Biosensor Response Research Articles

Electrochemical study of the effect of radiofrequency on glutamate oxidase activity using a glutamate oxidase-based biosensor

A biosensor based on glutamate oxidase (GluOx) was developed to measure glutamate concentration. The main function of this type of biosensor is related to the structure and catalytic activity of GluOx. Since radiofrequency, as the widest spectrum of electromagnetic fields, can affect the catalytic activity and structure of GluOx, in this study, the effect of these fields on the analytical parameters of the fabricated biosensor was investigated. To build the biosensor a sol-gel solution of chitosan and native GluOx were prepared and then immobilized on the surface of the platinum electrode. Similarly, to investigate the effect of radiofrequency fields on the analytical parameters of the biosensor, instead of the native GluOx, irradiated GluOx was used to build the biosensor. To evaluate the biosensor responses, cyclic voltammetry experiments were performed and voltammograms were considered as biosensor responses. To determine the analytical parameters including detection limit, linear range, and saturation region of the responses, calibration curves were drawn for each of the biosensors. Also the long-term stability and selectivity of the fabricated biosensor were evaluated. Thereafter, the optimum pH and temperature for each of these two biosensors were examined. The results showed that radiofrequency waves harmed the detection and response of biosensors in the saturation region, while they had little effect on the linear region. Such results could be due to the effect of radiofrequency waves on the structure and function of glutamate oxidase. In general, the results indicate that when a glutamate oxidase-based biosensor is used to measure glutamate in radiofrequency fields, corrective coefficients for this type of biosensor should be considered to accurately measure glutamate concentration.

Taguchi method: artificial neural network approach for the optimization of high-efficiency microfluidic biosensor for COVID-19.

COVID-19 is a pandemic disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This virus is mainly spread by droplets, respiratory secretions, and direct contact. Caused by the huge spread of the COVID-19 epidemic, research is focused on the study of biosensors as it presents a rapid solution for reducing incidents and fatality rates. In this paper, a microchip flow confinement method for the rapid transport of small sample volumes to sensor surfaces is optimized in terms of the confinement coefficient β, the position of the confinement flow X, and its inclination α relative to the main channel. A numerical simulation based on two-dimensional Navier-Stokes equations has been used. Taguchi's L9(33) orthogonal array was adopted to design the numerical assays taking into account the confining flow parameters (α, β, and X) on the response time of microfluidic biosensors. Analyzing the signal-to-noise ratio allowed us to determine the most effective combinations of control parameters for reducing the response time. The contribution of the control factors to the detection time was determined via analysis of variance (ANOVA). Numerical predictive models using multiple linear regression (MLR) and an artificial neural network (ANN) were developed to accurately predict microfluidic biosensor response time. This study concludes that the best combination of control factors is that corresponds to , and X = 40µm. Analysis of variance (ANOVA) shows that the position of the confinement channel (62% contribution) is the factor most responsible for the reduction in response time. Based on the correlation coefficient (R 2), and value adjustment factor (VAF), the ANN model performed better than the MLR model in terms of prediction accuracy.

Tear-based MMP-9 detection: A rapid antigen test for ocular inflammatory disorders using vanadium disulfide nanowires assisted chemi-resistive biosensor

A sensitive, non-invasive, and biomarker detection in tear fluids for inflammation in potentially blinding eye diseases could be of great significance as a rapid diagnostic tool for quick clinical decisions. In this work, we propose a tear-based MMP-9 antigen testing platform using hydrothermally synthesized vanadium disulfide nanowires. Also, various factors contributing to baseline drifts of the chemiresistive sensor including nanowire coverage on the interdigitated microelectrode of the sensor, sensor response duration, and effect of MMP-9 protein in different matrix solutions were identified. The drifts on the sensor baseline due to nanowire coverage on the sensor were corrected using substrate thermal treatment providing a more uniform distribution of nanowires on the electrode which brought the baseline drift to 18% (coefficient of variations, CV = 18%). This biosensor exhibited sub-femto level limits of detection (LODs) of 0.1344 fg/mL (0.4933 fmoL/l) and 0.2746 fg/mL (1.008 fmoL/l) in 10 mM phosphate buffer saline (PBS) and artificial tear solution, respectively. For a practical tear MMP-9 detection, the proposed biosensor response was validated with multiplex ELISA using tear samples from five healthy controls which showed excellent precision. This label-free and non-invasive platform can serve as an efficient diagnostic tool for the early detection and monitoring of various ocular inflammatory diseases.

Electrochemical Sensing and Biosensor Based on Functional Polymers

Functional polymers has great importance in the field of electrochemical sensing and biosensors due to adaptable chemical, electrical, and structural features, functional polymers are a significant class of materials that have been extensively used to create electrochemical biosensors. Additionally, conducting polymers can be made nanostructured, functional group-grafted chemically, or combined with other functional materials, like nanoparticles, to significantly enhance the sensitivity, selectivity, stability, and reproducibility of the biosensor's response to a variety of bioanalytes. Since these biosensors offer benefits such being affordable and having a low detection limit, they are anticipated to play an increasingly important role in providing diagnostic information and monitoring therapy. Because of this, this article begins with a description of the electroanalytical techniques (amperometry, potentiometry, conductometry, impedometry, voltammetry) used in electrochemical biosensors, and then moves on to a review of recent developments in the use of conducting polymers in the identification of bioanalytes that led to the development of enzyme-based biosensors, immunosensors, DNA biosensors, and whole-cell biosensors.

Microbial Biosensor for Characterization of a Microorganism: A Review focusing on the Biochemical Activity of Microbial Cells

Express assessment of the biochemical activity of microorganisms is important in both applied and fundamental research. A laboratory model of a microbial electrochemical sensor formed on the basis of the culture of interest is a device that provides rapidly information about the culture and is cost effective, simple to fabricate and easy to use. This paper describes the application of laboratory models of microbial sensors in which the Clark-type oxygen electrode was used as a transducer. The formation of the models of the reactor microbial sensor (RMS) and the membrane microbial sensor (MMS) and the formation of the response of biosensors are compared. RMS and MMS are based on intact or immobilized microbial cells, respectively. For MMS, the response of biosensor is caused both by the process of transport of substrate into microbial cells and by the process of the initial metabolism of substrate; and only initial substrate metabolism triggers the RMS response. The details of the application of biosensors for the study of allosteric enzymes and inhibition by substrate are discussed. For inducible enzymes, special attention is paid to the induction of microbial cells. This article addresses current problems related to implementation of the biosensor approach and discusses the ways how to overcome these problems.

Microfluidic Electrochemical Glucose Biosensor with In Situ Enzyme Immobilization

The development and characterization of a microfluidic electrochemical glucose biosensor are presented herein. The transducer part is based on thin-film metal electrodes on a glass substrate. The biological recognition element of the biosensor is the pyrroloquinoline quinone-glucose dehydrogenase (PQQ-GdhB) enzyme, selectively in situ immobilized via microcontact printing of a mixed self-assembling monolayer (SAM) on a gold working electrode, while the microfluidic part of the device comprises microchannel and microfluidic connections formed in a polydimethylsiloxane (PDMS) elastomer. The electrode properties throughout all steps of biosensor construction and the biosensor response to glucose concentration and analyte flow rate were characterized by cyclic voltammetry and chronoamperometry. A measurement range of up to 10 mM in glucose concentration with a linear range up to 200 μM was determined. A detection limit of 30 µM in glucose concentration was obtained. Respective biosensor sensitivities of 0.79 nA/µM/mm2 and 0.61 nA/µM/mm2 were estimated with and without a flow at 20 µL/min. The developed approach of in situ enzyme immobilization can find a wide number of applications in the development of microfluidic biosensors, offering a path towards continuous and time-independent detection.

Mass transport limitations for electrochemical sensing in low-flux excretory fluids

Despite non-invasive instant monitoring of sweat metabolites is becoming a general trend in early diagnostics and screening, the reliability and accuracy of the on-skin electrochemical biosensors in real-life scenarios still remain questionable. As a rule, mass transport effects in scantily excreted liquids are ignored, when considering the design of such wearable setups. Here we provide a comprehensive investigation of the disruption factors for commonly used Prussian Blue based (bio)sensors under different hydrodynamic conditions (2 × 10−5 – 5 × 100 mm s−1 electrolyte velocity). A huge effect of flow on the (bio)sensors response has been revealed and explained with transport limitations for both analyte influx and reaction product outflux. It suggests no need for improving the sensor sensitivity, while minimizing analyte consumption and enhancing product withdrawal. Some strategies concerning measurement schemes and sensor design ensuring reliable sweat analysis have been discussed and illustrated for lactate and glucose on-skin monitoring.

High-performance amperometric biosensor for flow injection analysis consisting of a replaceable lactate oxidase-based mini-reactor and a silver amalgam screen-printed electrode

L-lactic acid (LA) is an essential compound in the winemaking process; furthermore, it is considered a significant biomarker for the diagnosis of a wide range of diseases, including cancer. Therefore, a cost-effective and reliable biosensing system for FIA is developed here for rapid and selective determination of LA. The biosensor architecture includes an easily replaceable lactate oxidase (LOx)-based mini-reactor connected in front of a silver amalgam screen-printed electrode acting as a transducer. The mini-reactor represents a tube filled with mesoporous silica powder (SBA−15) coated with covalently immobilized LOx. The principle of LA determination is based on the amperometric monitoring of oxygen consumption by its four-electron reduction at −900 mV vs. Ag pseudo-reference electrode, which avoids the interferences from commonly interfering compounds. The application of a strategy of a spatially separated biorecognition and detection part allowed for the immobilization of a notably large amount of enzyme (one mini-reactor contains c.a. 270 μg of the LOx), which, in turn, ensured excellent operational (93.8% of the initial signal after 350 measurements) and storage (96.9% of the initial biosensor response after 7 months) stability. The newly developed high-performance biosensor was successfully tested for the quantification of LA in saliva, wine, and dairy products.

Comparative Surface Interaction Study to Detect Brucella melitensis 16M Using Biosensor Transducer Modifications with 4-MBA, ZnONPs/AuNPs, ZnONPs/AuNPs/4-MBA

Brucellosis is a zoonotic disease endemic in developing countries and caused by gram-negative bacteria of genus Brucella infecting both livestock and humans. Brucella melitensis and Brucella abortus are important species representing largest biotypes world-wide. Therefore, modified detection strategies and advancement in potential analytical tools are required to monitor its rapid prevalence. In this study, we aim to modify gold-transducer of Surface Plasmon Resonance biosensor with combination of metal oxide nanomaterials and chemical probe to detect recombinant outer membrane ‘rOmp28’ protein antigen of Brucella melitensis 16M in concentration-dependent surface interactions. We synthesized Zinc (ZnONPs) and Gold (AuNPs) nanoparticles using standard ‘Hydrothermal and Turkevich Methods’ and their crystalline structure, chemical property and morphology were analysed using UV-Visible Spectrophotometry, FT-IR, Powder-XRD, SEM-EDX and TEM-SAED. For immobilizing specific rOmp28 derived IgG-pAbs on modified Au-transducer, rOmp28 protein was expressed and purified using Ni-NTA gel affinity chromatography for producing pAbs in BALB/c mice. Three modifications of Au-transducer with 4-MBA, ZnONPs/AuNPs and ZnONPs/AuNPs/4-MBA were subjected for immobilization and SPR biosensing was performed with rOmp28 Ag at detection range of 0.1 μg mL-1 to 0.01 fg mL-1. Limit of detection observed with ZnONPs/AuNPs/4-MBA combination was 0.1 fg mL-1 by relative increase in SPR response angle at 0.1 μg mL-1 in the order 83.7° < 98.9° < 179.2° for 4-MBA < ZnONPs/AuNPs < ZnONPs/AuNPs/4-MBA respectively. In conclusion, metal oxide nanomaterials in combination with biosensor are suitable in sensitive and specific interaction of antigen displaying lowest LODs and enhanced biosensor response for on-field real-time Brucella detection in both clinical and non-clinical disease scenario.

ALP-Based Biosensors Employing Electrodes Modified with Carbon Nanomaterials for Pesticides Detection.

Due to the growing presence of pesticides in the environment and in food, the concern of their impact on human health is increasing. Therefore, the development of fast and reliable detection methods is needed. Enzymatic inhibition-based biosensors represent a good alternative for replacing the more complicated and time-consuming traditional methods (chromatography, spectrophotometry, etc.). This paper describes the development of an electrochemical biosensor exploiting alkaline phosphatase as the biological recognition element and a chemically modified glassy carbon electrode as the transducer. The biosensor was prepared modifying the GCE surface by a mixture of Multi-Walled-Carbon-Nanotubes (MWCNTs) and Electrochemically-Reduced-Graphene-Oxide (ERGO) followed by the immobilization of the enzyme by cross-linking with bovine serum albumin and glutaraldehyde. The inhibition of the biosensor response caused by pesticides was established using 2-phospho-L-ascorbic acid as the enzymatic substrate, whose dephosphorylation reaction produces ascorbic acid (AA). The MWCNTs/ERGO mixture shows a synergic effect in terms of increased sensitivity and decreased overpotential for AA oxidation. The response of the biosensor to the herbicide 2,4-dichloro-phenoxy-acetic-acid was evaluated and resulted in the concentration range 0.04-24 nM, with a limit of the detection of 16 pM. The determination of other pesticides was also achieved. The re-usability of the electrode was demonstrated by performing a washing procedure.

Influence of BSA Protein on Electrochemical Response of Genosensors

The bovine serum albumin (BSA) protein is employed in genosensors as nonspecific binding blocker. In this work, the influence of BSA on the electrochemical response of a DNA-based biosensor is explored by electrochemical impedance spectroscopy (EIS) and double pulse voltammetry technique. Each buffer component of the hybridization buffer is studied on bare gold electrodes and on single-stranded DNA (ssDNA)-functionalized electrodes. The BSA is found to have the higher influence on the sensor sensitivity. The superficial modification induced by BSA greatly affects the electrochemical signal due to steric hindrance, which decreases the device sensitivity. Thus, we propose a novel treatment method of the BSA by using the proteinase K. The proteinase K reduces the steric hindrance, improving the charge transfer at the electrode/solution interface and enhancing the genosensor sensitivity. The proposed method is based on a rapid BSA treatment. The BSA solution containing the proteinase K is kept for 1 h at 37 ° C to obtain BSA fragmentation and then heated up to 95 ° C for 2 min to inactivate the enzyme and denature the fragments. The method was tested successfully on a genosensor for the detection of the Campylobacter Jejuni. Double pulse current measurements clearly discriminate the presence of complementary DNA (cDNA) fragments with any other noncomplementary DNA (NcDNA) fragments.

Paper biosensors utilize silver nanoparticles for onsite pesticide residue detection

The improper use of pesticides and excessive doses in the long term contribute to climate change and even threaten human health, organisms, and the balance of the ecosystem. A pesticide detection device is needed to monitor its levels to minimize risks to human health and the environment. A paper biosensor was developed in this study to detect organophosphates by utilizing the enzyme acetylcholine esterase (AChE) and silver nanoparticles (AgNP). The Whatman filter paper was used as a visual OP detection zone. AgNP, as an indicator, is adsorbed and gives a brownish-yellow color to the paper, while AChE is immobilized into the film and layered on the paper. The addition of acetylthiocholine chloride as a substrate to the film released thiocholine products which could replace the capping AgNPs causing the AgNPs to aggregate and the paper color to pale. The presence of OP in the sample will inhibit AChE activity so that paper fading is reduced. The biosensor response is quantized as an RGB value, which is determined using an application on a smartphone. The resulting biosensor has excellent performance with a linear range of 0.05-2.00 mg/L, a detection limit of 0.04 mg/L, and a CV of 0.48%. Biosensor measurements on vegetable samples showed conformity with the GCMS results as the standard method. Therefore, this biosensor is suitable for the on-site detection of pesticides offering easy, fast, and inexpensive analysis.

Developing an improved optical biosensing system based on gold nanoparticles acting as interferometric enhancers in Lactoferrin detection.

We report for the first time the whole development of a biosensing system based on the Interferometric Optical Detection Method (IODM) enriched with gold nanoparticles (AuNPs), acting as interferometric enhancers for improving the performance of immunoassays. For this purpose, the Lactoferrin sandwich immunoassay model was employed. We describe in detail the entire value chain from the AuNPs production, its functionalization, and characterization with anti-Lactoferrin (anti-LF), the biosensing response of these conjugates as well as their corresponding calculation of the kinetic constants, performance comparison of the readout interferometric signals versus Scanning Electron Microscopy (SEM) and the percentage of the sensing surface covered. Finally, a Lactoferrin sandwich immunoassay was carried out and correlated with Enzyme-Linked ImmunoSorbent Assay (ELISA), and the Limit of Detection and sensitivity figures were obtained. As a result, we demonstrate how the AuNPs act as interferometric amplifiers of the IODM for improving the biosensing response, opening the possibility of being applied in multiple biological detection applications.

Efficient and rapid linker optimization with heterodimeric coiled coils improves the response of fluorescent biosensors comprising antibodies and protein M.

We developed a coiled Q-probe (CQ-probe), a fluorescent probe containing a coiled-coil peptide pair E4/K4, to convert antibodies into biosensors for homogeneous immunoassays. This probe consists of an antibody-binding protein, protein M (PM) with the E4 peptide and the K4 peptide with a fluorescent dye. Compared to PM Q-probes, which are generated by modifying the C-terminus of PM with a fluorescent dye, CQ-probe variants with various linkers are easy to prepare and therefore enable the establishment of biosensors with a significant fluorescence response by localizing the fluorescent dye at the optimal position for quenching and antigen-dependent release. The fluorescence changes of biosensors converted from anti-BGP, anti-cortisol, and anti-testosterone antibodies using the rhodamine 6G (or TAMRA)-labeled CQ-probe upon antigen addition were 13 (or 2.6), 9.7 (or 1.5), and 2.1 (or 1.2) times larger than that of the biosensors converted using the PM Q-probe. Furthermore, the CQ-probe converted anti-digoxin IgG into a functional biosensor, whereas the PM Q-probe/antibody complex showed an insufficient response. This technology exhibits a promising capacity to convert antibodies into high-response biosensors, which are expected to be applied in a wide range of fields, including clinical diagnosis, environmental surveys, food analysis, and biological research.

Kinetics of metal detection by luminescence-based whole-cell biosensors: connecting biosensor response to metal bioavailability, speciation and cell metabolism

A mechanistic theory is developed to formulate the bioluminescence signal produced by metal-responsive whole-cell lux-bacterial sensors. It details how the signal reflects metal bioavailability-speciation processes and cell metabolism contribution.

Chemometrics-assisted electrochemical biosensing of cholesterol as the sole precursor of steroids by a novel electrochemical biosensor

This project was performed with the aims of increasing the sensitivity of differential pulse voltammetry (DPV) which itself is a sensitive electroanalytical technique, and also to compare the area under peak (univariate calibration), height of peak (univariate calibration) and whole of vector (multivariate calibration) for calibration purposes. These topics were investigated by fabrication of a novel electrochemical biosensor for determination of cholesterol (CHO). The procedure used in this project was based on the synthesis of molecularly imprinted polymers (MIPs) to the preconcentration of CHO and its biosensing by a rotating glassy carbon electrode (GCE) modified by co-immobilization of cholesterol oxidase (CO), cholesterol esterase (CE) and horseradish peroxidase (HP) onto multiwalled carbon nanotubes-ionic liquid (COCEHP/MWCNTs-IL/GCE). The results showed that the hydrodynamic DPV (HYDPV) was much more sensitive than DPV and using the area under peak for univariate calibration purposes was more suitable than height of peak. Adsorption at the electrode surface is an important trouble which affects the height and position of voltammetric peaks, but the area under peak is not affected by adsorption therefore, it can be more suitable for univariate calibration purposes. The biosensor response was also calibrated by chronoamperometry and the results confirmed that the HYDPV was more sensitive than chronoamperometry. The next attempt was based on recording the biosensor responses based on second-order HYDPV data and modeling of them (whole of vectors) by three-way calibration methods which showed the best performance among the tested methods for determination of CHO. The biosensor response was long-term stable, repeatable and reproducible which was successfully applied to the analysis of serum sample towards determination of CHO whose results were comparable with a reference method.

An Amperometric Glucose Biosensor Composed of Prussian Blue, Nafion, and Glucose Oxidase Studied by Flow Injection Analysis

The construction and characterization of a glucose biosensor has been implemented in an instrumental chemical analysis course to provide students experience with chemical sensing, electroanalytical methods, and flow injection analysis (FIA). The glucose biosensor was assembled on a glassy carbon electrode by depositing a layer of Prussian blue (PB) electrocatalyst followed by a layer of glucose oxidase (GOx) immobilized in a Nafion film. GOx catalyzed the oxidation of glucose in the presence of dissolved oxygen to yield gluconic acid and hydrogen peroxide. The hydrogen peroxide produced was then electrocatalytically reduced at the PB film with a mild applied potential. The measured cathodic current provided the analytical signal for the glucose biosensor, which was applied to the quantification of glucose levels in soft drinks using FIA techniques. This experiment exposes students to several electroanalytical methods, including amperometry, cyclic voltammetry, and chronocoulometry, as well as methods that are used to measure biosensor response, deposit the PB film, measure the PB film coverage, and examine its electrocatalytic properties. In addition, students gain experience with FIA and the application of an enzyme-based biosensor.

Wearable potentiometric biosensor for analysis of urea in sweat

Herein, we introduce wearable potentiometric biosensors on screen-printed carbon electrodes (SPCEs) for on-body and on-site monitoring of urea in sweat. The biosensor architecture was judiciously designed to detect urea at different pHs and incorporate a pH sensor, thus containing polyaniline ink, urease bioink and a polyvinylchloride membrane. Urea detection could be performed in the wide range from 5 to 200 mM at pH 7.0, encompassing urea levels in human sweat. The biosensor response was fast (incubation time 5 min), with no interference from other substances in sweat. Reliable urea detection could be done in undiluted human sweat with a skin-worn flexible device using the pH correction strategy afforded by the pH sensor. The performance of the epidermal biosensor was not affected by severe bending strains. The feasibility of mass production was demonstrated by fabricating epidermal flexible biosensors using slot-die coating with a roll-to-roll technique.

Sensitive and Specific Detection of Estrogens Featuring Doped Silicon Nanowire Arrays.

Estrogens and estrogen-mimicking compounds in the aquatic environment are known to cause negative impacts to both ecosystems and human health. In this initial proof-of-principle study, we developed a novel vertically oriented silicon nanowire (vSiNW) array-based biosensor for low-cost, highly sensitive and selective detection of estrogens. The vSiNW arrays were formed using an inexpensive and scalable metal-assisted chemical etching (MACE) process. A vSiNW array-based p-n junction diode (vSiNW-diode) transducer design for the biosensor was used and functionalized via 3-aminopropyltriethoxysilane (APTES)-based silane chemistry to bond estrogen receptor-alpha (ER-α) to the surface of the vSiNWs. Following receptor conjugation, the biosensors were exposed to increasing concentrations of estradiol (E2), resulting in a well-calibrated sensor response (R 2 ≥ 0.84, 1-100 ng/mL concentration range). Fluorescence measurements quantified the distribution of estrogen receptors across the vSiNW array compared to planar Si, indicating an average of 7 times higher receptor presence on the vSiNW array surface. We tested the biosensor's target selectivity by comparing it to another estrogen (estrone [E1]) and an androgen (testosterone), where we measured a high positive electrical biosensor response after E1 exposure and a minimal response after testosterone. The regeneration capacity of the biosensor was tested following three successive rinses with phosphate buffer solution (PBS) between hormone exposure. Traditional horizontally oriented Si NW field effect transistor (hSiNW-FET)-based biosensors report electrical current changes at the nanoampere (nA) level that require bulky and expensive measurement equipment making them unsuitable for field measurements, whereas the reported vSiNW-diode biosensor exhibits current changes in the microampere (μA) range, demonstrating up to 100-fold electrical signal amplification, thus enabling sensor signal measurement using inexpensive electronics. The highly sensitive and specific vSiNW-diode biosensor developed here will enable the creation of low-cost, portable, field-deployable biosensors that can detect estrogenic compounds in waterways in real-time.

Surface construction of catalase-immobilized Au/PEDOT nanocomposite on phase-change microcapsules for enhancing electrochemical biosensing detection of hydrogen peroxide

To address the deterioration in electrochemical biosensing detection of hydrogen peroxide (H2O2) caused by enzymatic inactivation in high-temperature environments, we have developed a novel type of catalase-immobilized electroactive phase-change microcapsules based on a phase change material (PCM) core, an SiO2 shell, and a poly(3,4-ethylenedioxythiophene)/Au nanocomposite layer with the surface-immobilized catalase. The microcapsules not only show a well-defined core–shell microstructure with the desired chemical compositions, but also present a high latent heat capacity of over 130 J/g for thermal regulation. An electrochemical biosensor was established using the microcapsule-modified working electrode to obtain a high electrochemical response to H2O2 through combining the highly electroactive layer with covalently bonded catalase. The developed biosensor achieved an enhanced electrochemical biosensing response to H2O2 through regulating the microenvironmental temperature surrounding the working electrode by the latent heat release of its PCM core to prevent enzymatic inactivation at high temperature. Compared to conventional H2O2 biosensors without thermal management by PCMs, the developed biosensor exhibits a higher sensitivity of 42.85 µA·mM−1·cm−2 and a lower detection limit of 3.56 µM at a high working temperature of 50 °C. The developed biosensor has found practical applications for sensitive detection and accurate determination of H2O2 level in real food and biological samples over a wide temperature range.