Fabrication of a Nanobiocomposite Film Containing Heme Proteins and Carbon Nanotubes on a Choline Modified Glassy Carbon Electrode: Direct Electrochemistry and Electrochemical Catalysis

A nanocomposite electrochemical sensing film is assembled on choline (Ch) modified glassy carbon electrode (GCE), which contains multiwalled carbon nanotubes (MWNTs), Nafion cation exchanger, and myoglobin (Mb) or hemoglobin (Hb). The MWNTs provide a 3D porous and conductive network for the enzyme immobilization and Nafion acts as polymeric binder to give cast thin films. Both MWNTs and Nafion provide negative functionalities to bind to the positively charged redox proteins and to attach at the positively charged Ch modified layer, and drive the formation of homogeneous and stable nanocomposite film, the MWNT‐Nafion‐Mb. The nanocomposite film was characterized by field emission scanning electron microscope (FE‐SEM). The Mb in the nanocomposite film showed a pair of well‐defined and nearly reversible cyclic voltammetric peaks at about ?0.32 V vs. SCE at pH 7.0 solution for the heme Fe(III)/Fe(II) redox couple. The immobilized heme proteins can display the features of peroxidase in electrocatalytic reductions of oxygen, hydrogen peroxide, nitric oxide, trichloroacetic acid (TCA), and bromate.     

Electrodeposited nanogold decorated graphene modified carbon ionic liquid electrode for the electrochemical myoglobin biosensor

In this paper, a carbon ionic liquid electrode (CILE) was fabricated using ionic liquid 1-hexylpyridinium hexafluorophosphate as modifier, which was further in situ electrodeposited with graphene (GR) and gold nanoparticles step by step to get an Au/GR nanocomposite modified CILE. Myoglobin (Mb) was further immobilized on the Au/GR/CILE surface with Nafion film to get the modified electrode denoted as Nafion/Mb/Au/GR/CILE. Cyclic voltammetric experiments indicated that a pair of well-defined quasi-reversible redox peaks appeared in pH 3.0 phosphate buffer solution with the formal potential (E ^0′) located at ?0.197 V (vs. saturated calomel electrode), which was the typical characteristics of Mb heme Fe(III)/Fe(II) redox couples. Thus, the direct electron transfer rate between Mb and the modified electrode was promoted due to the high conductivity and increased surface area of Au/GR nanocomposite present on electrode surface. Based on the cyclic voltammetric data, the electrochemical parameters of Mb on the modified electrode were calculated. The Mb-modified electrode showed excellent electrocatalytic activities towards the reduction of trichloroacetic acid and H₂O₂ with wider linear range and lower detection limit. Using GR and Au nanoparticles modified CILE, a new third-generation electrochemical Mb biosensor was constructed with good stability and reproducibility.     

基于AuNPs/PDDA-GO纳米复合物的电化学免疫传感器的构建及对SirT1蛋白的检测

基于AuNPs/PDDA-GO纳米复合物制备了一种新型电化学免疫传感器, 并将其用于SirT1的检测. 首先, 在电极表面修饰复合材料AuNPs/PDDA-GO, 然后将目标蛋白SirT1固定到修饰了AuNPs/PDDA-GO的电极表面, 再通过特异性免疫反应结合一抗(Ab₁)和辣根过氧化酶标记的二抗分子(HRP-Ab₂), 最后用示差脉冲伏安法检测电流信号, 实现了对SirT1蛋白水平的测定. 在优化的实验条件下, SirT1蛋白的浓度在0.1~100 ng/mL范围内与响应电流呈良好线性关系, 检出限为0.029 ng/mL.     

Direct electrochemistry and electrochemical catalysis of myoglobin-TiO2 coated multiwalled carbon nanotubes modified electrode

TiO(2) nanoparticles were homogeneously coated on multiwalled carbon nanotubes (MWCNTs) by hydrothermal deposition, and this nanocomposite might be a promising material for myoglobin (Mb) immobilization in view of its high biocompatibility and large surface. The glassy carbon (GC) electrode modified with Mb-TiO(2)/MWCNTs films exhibited a pair of well-defined, stable and nearly reversible cycle voltammetric peaks. The formal potential of Mb in TiO(2)/MWCNTs film was linearly varied in the range of pH 3-10 with a slope of 48.65 mV/pH, indicating that the electron transfer was accompanied by single proton transportation. The electron transfer between Mb and electrode surface, k(s) of 3.08 s(-1), was greatly facilitated in the TiO(2)/MWCNTs film. The electrocatalytic reductions of hydrogen peroxide were also studied, and the apparent Michaelis-Menten constant is calculated to be 83.10 microM, which shows a large catalytic activity of Mb in the TiO(2)/MWCNTs film to H(2)O(2). The modified GC electrode shows good analytical performance for amperometric determination of hydrogen peroxide. The resultant Mb-TiO(2)/MWCNTs modified glassy carbon electrode exhibited fast amperometric response to hydrogen peroxide reduction, long term life and excellent stability. Finally the activity of the sensor for nitric oxide reduction was also investigated.     

Exploiting multi-function Metal-Organic Framework nanocomposite Ag@Zn-TSA as highly efficient immobilization matrixes for sensitive electrochemical biosensing

A novel multi-function Metal-Organic Framework composite Ag@Zn-TSA (zinc thiosalicylate, Zn(C₇H₄O₂S), Zn-TSA) was synthesized as highly efficient immobilization matrixes of myoglobin (Mb)/glucose oxidase (GOx) for electrochemical biosensing. The electrochemical biosensors based on Ag@Zn-TSA composite and ionic liquid (IL) modified carbon paste electrode (CPE) were fabricated successfully. Furthermore, the properties of the sensors were discussed by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and amperometric current-time curve, respectively. The results showed the proposed biosensors had wide linear response to hydrogen peroxide (H₂O₂) in the range of 0.3–20,000 μM, to nitrite (NO₂⁻) for 1.3 μM–1660 μM and 2262 μM–1,33,000 μM, to glucose for 2.0–1022 μM, with a low detection limit of 0.08 μM for H₂O₂, 0.5 μM for NO₂⁻, 0.8 μM for glucose. The values of the apparent heterogeneous electron transfer rate constant (k_s) for Mb and GOx were estimated as 2.05 s⁻¹ and 2.45 s⁻¹, respectively. Thus, Ag@Zn-TSA was a kind of ideal material as highly efficient immobilization matrixes for sensitive electrochemical biosensing. In addition, this work indicated that MOF nanocomposite had a great potential for constructing wide range of sensing interface.     

One step electrochemically deposited nanocomposite film of chitosan-carbon nanotubes-gold nanoparticles for carcinoembryonic antigen immunosensor application

A simple and controllable one-step electrodeposition method for the preparation of a chitosan-carbon nanotubes-gold nanoparticles (CS-CNTs-GNPs) nanocomposite film was used to fabricate an immunosensor for detection of carcinoembryonic antigen (CEA). The porous three-dimensional CS-CNTs-GNPs nanocomposite film, which offered a large specific surface area for immobilization of antibodies, exhibited improved conductivity, high stability and good biocompatibility. The morphology of the formed nanocomposite film was investigated by scanning electron microscopy (SEM), and the electrochemical behaviors of the immunosensor were characterized by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Under the optimal conditions, the proposed immunosensor could detect CEA in two linear ranges from 0.1 to 2.0 ng mL⁻¹ and from 2.0 to 200.0 ng mL⁻¹, with a detection limit of 0.04 ng mL⁻¹. The immunosensor based on CS-CNTs-GNPs nanocomposite film as the antibody immobilization matrix could exhibit good sensitivity, stability, and reproducibility for the determination of CEA.     

Electrochemical biosensors based on Ti3C2Tx MXene: future perspectives for on-site analysis

MXenes are recently developed two-dimensional layered materials composed of early transition metal carbides and/or nitrides that provide unique characteristics for biosensor applications. This review presents the recent progress made on the usage and applications of MXenes in the field of electrochemical biosensors, including microfluidic biosensors and wearable microfluidic biosensors, and highlights the challenges with possible solutions and future needs. The multilayered configuration and high conductivity make these materials as an immobilization matrix for the biomolecule immobilization with activity retention and to be explored in the fabrication of electrochemical sensors, respectively. First, how the MXene nanocomposite as an electrode modifier affects the sensing performance of the electrochemical biosensors based on enzymes, aptamer/DNA, and immunoassays is well described. Second, recent developments in MXene nanocomposites as wearable biosensing platforms for the biomolecule detection are highlighted. This review pointed out the future concerns and directions for the use of MXene nanocomposites to fabricate advanced electrochemical biosensors with high sensitivity and selectivity. Specifically, possibilities for developing microfluidic electrochemical sensors and wearable electrochemical microfluidic sensors with integrated biomolecule detection are emphasized.     

Amperometric hydrogen peroxide biosensor based on the immobilization of heme proteins on gold nanoparticles-bacteria cellulose nanofibers nanocomposite

A novel matrix, gold nanoparticles-bacterial cellulose nanofibers (Au-BC) nanocomposite was developed for enzyme immobilization and biosensor fabrication due to its unique properties such as satisfying biocompatibility, good conductivity and extensive surface area, which were inherited from both gold nanoparticles (AuNPs) and bacterial cellulose nanofibers (BC). Heme proteins such as horseradish peroxidase (HRP), hemoglobin (Hb) and myoglobin (Mb) were successfully immobilized on the surface of Au-BC nanocomposite modified glassy carbon electrode (GCE). The immobilized heme proteins showed electrocatalytic activities to the reduction of H₂O₂ in the presence of the mediator hydroquinone (HQ), which might be due to the fact that heme proteins retained the near-native secondary structures in the Au-BC nanocomposite which was proved by UV-vis and IR spectra. The response of the developed biosensor to H₂O₂ was related to the amount of AuNPs in Au-BC nanocomposite, indicating that the AuNPs in BC network played an important role in the biosensor performance. Under the optimum conditions, the biosensor based on HRP exhibited a fast amperometric response (within 1 s) to H₂O₂, a good linear response over a wide range of concentration from 0.3 μM to 1.00 mM, and a low detection limit of 0.1 μM based on S/N = 3. The high performance of the biosensor made Au-BC nanocomposite superior to other materials as immobilization matrix.     

Electrochemical detection of DNA damage induced by in situ generated bisphenol A radicals through electro-oxidation

A glassy carbon electrode was modified with dsDNA and a nanocomposite composed of multi-walled carbon nanotubes and chitosan (MWNT-chit). The electrode was applied to the electrochemical detection of DNA damage as induced by in situ generated bisphenol A (BPA) radicals through electro-oxidation. The modified electrode was characterized by cyclic voltammetry and electrochemical impedance spectroscopy. The results indicate that MWNT-chit nanocomposite represents a viable platform for the immobilization of DNA that effectively promotes electron transfer between DNA and the electrode. The mode of interaction between DNA and BPA was investigated by differential pulse voltammetry and UV-vis spectrophotometry, indicating that the dominant interaction is intercalation. In order to explore the mechanism of damage caused by BPA radicals, the electro-oxidation of BPA at the modified glass electrode was investigated. Based on the signal for guanine without any other external indicator, DNA damage was investigated through the electro-oxidation of BPA.     

Direct Electrochemistry and Electrocatalysis of Myoglobin Immobilized on Gold Nanoparticles/Carbon Nanotubes Nanohybrid Film

A novel nanohybrid material, constructed by gold nanoparticles (GNPs) and multiwalled carbon nanotubes (MWNTs), was designed for immobilization and biosensing of myoglobin (Mb). Morphology of the nanohybrid film was characterized by SEM. UV‐vis spectroscopy demonstrated that Mb on the composite film could retain its native structure. Direct electrochemistry of Mb immobilized on the GNPs/MWNTs film was investigated. The immobilized Mb showed a couple of quasireversible and well‐defined cyclic voltammetry peaks with a formal potential of about ?0.35 V (vs. Ag/AgCl) in pH 6.0 phosphate buffer solution (PBS) solution. Furthermore, the modified electrode also displayed good sensitivity, wide linear range and long‐term stability to the detection of hydrogen peroxide. The experiment results demonstrated that the hybrid matrix provided a biocompatible microenvironment for protein and supplied a necessary pathway for its direct electron transfer.     

Ultrasensitive Electrochemical Immunosensor for Detection of Tumor Necrosis Factor‐α Based on Functionalized MWCNT‐Gold Nanoparticle/Ionic Liquid Nanocomposite

A novel and highly sensitive electrochemical immunosensor was developed for the detection of protein biomarker tumor necrosis factor‐alpha (TNF‐α) based on immobilization of TNF‐α‐antibody (anti‐TNF‐α) onto robust nanocomposite containing gold nanoparticles (AuNP), multiwalled carbon nanotubes (MWCNTs) and ionic liquid (1‐buthyl‐3‐methylimidazolium bis (trifluoromethyl sulfonyl)imide). Functionalized MWCNT‐gold nanoparticle was produced by one‐step synthesis based on the direct redox reaction. The electrochemical properties of nanocomposite were characterized by electrochemical impedance spectroscopy and cyclic voltammetry. The anti‐TNF‐α was immobilized or entrapped in the nanocomposite and used in a sandwich type complex immunoassay with anti‐TNF‐α labeled with horseradish peroxidase as secondary antibody. Under optimum conditions, the immunosensor could detect TNF‐α in a linear range from 6.0 to 100 pg mL^?1 with a low detection limit of 2.0 pg mL^?1. The simple fabrication method, high sensitivity, good reproducibility, stability, as well as acceptable accuracy for TNF‐α detection in human serum samples are the main advantages of this immunosensor, which might have broad applications in protein diagnostics and bioassay.     

Electrochemical performance and electrocatalytic behavior of myoglobin on graphene tube-modified electrode

A new protein biosensing strategy based on a graphene tube (GT)-modified electrode was developed in this article. GT as a fixed material and signal amplifier was loaded on the carbon ionic liquid electrode (CILE) surface, which could provide a suitable environment for myoglobin (Mb) immobilization and promote electron transfer between Mb and the electrode. Fourier-transform infrared spectroscopy (FT-IR) of Mb before and after mixing with GT was checked and showed that the native structure of Mb was maintained. The direct electrochemistry of Mb was investigated with a pair of obtained well-defined and quasireversible redox peaks. The modified electrode (Nafion/Mb/GT/CILE) also showed excellent electrocatalytic activity for the reduction of trichloroacetic acid (TCA) and NaNO₂, which could be utilized to determine the concentrations of TCA and NaNO₂ with wide detection range and low detection limit. The biosensor was further applied to the detection of a real sample and obtained satisfactory experimental results.     

A Label-Free Amperometric Immunoassay for Thrombomodulin Using Graphene/Silver-Silver Oxide Nanoparticles as a Immobilization Matrix

A simple, sensitive, and label-free electrochemical immunosensor has been developed for the measurement of serum thrombomodulin (TM), an endothelial glycoprotein which is associated with the progression and metastasis of tumors. At first, the graphene nanosheets, which were dispersed in Nafion solution, were used to coat the bare gold electrode. Then, silver-silver oxide nanoparticles (Ag-Ag₂O NPs) were immobilized on the graphene-modified electrode by a one-step electrochemical deposition method. Lastly, a thrombomodulin antibody (anti-TM) was attached via amido-Ag affinity. This strategy combines graphene/Ag-Ag₂O NPs hybrid materials as an immobilization matrix and Ag-Ag₂O NPs also as an electrochemical signal indication reagent. The main advantage of this strategy has two important aspects. One is the high stability and unique electronic properties of the graphene nanostructure. The other is the use of Ag-Ag₂O NPs as the immobilization matrix and redox probes, thus avoiding the laborious labeling protein operation. Using this strategy, the concentration of TM in the range of 0.1 to 20 ng/mL was detected, with a detection limit of 31.5 pg/mL (at 3σ). The proposed methodology demonstrates that the nanocomposite film composed of graphene and Ag-Ag₂O NPs is a potential for biosensor applications.     

Acetylcholinesterase Immobilization on Polyacrylamide/Functionalized Multi-walled Carbon Nanotube Nanocomposite Nanofibrous Membrane

In this work, polyacrylamide/multi-walled carbon nanotubes (MWCNT) solution is electrospun to nanocomposite nanofibrous membranes for acetylcholinesterase enzyme immobilization. A new method for enzyme immobilization is proposed, and the results of analysis show successful covalent bonding of enzymes on electrospun membrane surface besides their non-covalent entrapment. Fourier transform infrared spectroscopy, mechanical and thermal investigations of nanofibrous membrane approve successful cross-linking and enzyme immobilization. The enzyme relative activity and kinetic on both pure and nanocomposite membranes is investigated, and the results show proper performance of designed membrane to even improve the enzyme activity followed by immobilization compared to free enzyme. Scanning electron microscopy images show nanofibrous web of 3D structure with a low shrinkage and hydrogel structure followed by enzyme immobilization and cross-linking. Moreover, the important role of functionalized carbon nanotubes on final nanofibrous membrane functionality as a media for enzyme immobilization is investigated. The results show that MWCNT could act effectively for enzyme immobilization improvement via both physical (enhanced fibers’ morphology and conductivity) and chemical (enzyme entrapment) methods.

Renewable nanocomposite layer-by-layer assembled catalytic interfaces for biosensing applications

A novel, easily renewable nanocomposite interface based on layer-by-layer (LbL) assembled cationic/anionic layers of carbon nanotubes customized with biopolymers is reported. A simple approach is proposed to fabricate a nanoscale structure composed of alternating layers of oxidized multiwalled carbon nanotubes upon which is immobilized either the cationic enzyme organophosphorus hydrolase (OPH; MWNT-OPH) or the anionic DNA (MWNT-DNA). The presence of carbon nanotubes with large surface area, high aspect ratio and excellent conductivity provides reliable immobilization of enzyme at the interface and promotes better electron transfer rates. The oxidized MWNTs were characterized by thermogravimetric analysis and Raman spectroscopy. Fourier transform infrared spectroscopy showed the surface functionalization of the MWNTs and successful immobilization of OPH on the MWNTs. Scanning electron microscopy images revealed that MWNTs were shortened during sonication and that LbL of the MWNT/biopolymer conjugates resulted in a continuous surface with a layered structure. The catalytic activity of the biopolymer layers was characterized using absorption spectroscopy and electrochemical analysis. Experimental results show that this approach yields an easily fabricated catalytic multilayer with well-defined structures and properties for biosensing applications whose interface can be reactivated via a simple procedure. In addition, this approach results in a biosensor with excellent sensitivity, a reliable calibration profile, and stable electrochemical response.     

Dual Amplified Electrochemical Immunosensor for Hepatitis B Virus Surface Antigen Detection Using Hemin/G‐Quadruplex Immobilized onto Fe3O4‐AuNPs or (Hemin‐Amino‐rGO‐Au) Nanohybrid

A sensitive electrochemical immunosensor for Hepatitis B virus surface antigen (HBsAg) detection was fabricated based on hemin/G‐quadruplex interlaced onto Fe₃O₄‐AuNPs or hemin ‐amino‐reduced graphene oxide nanocomposite (H‐amino‐rGO‐Au). G‐quadruplex DNAzyme, which is composed of hemin and guanine‐rich nucleic acid, is an effective signal amplified tool for its outstanding peroxidase activity and Fe₃O₄‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites with quasi‐enzyme activity provide appropriate support for the immobilization of hemin/G‐quadruplex. The target protein was sandwiched between the primary antibody immobilized on the GO and secondary antibody immobilized on the Fe₃O₄‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites and glutaraldehyde was used as linking agent for the immobilization of primary antibody on the surface of GO. Both Fe₃O₄‐AuNPs and H‐amino‐rGO‐Au nanocomposite and also hemin/G‐quadruplex can cooperate the electrocatalytic reduction of H₂O₂ in the presence of methylene blue as mediator. The proposed immunosensor has a wide linear dynamic range of 0.1 pg/ml to 300 pg/ml with a detection limit of 60 fg/ml when Fe₃O₄‐AuNPs was used for immobilization of hemin/G‐quadruplex, while the dynamic range and DL were 0. 1–1000 pg/mL and 10 fg/mL, respectively in the presence of H‐amino‐rGO‐ Au nanocomposite as platform for immobilizing of hemin/G‐quadruplex. The proposed immunosensor was also used for analysis of HBsAg in spiked human serum samples with satisfactory results.     

Myoglobin/sol-gel film modified electrode: direct electrochemistry and electrochemical catalysis

Direct electrochemical and electrocatalytic behavior of myoglobin (Mb) immobilized on carbon paste electrode (CPE) by a silica sol-gel film derived from tetraethyl orthosilicate was investigated for the first time. Mb/sol-gel film modified electrodes show a pair of well-defined and nearly reversible cyclic voltammetric peaks for the Mb Fe(III)/Fe(II) redox couple at about -0.298 V (vs Ag/AgCl) in a pH 7.0 phosphate buffer solution. The formal potential of the Mb heme Fe(III)/Fe(II) couple shifted linearly with pH with a slope of 52.4 mV/pH, denoting that an electron transfer accompanies single-proton transportation. An FTIR and UV-vis spectroscopy study confirms that the secondary structure of Mb immobilized on an electrode by a sol-gel film still maintains the original arrangement. The immobilized Mb displays the features of a peroxidase and acts in an electrocatalytic manner in the reduction of oxygen, trichloroacetic acid (TCA), and nitrite. In comparison to other electrodes, the chemically modified electrodes used in this study for direct electrochemistry and electrocatalysis of Mb are easy to fabricate and fairly inexpensive. Consequently, the Mb/sol-gel film modified electrode provides a convenient way to perform electrochemical research on this kind of protein. It also has potential use in the fabrication of bioreactors and third-generation biosensors.     

Nonenzymatic sensor for hydrogen peroxide based on the electrodeposition of silver nanoparticles on poly(ionic liquid)-stabilized graphene sheets

We have developed a nonenzymatic sensor for hydrogen peroxide (HP) that is based on a new kind of nanocomposite consisting of silver nanoparticles (AgNPs) electrodeposited on a basic film of a poly(ionic liquid) containing graphene. The nanocomposite was characterized by scanning electron microscopy, energy dispersive X-ray studies, cyclic voltammetry, and chronoamperometry. The AgNPs on the basic composite film provide the electrode with enhanced sensitivity in that the signal obtained for HP is 10-fold improved in the best case. The sensor exhibits good linear response in the 0.1 μM to 2.2 mM HP concentration range, and the detection limit is 0.05 μM (at S/N?=?3).      

Indicator-free electrochemical genosensing originated from the self-signal of poly-xanthurenic acid enhanced by Fe3O4/reduced graphene oxide

In this paper, an indicator-free electrochemical genosensing platform based on the self-signal changes of poly-xanthurenic acid (PXa) enhanced by Fe₃O₄/reduced graphene oxide (Fe₃O₄/RGO) was constructed. The resulting nanocomposite (PXa-Fe₃O₄/RGO) was characterized by transmission electron microscopy, cyclic voltammetry, and electrochemical impedance spectroscopy (EIS). The π–π* stacking and hydrogen bonding between the conjugated Fe₃O₄/GO and aromatic ring of xanthurenic acid monomer promoted the electropolymerization efficiency accompanied with an increased electrochemical response of PXa. The immobilization of the specific probe DNA was successfully realized via the noncovalent method due to the π–π* interaction between the conjugated nanostructure of PXa-Fe₃O₄/RGO and DNA bases. The hybridization between the probe DNA and target DNA induced the resulted double-stranded (ds)DNA to be released from the conjugated nanocomposite, accompanied with the self-signal regeneration of nanocomposite (“signal-on”). The self-signal changes could serve as a powerful tool for indicator-free and freely switchable detection of different target genes, and the synergistic effect of the integrated graphene-based nanocomposite effectively improved the sensitivity for the target DNA detection via EIS.     

Microbiosensors based on DNA modified single-walled carbon nanotube and Pt black nanocomposites

Glucose and ATP biosensors have important applications in diagnostics and research. Biosensors based on conventional materials suffer from low sensitivity and low spatial resolution. Our previous work has shown that combining single-walled carbon nanotubes (SWCNTs) with Pt nanoparticles can significantly enhance the performance of electrochemical biosensors. The immobilization of SWCNTs on biosensors remains challenging due to the aqueous insolubility originating from van der Waals forces. In this study, we used single-stranded DNA (ssDNA) to modify SWCNTs to increase solubility in water. This allowed us to explore new schemes of combining ssDNA-SWCNT and Pt black in aqueous media systems. The result is a nanocomposite with enhanced biosensor performance. The surface morphology, electroactive surface area, and electrocatalytic performance of different fabrication protocols were studied and compared. The ssDNA-SWCNT/Pt black nanocomposite constructed by a layered scheme proved most effective in terms of biosensor activity. The key feature of this protocol is the exploitation of ssDNA-SWCNTs as molecular templates for Pt black electrodeposition. The glucose and ATP microbiosensors fabricated on this platform exhibited high sensitivity (817.3 nA/mM and 45.6 nA/mM, respectively), wide linear range (up to 7 mM and 510 μM), low limit of detection (1 μM and 2 μM) and desirable selectivity. This work is significant to biosensor development because this is the first demonstration of ssDNA-SWCNT/Pt black nanocomposite as a platform for constructing both single-enzyme and multi-enzyme biosensors for physiological applications.