemically reduced at the modified gold electrode, producing a readout signal for quantitative detection of Hg(2+). The results showed that the enzyme-amplified electrochemical sensor system was highly sensitive to Hg(2+) in the concentration of 0.5 nM to 1 μM with a detection limit of 0.3 nM, and it also demonstrated excellent selectivity against other interferential metal ions.     

Design and development of a highly stable hydrogen peroxide biosensor on screen printed carbon electrode based on horseradish peroxidase bound with gold nanoparticles in the matrix of chitosan

The design and development of a screen printed carbon electrode (SPCE) on a polyvinyl chloride substrate as a disposable sensor is described. Six configurations were designed on silk screen frames. The SPCEs were printed with four inks: silver ink as the conducting track, carbon ink as the working and counter electrodes, silver/silver chloride ink as the reference electrode and insulating ink as the insulator layer. Selection of the best configuration was done by comparing slopes from the calibration plots generated by the cyclic voltammograms at 10, 20 and 30 mM K(3)Fe(CN)(6) for each configuration. The electrodes with similar configurations gave similar slopes. The 5th configuration was the best electrode that gave the highest slope. Modifying the best SPCE configuration for use as a biosensor, horseradish peroxidase (HRP) was selected as a biomaterial bound with gold nanoparticles (AuNP) in the matrix of chitosan (HRP/AuNP/CHIT). Biosensors of HRP/SPCE, HRP/CHIT/SPCE and HRP/AuNP/CHIT/SPCE were used in the amperometric detection of H(2)O(2) in a solution of 0.1M citrate buffer, pH 6.5, by applying a potential of -0.4V at the working electrode. All the biosensors showed an immediate response to H(2)O(2). The effect of HRP/AuNP incorporated with CHIT (HRP/AuNP/CHIT/SPCE) yielded the highest performance. The amperometric response of HRP/AuNP/CHIT/SPCE retained over 95% of the initial current of the 1st day up to 30 days of storage at 4 degrees C. The biosensor showed a linear range of 0.01-11.3mM H(2)O(2), with a detection limit of 0.65 microM H(2)O(2) (S/N=3). The low detection limit, long storage life and wide linear range of this biosensor make it advantageous in many applications, including bioreactors and biosensors.     

Direct electrochemistry of horseradish peroxidase immobilized on a colloid/cysteamine-modified gold electrode

Direct electron transfer of immobilized horseradish peroxidase on gold colloid and its application as a biosensor were investigated by using electrochemical methods. The Au colloids were associated with a cysteamine monolayer on the gold electrode surface. A pair of redox peaks attributed to the direct redox reaction of horseradish peroxidase (HRP) were observed at the HRP/Au colloid/cysteamine-modified electrode in 0.1 M phosphate buffer (pH 7.0). The surface coverage of HRP immobilized on Au colloid was about 7.6 x 10(-10) mol/cm(2). The sensor displayed an excellent electrocatalytic response to the reduction of H(2)O(2) without the aid of an electron mediator. The calibration range of H(2)O(2) was 1. 4 microM to 9.2 mM with good linear relation from 1.4 microM to 2.8 mM. A detection limit of 0.58 microM was estimated at a signal-to-noise ratio of 3. The sensor showed good reproducibility for the determination of H(2)O(2). The variation coefficients were 3. 1 and 3.9% (n = 10) at 46 microM and 2.8 mM H(2)O(2), respectively. The response showed a Michaelis-Menten behavior at higher H(2)O(2) concentrations. The K(app)(M) value for the H(2)O(2) sensor was found to be 2.3 mM.     

Multienzyme-nanoparticles amplification for sensitive virus genotyping in microfluidic microbeads array using Au nanoparticle probes and quantum dots as labels

A novel microfluidic device with microbeads array was developed and sensitive genotyping of human papillomavirus was demonstrated using a multiple-enzyme labeled oligonucleotide-Au nanoparticle bioconjugate as the detection tool. This method utilizes microbeads as sensing platform that was functionalized with the capture probes and modified electron rich proteins, and uses the horseradish peroxidase (HRP)-functionalized gold nanoparticles as label with a secondary DNA probe. The functionalized microbeads were independently introduced into the arrayed chambers using the loading chip slab. A single channel was used to generate weir structures to confine the microbeads and make the beads array accessible by microfluidics. Through "sandwich" hybridization, the enzyme-functionalized Au nanoparticles labels were brought close to the surface of microbeads. The oxidation of biotin-tyramine by hydrogen peroxide resulted in the deposition of multiple biotin moieties onto the surface of beads. This deposition is markedly increased in the presence of immobilized electron rich proteins. Streptavidin-labeled quantum dots were then allowed to bind to the deposited biotin moieties and displayed the signal. Enhanced detection sensitivity was achieved where the large surface area of Au nanoparticle carriers increased the amount HRP bound per sandwiched hybridization. The on-chip genotyping method could discriminate as low as 1fmol/L (10zmol/chip, SNR>3) synthesized HPV oligonucleotides DNA. The chip-based signal enhancement of the amplified assay resulted in 1000 times higher sensitivity than that of off-chip test. In addition, this on-chip format could discriminate and genotype 10copies/μL HPV genomic DNA using the PCR products. These results demonstrated that this on-chip approach can achieve highly sensitive detection and genotyping of target DNA and can be further developed for detection of disease-related biomolecules at the lowest level at their earliest incidence.     

Covalent immobilization of cholesterol oxidase on self-assembled gold nanoparticles for highly sensitive amperometric detection of cholesterol in real samples

A novel scheme for the fabrication of gold nanoparticle modified cholesterol oxidase based bioelectrode is presented and its application potential for cholesterol biosensor is investigated. The fabrication procedure is based on the deposition of gold nanoparticles on the 1,6-hexanedithiol modified gold electrode, functionalization of the surface of deposited gold nanoparticles with carboxyl groups using 11-mercaptoundecanoic acid and then covalent immobilization of cholesterol oxidase on the surface of gold nanoparticle film using the N-ethyl-N'-(3-dimethylaminopropyl carbodimide) and N-hydroxysuccinimide ligand chemistry. The assembly process of the bioelectrode is investigated using atomic force microscopy, cyclic voltammetry and electrochemical impedance spectroscopy. The gold nanoparticle film on the electrode surface provided an environment for the enhanced electrocatalytic activities and thus resulted in enhanced analytical response. The resulting bioelectrode is further applied to the amperometric detection of cholesterol and exhibited a linear response to cholesterol in the range of 0.04-0.22 mM with a detection limit of 34.6 μM, apparent Michaelis-Menten constant (K(m)(app)) of 0.062 mM and a high sensitivity of 9.02 μA mM(-1). The fabricated bioelectrode is successfully used for the selective determination of cholesterol in human serum samples.     

A third-generation H2O2 biosensor based on horseradish peroxidase-labeled Au nanoparticles self-assembled to hollow porous polymeric nanopheres

A novel third-generation biosensor for hydrogen peroxide (H2O2) was developed by self-assembling gold nanoparticles to hollow porous thiol-functionalized poly(divinylbenzene-co-acrylic acid) (DVB-co-AA) nanospheres. At first, a cleaned gold electrode was immersed in hollow porous thiol-functionalized poly(DVB-co-AA) nanosphere latex to assemble the nanospheres, then gold nanoparticles were chemisorbed onto the thiol groups of the nanospheres. Finally, horseradish peroxidase (HRP) was immobilized on the surface of the gold nanoparticles. The immobilized horseradish peroxidase exhibited direct electrochemical behavior toward the reduction of hydrogen peroxide. The resulting biosensor showed a wide linear range of 1.0 microM-8.0mM and a detection limit of 0.5 microM estimated at a signal-to-noise ratio of 3. Moreover, the studied biosensor exhibited high sensitivity, good reproducibility, and long-term stability.     

Mediatorless biosensor for H(2)O(2) based on recombinant forms of horseradish peroxidase directly adsorbed on polycrystalline gold

Four forms of horseradish peroxidase (HRP) have been used to prepare peroxidase-modified gold electrodes for mediatorless detection of peroxide: native HRP, wild type recombinant HRP, and two recombinant forms containing six-His tag at the C-terminus and at the N-terminus, respectively. The adsorption of the enzyme molecules on gold was studied by direct mass measurements with electrochemical quartz crystal microbalance. All the forms of HRP formed a monolayer coverage of the enzyme on the gold surface. However, only gold electrodes with adsorbed recombinant HRP forms exhibited high and stable current response to H(2)O(2) due to its bioelectrocatalytic reduction based on direct electron transfer between gold and HRP. The sensitivity of the gold electrodes modified with recombinant HRPs was in the range of 1.4-1.5 A M(-1) cm(-2) at -50 mV versus Agmid R:AgCl. The response to H(2)O(2) in the concentration range 0.1-40 microM was not dependent on the presence of a mediator (i.e. catechol) giving strong evidence that the electrode currents are diffusion limited. Lower detection limit for H(2)O(2) detection was 10 nM at the electrodes modified with recombinant HRPs.     

A novel and simple biomolecules immobilization method: electro-deposition ZrO2 doped with HRP for fabrication of hydrogen peroxide biosensor

For the first time, a very novel and simple immobilization method for fabrication of hydrogen peroxide biosensor was reported in this paper. The biocompatible composite HRP-ZrO(2) thin films were synthesized on gold electrode surface based on electro-deposition zirconia doped with horseradish peroxidase (HRP) by cyclic voltammetry scanning in KCl solution containing ZrO(2) and HRP. The fabricated process of biosensor was characterized by electrochemical impedance spectroscopy (EIS) and the surface topography of the prepared films was imaged by atomic force microscope (AFM). The HRP in HRP-ZrO(2) thin films kept its bioactivity and exhibited excellent electrocatalytical response to the reduction of H(2)O(2). Experimental conditions influencing the biosensor performance such as pH, potential were optimized. The resulting biosensor (HRP-ZrO(2)/Au electrode) showed a linear response to H(2)O(2) over a concentration range from 0.02 to 9.45mM with a detection limit of 2muM based on a signal-to-noise ratio of 3 under optimized conditions. The apparent Michaelis-Menten constant (K(M)(app)) was evaluated to be 8.01mM, which indicated the HRP in HRP-ZrO(2) thin films kept its native bioactivity and had high affinity for H(2)O(2). Moreover, the proposed biosensor showed high sensitivity, good reproducibility and long-term stability. What is more, this immobilization methodology widened biosensor application in biomolecules immobilization and could further develop for other protein and biomolecules immobilization.     

Direct electrochemistry and electrocatalysis based on a film of horseradish peroxidase intercalated into Ni-Al layered double hydroxide nanosheets

Positively charged Ni-Al layered double hydroxide nanosheets (Ni-Al LDHNS) have been used for the first time as matrices for immobilization of horseradish peroxidase (HRP) in order to fabricate enzyme electrodes for the purpose of studying direct electron transfer between the redox centers of proteins and underlying electrodes. X-ray diffraction (XRD) and high-resolution transmission electron microscopy (HRTEM) revealed that the HRP-Ni-Al LDHNS film had an ordered structure and that HRP was intercalated into Ni-Al LDHNS with a monolayer arrangement. Field emission scanning electron microscopy (FESEM) showed that the HRP-Ni-Al LDHNS film had a uniform, porous morphology. UV-vis spectroscopy indicated that the intercalated HRP retained its native structure after incorporation in the Ni-Al LDHNS film. The immobilized HRP in Ni-Al LDHNS on the surface of a glassy carbon electrode (GCE) exhibited good direct electrochemical and electrocatalytic responses to the reduction of hydrogen peroxide (H(2)O(2)) and trichloroacetic acid (TCA). The resulting H(2)O(2) biosensor showed a wide linear range from 6.00x10(-7)M to 1.92x10(-4)M, low detection limit (4.00x10(-7)M) and good stability. The results show that Ni-Al LDHNS provide a novel and efficient platform for the immobilization of enzymes and realizing direct electrochemistry and that the materials have potential applications in the fabrication of third-generation biosensors.     

A label-free electrochemical immunoassay for carcinoembryonic antigen (CEA) based on gold nanoparticles (AuNPs) and nonconductive polymer film

A simple and sensitive label-free electrochemical immunoassay electrode for detection of carcinoembryonic antigen (CEA) has been developed. CEA antibody (CEAAb) was covalently attached on glutathione (GSH) monolayer-modified gold nanoparticle (AuNP) and the resulting CEAAb-AuNP bioconjugates were immobilized on Au electrode by electro-copolymerization with o-aminophenol (OAP). Electrochemical impedance spectroscopy and cyclic voltammetry studies demonstrate that the formation of CEA antibody-antigen complexes increases the electron transfer resistance of [Fe(CN)(6)](3-/4-) redox pair at the poly-OAP/CEAAb-AuNP/Au electrode. The use of CEA antibody-AuNP bioconjugates and poly-OAP film could enhance the sensitivity and anti-nonspecific binding of the resulting immunoassay electrode. The preliminary application of poly-OAP/CEAAb-AuNP/Au electrode for detection of CEA was also evaluated.     

Characterization and electrocatalytic properties of Prussian blue electrochemically deposited on nano-Au/PAMAM dendrimer-modified gold electrode

Gold electrode was modified with 3-mercaptopropionic acid (MPA) and further reacted with poly(amidoamine) (PAMAM) dendrimer (generation 4.0) then attached the nano-Au to obtain films on which Prussian blue (PB) was electrochemically deposited to afford much wider pH adaptive range, much better electrochemical stability and excellent electrochemical response. The microstructure and electrochemical behavior of Au/MPA/PAMAM/nano-Au/PB electrode were investigated by scanning electron microscopy (SEM) and cyclic voltammetry. The electrochemical response of the Au/MPA/PAMAM/nano-Au/PB-modified electrode for the electrocatalytic reduction of hydrogen peroxide was investigated, and it was found that the sensitivity as well as the corresponding detection limits were improved as compared to the voltammetric response of a Au/PB-modified electrode and Au/MPA/PAMAM/PB electrode. Based on this, a new electrochemical sensor for determination of hydrogen peroxide has been developed.     

Novel amperometric assay for drug-DNA interaction based on an inhibitory effect on an electrocatalytic activity of DNA-Cu(II) complex

A novel strategy of amperometric assay for drug-dsDNA interactions was developed based on an inhibitory effect of antimararial drug (quinacrine) on an electrocatalytic activity of DNA-Cu(II) complex. In this method, a DNA-Cu(II) complex immobilized DNA/polyallylamine(PAA) polyion complex membrane was used as a sensing element. The electrocatalytic activity of a DNA-Cu(II) complex for hydrogen peroxide reduction was reversibly inhibited by electron blocking effect of quinacrine-dsDNA interaction and this inhibitory effect was amplified by the hydrogen peroxide reduction. This phenomenon was utilized for development of a novel amperometric biosensor for DNA-binding drug. From the amperometric current-time curves, the response time of the sensor to 20 μM quinacrine was obtained about 20s, and the detection limit of the quinacrine was found to be 10 μM estimated to a signal-to-noise ratio of 3.0. Based on the change of steady-state catalytic current, the kinetic analysis of drug-dsDNA interaction can be done in a similar manner of enzyme inhibition, and the binding constant of the quinacrine with DNA can be calculated. This measurement method would be useful for screening of wide variety of DNA-binding drugs and highly toxic pollutants.     

P-chip and P-chip bienzyme electrodes based on recombinant forms of horseradish peroxidase immobilized on gold electrodes

Adsorption and bioelectrocatalytic activity of native horseradish peroxidase (HRP) and its recombinant forms on polycrystalline gold electrodes were studied. Recombinant forms of HRP were produced by a genetic engineering approach using an E. coli expression system. According to direct mass measurements with a quartz crystal microbalance, all the forms of HRP formed monolayer coverage of the enzyme on the gold surface. However, only gold electrodes modified with the recombinant HRP forms (non-glycosylated) exhibited high and stable current response to H₂O₂ due to its bioelectrocatalytic reduction based on direct electron transfer (ET) between gold and the active site of the enzyme. Introduction of a six-His tag either at the C-terminus or at the N-terminus of the enzyme molecule additionally increased the strength of the enzyme binding with the gold surface and the efficiency of direct ET. Immobilization of recombinant forms of HRP containing histidine functional groups on the surface of the gold electrode was used both for the development of a P-chip, a biosensor for hydrogen peroxide determination based on direct ET, and for the development of a bienzyme biosensor electrode for the determination of L-lysine based on co-immobilized recombinant forms of HRP and L-lysine--oxidase.     

Nanodiagnostic Method for Colorimetric Detection of Mycobacterium tuberculosis 16S rRNA

A nanodiagnostic method using nucleic acid sequence-based amplification (NASBA) and gold nanoparticle probes (AuNP probes) was developed for colorimetric detection of Mycobacterium tuberculosis. The primers targeting 16S rRNA were used for the amplification of mycobacterial RNA by the isothermal NASBA process. The amplicons were hybridized with specific gold nanoparticle probes. The RNA–DNA hybrids were colorimetrically detected by the accumulation of gold nanoparticles. Using this method, 10 CFU ml^?1 of M. tuberculosis was detected within less than 1 h. Results obtained from the clinical specimens showed 94.7% and 96% sensitivity and specificity, respectively. No interference was encountered in the amplification and detection of M. tuberculosis in the presence of non-target bacteria, confirming the specificity of the method.     

Amperometric biosensor for hydrogen peroxide based on horseradish peroxidase onto gold nanowires and TiO2 nanoparticles

An electrochemical biosensor for determination of hydrogen peroxide (H₂O₂) was fabricated, based on the electrostatic immobilization of horseradish peroxidase (HRP) with one-dimensional gold nanowires (Au NWs) and TiO₂ nanoparticles (nano-TiO₂) on a gold electrode. The nano-TiO₂ can give a biocompatible microenvironment and compact film, and the Au NWs can provide fast electron transferring rate and greatly add the amount of HRP molecules immobilized on the electrode surface. Au NWs were characterized by ultraviolet–visible spectra and transmission electron microscope. The electrode modification process was probed by cyclic voltammetry and electrochemical impedance spectroscopy. Chronoamperometry was used to study the electrochemical performance of the resulting biosensor. Under optimal conditions, the linear range for the determination of H₂O₂ was from 2.3 × 10⁻⁶ to 2.4 × 10⁻³ M with a detection limit of 7.0 × 10⁻⁷ M (S/N = 3). Moreover, the proposed biosensor showed superior stability and high sensitivity.     

Nanomolar detection of hydrogen peroxide at a new polynuclear cluster of tin pentacyanonitrosylferrate nanoparticle-modified carbon ceramic electrode

This work describes a new electrochemical sensor for hydrogen peroxide based on tin pentacyanonitrosylferrate (SnPCNF)-modified carbon ceramic electrode (CCE). The modified electrode was constructed by using a sol-gel technique involving two steps: construction of CCE containing metallic tin (Sn) powder and then electrochemical creation of SnPCNF film on the surface of CCE. The modified electrode was characterized by energy-dispersive X-ray, Fourier transform infrared, scanning electron microscopy, and cyclic voltammetry (CV) techniques. The charge transfer coefficient (α) and charge transfer rate constant (k_s) for the modifying film were calculated. The electrocatalytic activity of the modified electrode toward the reduction of hydrogen peroxide was studied by CV and chronoamperometry. A linear calibration curve was obtained over the hydrogen peroxide concentration range of 0.5 to 69.4 μM using a hydrodynamic amperometric technique. The limit of detection (for a signal-to-noise ratio of 3) and sensitivity were found to be 92 nM and 0.89 μA/μM, respectively. Furthermore, the diffusion coefficient of hydrogen peroxide (D) and catalytic rate constant (k_cat) were calculated.