Biosensing Element Research Articles

Aptasensors for full body health checkup

In the last couple of decades, aptamers have been one of the preferred biomolecular recognition elements in biosensors. The platform of aptasensors (aptamer-based biosensors) is demonstrated to solve practical research problems in various areas. Aptamers and biosensors altogether gift a specific, selective, sensitive, affordable, easy to operate, robust, rapid, stable, and low sample volume analytical platform. In diagnostics, numerous review articles are published discussing the detection of various types of non-infectious and infectious diseases using aptasensors, but to the best of our knowledge, no one has reviewed the available information from the perspective of the utility of aptasensors for a full body health checkup. Therefore, to bridge this existing gap in knowledge in terms of the utility of aptasensors for full body health checkup and to provide a new perspective to the aptasensor community, in this review article, we are reviewing a battery of aptasensors, which has been utilized to detect biomarkers that correlate well with an individual's health status. Our motive in this review is to demonstrate the expansion of utility of aptasensors in routine full body checkups. Systematic Evolution of Ligands by Exponential Enrichment (SELEX), design configurations, and various detection methods employed for aptasensors are also discussed. Along with, the existing gap in the field, current challenges, and future of aptamers in diagnostics with concluding remarks are also presented in this review article.

Aptamer-Based Biosensors for the Colorimetric Detection of Blood Biomarkers: Paving the Way to Clinical Laboratory Testing.

Clinical diagnostics for human diseases rely largely on enzyme immunoassays for the detection of blood biomarkers. Nevertheless, antibody-based test systems have a number of shortcomings that have stimulated a search for alternative diagnostic assays. Oligonucleotide aptamers are now considered as promising molecular recognizing elements for biosensors (aptasensors) due to their high affinity and specificity of target binding. At the moment, a huge variety of aptasensors have been engineered for the detection of various analytes, especially disease biomarkers. However, despite their great potential and excellent characteristics in model systems, only a few of these aptamer-based assays have been translated into practice as diagnostic kits. Here, we will review the current progress in the engineering of aptamer-based colorimetric assays as the most suitable format for clinical lab diagnostics. In particular, we will focus on aptasensors for the detection of blood biomarkers of cardiovascular, malignant, and neurodegenerative diseases along with common inflammation biomarkers. We will also analyze the main obstacles that have to be overcome before aptamer test systems can become tantamount to ELISA for clinical diagnosis purposes.

Electrochemical Biosensor for Rapid Detection of Listeria monocytogenes

Listeria monocytogenes (L. monocytogenes) is a common foodborne pathogen that has been responsible for many foodborne illness outbreaks in recent years. Currently, Listeria contamination in food products is identified only through molecular tests conducted in diagnostic laboratories. No established phage based diagnostic methods for L. monocytogenes during food production or processing are used. Here we report a potentially disruptive rapid diagnostic method based on electrochemical biosensing principles that use bacteriophages as bioreceptors for selective identification and quantification of L. monocytogenes. Electrochemical biosensors are good alternatives to molecular detection methods due to their ease of use, high specificity, sensitivity, and low cost. Bacteriophages can serve as excellent biorecognition elements in biosensors due to their robust stability in a range of environmental conditions and their ability to distinguish between live and dead bacterial cells. The impedimetric biosensing platform for L. monocytogenes detection was developed by immobilizing P100 bacteriophage onto quarternized polyethylenimine modified carbon nanotubes using an in-house developed molecular tethering method. The resulting sensor showed high selectivity and sensitivity toward L. monocytogenes with a limit of detection of 8.4 CFU/ml. Initial results demonstrate that the biosensing platform is highly reliable in its selectivity towards its target analyte, L. monocytogenes.

New PEPTIR-2.0 Peptide Designed for Use as Recognition Element in Electrochemical Biosensors with Improved Specificity towards E. coli O157:H7.

The detection of pathogens through alternative methodologies based on electrochemical biosensors is being studied. These devices exhibit remarkable properties, such as simplicity, specificity, and high sensitivity in monitoring pathogens. However, it is necessary to continue conducting studies that adequately improve these characteristics, especially the recognition molecule. This work aims to design and evaluate a new peptide, named PEPTIR-2.0, as a recognition molecule in electrochemical biosensors to detect E. coli O157:H7 in water. PEPTIR-2.0 was obtained from modifications of the PEPTIR-1.0 peptide sequence, which was previously reported and exhibited excellent properties for detecting and quantifying this pathogenic microorganism. PEPTIR-1.0 is a peptide analogous to the TIR (Translocated Intimin Receptor) protein capable of interacting with the Intimin outer membrane. The basis of this study was to obtain, by using bioinformatics tools, a molecule analogous to PEPTIR-1.0 that maintains its three-dimensional structure but increases the hydrophobic interactions between it and Intimin, since these intermolecular forces are the predominant ones. The designed PEPTIR-2.0 peptide was immobilized on screen-printed electrodes modified with gold nanoparticles. The detection capacity of E. coli O157:H7 in water was evaluated using electrochemical impedance spectroscopy in the presence of other microorganisms, such as P. aeruginosa, S. aureus, and non-pathogenic E. coli. The results showed that PEPTIR-2.0 confers remarkable specificity to the biosensor towards detecting E. coli, even higher than PEPTIR-1.0.

Formation of Island SERS Films on Surfaces of Track Membranes and Silicon Substrates

It is shown that metallic nano-island SERS-active layers can be created on the surfaces of porous track membranes made of polyethylene terephthalate and silicon substrates optimized for exciting laser radiation with wavelengths of 532 and 785 nm. Characteristic patterns of changes in the optical properties of SERS structures upon altering the morphology of metal nanoparticles on a substrate’s surface are established. Such structures can be used as SERS chips and highly sensitive filtering SERS elements in optical biosensors.

Serum Metal Ion-Induced Cross-Linking of Photoelectrochemical Peptides and Circulating Proteins for Evaluating Cardiac Ischemia/Reperfusion.

Patients having experienced the ischemia-reperfusion process are particularly vulnerable to subsequent heart attacks because this process can induce myocardial fibrosis, hallmarked by the release of reactive oxygen species and some proteases, such as cathepsin G, into the circulating blood. If these risk indicators can be monitored from the peripheral serum, early diagnosis and intervention may become a reality. For this purpose, we have designed an assay of free copper ions and cathepsin G in serum using only synthetic small molecules as the biosensing elements. No antibodies are needed to recognize the target protein, and no enzymes are needed to generate and amplify the biosensing signal. In this design, a short peptide can target-specifically recognize protease, while the copper ion in the serum can stimulate the photoelectrochemical activity of the probe, resulting in cross-linking of the serum proteins in a target protein-specific manner. Using this method, serum cathepsin G and free copper ion are found to be significantly elevated in the blood samples collected from patients with acute myocardial infarction and successful percutaneous coronary intervention in comparison with healthy controls, indicating a higher risk of subsequent myocardial injury and cardiovascular events. These results may point to the possible application of the proposed assay to evaluate the severity and prognosis of cardiac ischemia/reperfusion in the near future.

A Strategy for Accessing Nanobody-Based Electrochemical Sensors for Analyte Detection in Complex Media.

Nanobodies are single variable domain antibodies isolated from camelids and are rapidly distinguishing themselves as ideal recognition elements in biosensors due to their comparative stability, ease of production and isolation, and high binding affinities. However, transducing analyte binding by nanobodies in real time is challenging, as most nanobodies do not directly produce an optical or electrical signal upon target recognition. Here, we report a general strategy to fabricate sensitive and selective electrochemical sensors incorporating nanobodies for detecting target analytes in heterogeneous media, such as cell lysate. Graphite felt can be covalently functionalized with recombinant HaloTag-modified nanobodies. Subsequent encapsulation with a thin layer of a hydrogel using a vapor deposition process affords encapsulated electrodes that directly display a decrease in current upon antigen binding, without added redox mediators. Differential pulse voltammetry affords clear and consistent decreases in electrode current across multiple electrode samples for specific antigen concentrations. The change in observed current vs increasing antigen concentration follows Langmuir binding characteristics, as expected. Importantly, selective and repeatable target binding in unpurified cell lysate is only demonstrated by the encapsulated electrode, with an antigen detection limit of ca. 30 pmol, whereas bare electrodes lacking encapsulation produce numerous false positive signals in control experiments.

An all-in-one approach for self-powered sensing: A methanol fuel cell modified with a molecularly imprinted polymer for cancer biomarker detection

• A hybrid methanol fuel cell operating with a biosensor. • One electrode of the fuel cell is the biosensing element. • A molecularly-imprinted polymer tailored in situ for carcinoembryonic antigen. • The overall power of the hybrid cell is carcinoembryonic antigen concentration dependent. • This hybrid fuel cell pursues an application in point-of-care. This work describes the development of an innovative electrochemical biosensor comprehending a passive direct methanol fuel cell (DMFC) assembly, modified by a layer of a molecularly imprinted polymer (MIP) on a carbon fabric anode electrode containing Pt/Ru nanoparticles. This MIP film was prepared from poly(3,4-ethylenedioxythiophene) (PEDOT) and polypyrrole (PPy) obtained by in situ electropolymerization of the corresponding monomers on the anode electrode surface. This MIP film is designed to detect an important cancer biomarker- carcinoembryonic antigen (CEA). This innovative, all-in-one device works in a simple way. First, CEA is incubated on the anode container of the fuel cell, then methanol is added, followed by the response evaluation (polarization curves determination). As CEA selectively interacts with the MIP film, it blocks the methanol's access to the Pt catalyst, remains specific bonded, and interferes with the subsequent polarization curves of the DMFC. Polarization curves obtained in the presence of standard solutions prepared in buffer and human serum confirmed linear responses of log CEA concentration ranging from 30 to 30 000 ng/mL in both media. The biosensor DMFC showed a sensitive response with a detection limit of 4.41 ng/mL when an aqueous 0.05 M methanol solution was used as fuel. When methanol was replaced by an ethanol solution of the same concentration (using the same setup developed for the DMFC), the lower detection limit of 3.52 ng/mL was obtained. Overall, the obtained results show that methanol/ethanol fuel cells operating without flow-through can be successfully used for the fabrication of self-powered biosensors. The novel biosensor concept presented here is simple, inexpensive, and effective, and can be further developed to meet point-of-care requirements.

Recent progress in the design of G-quadruplex–based electrochemical aptasensors

Aptamer-based electrochemical sensors are now developed for the detection of a wide variety of analytes including ions, low-molecular-weight molecules, proteins, and living cells. An aptamer-based sensor is an analytical device whose bio-sensing element (i.e. the aptamer) is immobilized on a transducer surface. Aptasensors have attracted great attention because of their high selectivity, sensitivity, and stability; they could be miniaturized and are of low production cost and offer extraordinary flexibility in the design of their assemblies. This review will emphasize recent developments of aptasensors using aptamers that are able to adopt the particular G-quadruplex (G4) conformations, which are secondary DNA structures formed from guanine-rich sequences. Indeed, G4 exhibits notable recognition properties inherent to their particular structuration. • G4 is involved in the design of electrochem for the sensitive detection of a wide variety of analytes.

Aptamer‐based biosensors and application in tumor theranostics

An aptamer is a short oligonucleotide chain that can specifically recognize targeting analytes. Due to its high specificity, low cost, and good biocompatibility, aptamers as the targeting elements of biosensors have been applied widely in non‐invasive tumor imaging and treatment in situ to replace traditional methods. In this review, we will summarize recent advances in using aptamer‐based biosensors in tumor diagnosis. After a brief introduction of the advantage of aptamers compared with enzyme sensors and immune sensors, the different sensing designs and mechanisms based on 3 signal transduction modes will be reviewed to cover different kinds of analytical methods, including: electrochemistry analysis, colorimetry analysis, and fluorescence analysis. Finally, the prospective advantages of aptamer‐based biosensors in tumor theranostics and post‐treatment monitoring are also evaluated in this review.

Design and Investigation of Dielectrically Modulated Dual-Material Gate-Oxide-Stack Double-Gate TFET for Label-Free Detection of Biomolecules

This article investigates the applicability of dual-material gate-oxide-stack double-gate tunnel field effect transistor (DMGOSDG-TFET) as a biosensing element with the ability to assess the health parameters and disease onset. For this, employment of gate work-function engineering along with the gate-oxide-stack approach and asymmetrical doping at <inline-formula xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"> <tex-math notation="LaTeX">${p}^{+}$ </tex-math></inline-formula> source and <inline-formula xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"> <tex-math notation="LaTeX">${n}^{+}$ </tex-math></inline-formula> drain region are introduced for the first time to implement DMGOSDG-TFET-based biosensor. Also, a nanogap cavity is created by etching a portion of gate dielectric material toward the source end for the accomplishment of biomolecules conjugation in the proposed device. The main focus of this article is to estimate the underlying device sensitivity in the presence of different charged as well as neutral biomolecules. To explore such effects, different dielectric constants and negative charge densities of the biomolecules are considered independently in the nanogap cavity. Next, the sensing performance of the presented device is analyzed in terms of switching-ratio ( <inline-formula xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"> <tex-math notation="LaTeX">${I}_{ \mathrm{\scriptscriptstyle ON}}/{I}_{ \mathrm{\scriptscriptstyle OFF}}$ </tex-math></inline-formula> ), transconductance-to-current ratio ( <inline-formula xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink"> <tex-math notation="LaTeX">${g}_{m}/{I}_{ds}$ </tex-math></inline-formula> ), and average subthreshold-swing. A deep investigation of device performance is also performed with different fillings of the nanogap cavity and step-profiles arising out from the steric hindrance. The device sensitivity is analyzed for different cavity lengths and cavity thicknesses for the best outcomes. In addition, a comparative sensitivity analysis of DMGOSDG-TFET with single-material gate-oxide-stack double-gate tunnel field effect transistor (SMGOSDG-TFET) and metal–oxide–semiconductor field effect transistor (MOSFET)-based biosensor is also presented in this work. The device implementation and all the simulations are carried out using technology computer-aided design (TCAD) tool. All of the sensitivity-assessments disclose that DMGOSDG-TFET can be a good candidate for biosensing applications.

Construction of a p-coumaric and ferulic acid auto-regulatory system in Pseudomonas putida KT2440 for protocatechuate production from lignin-derived aromatics

Lignin is a robust and underutilized aromatic heteropolymer on earth. The effective valorization of lignin into value-added products is an attractive research topic in lignocellulosic biorefineries. However, a low bioconversion rate, high process cost, low yield, and high toxicity of substrates hinder its further applications. In this study, an auto-regulatory system was developed and identified as an effective solution to diminish these challenging bottlenecks. First, a lignin-derived standard model aromatic p-coumaric acid (p-CA)-responsive biosensor was successfully developed through a series of rational engineering approaches in Pseudomonas putida KT2440. Furthermore, an auto-regulatory system was established, which rapidly coexpressed key rate-limiting enzymes, 4-hydroxybenzoate hydroxylase, vanillate-O-demethylase, and transporter HcnK under the biosensor element, to convert the mixture of p-CA and ferulic acid into a value-added platform chemical protocatechuate with a titer of 12.7 g/L. This study demonstrated that the constructed auto-regulatory platform is effective and economical for lignin valorization.

Improving the Room-Temperature Ferromagnetism in ZnO and Low-Doped ZnO:Ag Films Using GLAD Sputtering.

ZnO and doped ZnO films with non-ferromagnetic metal have been widely used as biosensor elements. In these studies, the electrochemical measurements are explored, though the electrical impedance of the system. In this sense, the ferromagnetic properties of the material can be used for multifunctionalization of the sensor element using external magnetic fields during the measurements. Within this context, we investigate the room-temperature ferromagnetism in pure ZnO and Ag-doped ZnO films presenting zigzag-like columnar geometry. Specifically, we focus on the films’ structural and quasi-static magnetic properties and disclose that they evolve with the doping of low-Ag concentrations and the columnar geometry employed during the deposition. The magnetic characterization reveals ferromagnetic behavior at room temperature for all studied samples, including the pure ZnO one. By considering computational simulations, we address the origin of ferromagnetism in ZnO and Ag-doped ZnO and interpret our results in terms of the Zn vacancy dynamics, its substitution by an Ag atom in the site, and the influence of the columnar geometry on the magnetic properties of the films. Our findings bring to light an exciting way to induce/explore the room-temperature ferromagnetism of a non-ferromagnetic metal-doped semiconductor as a promising candidate for biosensor applications.

Electroantennography-based Bio-hybrid Odor-detecting Drone using Silkmoth Antennae for Odor Source Localization

Small drones with chemical or biosensor devices that can detect airborne odorant molecules have attracted considerable attention owing to their applicability in environmental and security monitoring and search-and-rescue operations. Small drones with commercial metal-oxide-semiconductor (MOX) gas sensors have been developed for odor source localization; however, their real-time-odor-detection performance has proven inadequate. However, biosensing technologies based on insect olfactory systems exhibit relatively high sensitivity, selectivity, and real-time response with respect to odorant molecules compared to commercial MOX gas sensors. In such devices, excised insect antennae function as portable odorant biosensor elements and have been found to deliver excellent sensing performance. This study presents experimental protocols for odorant-molecule detection in the air using a small autonomous bio-hybrid drone based on a mountable electroantennography (EAG) device incorporating silkmoth antennae. We developed a mountable EAG device including sensing/processing parts with a Wi-Fi module. The device was equipped with a simple sensor enclosure to enhance the sensor directivity. Thus, odor source localization was conducted using the spiral-surge algorithm, which does not assume an upwind direction. The experimental bio-hybrid odor-detecting drone identified real-time odorant-concentration differences in a pseudo-open environment (outside a wind tunnel) and localized the source. The developed drone and associated system can serve as an efficient odorant molecule-detection tool and a suitable flight platform for developing odor source localization algorithms owing to its high programmability.

Electroantennography-based Bio-hybrid Odor-detecting Drone using Silkmoth Antennae for Odor Source Localization

Small drones with chemical or biosensor devices that can detect airborne odorant molecules have attracted considerable attention owing to their applicability in environmental and security monitoring and search-and-rescue operations. Small drones with commercial metal-oxide-semiconductor (MOX) gas sensors have been developed for odor source localization; however, their real-time-odor-detection performance has proven inadequate. However, biosensing technologies based on insect olfactory systems exhibit relatively high sensitivity, selectivity, and real-time response with respect to odorant molecules compared to commercial MOX gas sensors. In such devices, excised insect antennae function as portable odorant biosensor elements and have been found to deliver excellent sensing performance. This study presents experimental protocols for odorant-molecule detection in the air using a small autonomous bio-hybrid drone based on a mountable electroantennography (EAG) device incorporating silkmoth antennae. We developed a mountable EAG device including sensing/processing parts with a Wi-Fi module. The device was equipped with a simple sensor enclosure to enhance the sensor directivity. Thus, odor source localization was conducted using the spiral-surge algorithm, which does not assume an upwind direction. The experimental bio-hybrid odor-detecting drone identified real-time odorant-concentration differences in a pseudo-open environment (outside a wind tunnel) and localized the source. The developed drone and associated system can serve as an efficient odorant molecule-detection tool and a suitable flight platform for developing odor source localization algorithms owing to its high programmability.

Studies towards the Synthesis of Novel 3-Aminopropoxy-Substituted Dioxins Suitable for the Development of Aptamers for Photonic Biosensor Applications.

Hydroxy-substituted tetrachlorodibenzo[b,e][1,4]dioxin and tetrachlorodibenzo[b,d]furans have been synthesized using 3,4-dichloroanisole, 2,3,6-trichlorophenol and 4,5-dichlorocatechol as starting materials and electrophilic and/or nucleophilic aromatic substitution reactions for the assembly of the dibenzo[b,e][1,4]dioxin and dibenzo[b,d]furan systems. The thus-obtained phenolic compounds were then alkylated with N-1-(4,4-dimethyl-2,6-dioxocyclohexylidene)ethyl (Dde)-protected 3-bromopropan-1-amine to give the corresponding N-Dde protected 3-aminopropoxy-substituted tetrachlorodibenzo[b,e][1,4]dioxin and tetrachlorodibenzo[b,d]furans, respectively. Hydrazinolysis-mediated Dde removal from the former compound provided the corresponding amino-substituted dioxin, which was coupled to carboxy-substituted magnetic beads affording magnetic beads coated by the amino-substituted dioxin. The latter is an attractive intermediate for the development of selective single-standard DNA (ssDNA) aptamers, which constitute molecular recognition elements in photonic biosensors with potential application to the monitoring of the dangerous environmental pollutants, dioxins having serious implications in human health.

Amperometric Biosensors Based on Direct Electron Transfer Enzymes

The accurate determination of analyte concentrations with selective, fast, and robust methods is the key for process control, product analysis, environmental compliance, and medical applications. Enzyme-based biosensors meet these requirements to a high degree and can be operated with simple, cost efficient, and easy to use devices. This review focuses on enzymes capable of direct electron transfer (DET) to electrodes and also the electrode materials which can enable or enhance the DET type bioelectrocatalysis. It presents amperometric biosensors for the quantification of important medical, technical, and environmental analytes and it carves out the requirements for enzymes and electrode materials in DET-based third generation biosensors. This review critically surveys enzymes and biosensors for which DET has been reported. Single- or multi-cofactor enzymes featuring copper centers, hemes, FAD, FMN, or PQQ as prosthetic groups as well as fusion enzymes are presented. Nanomaterials, nanostructured electrodes, chemical surface modifications, and protein immobilization strategies are reviewed for their ability to support direct electrochemistry of enzymes. The combination of both biosensor elements—enzymes and electrodes—is evaluated by comparison of substrate specificity, current density, sensitivity, and the range of detection.

Benchmarking Electrolyte‐Gated Monolayer MoS2 Field‐Effect Transistors in Aqueous Environments

Most electrical sensor and biosensor elements require reliable transducing elements to convert small potential changes into easy to read out current signals. Offering inherent signal magnification and being operable in many relevant environments field‐effect transistors (FETs) are the element of choice in many cases. In particular, using electrolyte gating, numerous sensors and biosensors have been realized in aqueous environments. Over the past years, electrolyte‐gated FETs have been fabricated using a variety of semiconducting materials, including graphene, ZnO, as well as conjugated molecules and polymers. Above all, using conducting polymers top‐performing devices have been achieved. Herein, an approach to use a transition metal dichalcogenide (TMDC)‐based monolayer device as a transducing element is presented. Using MoS2 monolayers, it is shown that such electrolyte‐gated devices may be regarded as very promising transducing elements for sensor and biosensor applications, enabled by their high sensitivity for environmental changes and the possibility of using the naturally occurring sulfur vacancies as grafting points of biorecognition layers. Furthermore, the behavior of such a device under prolonged operation in a dilute biologically relevant electrolyte such as phosphate buffered saline solution (PBS) is reported.

Effect of Spherical Elements of Biosensors and Bioreactors on the Physicochemical Properties of a Peroxidase Protein.

External electromagnetic fields are known to be able to concentrate inside the construction elements of biosensors and bioreactors owing to reflection from their surface. This can lead to changes in the structure of biopolymers (such as proteins), incubated inside these elements, thus influencing their functional properties. Our present study concerned the revelation of the effect of spherical elements, commonly employed in biosensors and bioreactors, on the physicochemical properties of proteins with the example of the horseradish peroxidase (HRP) enzyme. In our experiments, a solution of HRP was incubated within a 30 cm-diameter titanium half-sphere, which was used as a model construction element. Atomic force microscopy (AFM) was employed for the single-molecule visualization of the HRP macromolecules, adsorbed from the test solution onto mica substrates in order to find out whether the incubation of the test HRP solution within the half-sphere influenced the HRP aggregation state. Attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) was employed in order to reveal whether the incubation of HRP solution within the half-sphere led to any changes in its secondary structure. In parallel, spectrophotometry-based estimation of the HRP enzymatic activity was performed in order to find out if the HRP active site was affected by the electromagnetic field under the conditions of our experiments. We revealed an increased aggregation of HRP after the incubation of its solution within the half-sphere in comparison with the control sample incubated far outside the half-sphere. ATR-FTIR allowed us to reveal alterations in HRP’s secondary structure. Such changes in the protein structure did not affect its active site, as was confirmed by spectrophotometry. The effect of spherical elements on a protein solution should be taken into account in the development of the optimized design of biosensors and bioreactors, intended for performing processes involving proteins in biomedicine and biotechnology, including highly sensitive biosensors intended for the diagnosis of socially significant diseases in humans (including oncology, cardiovascular diseases, etc.) at early stages.

Development of three-enzyme lactose amperometric biosensor modified by nanosized poly (meta-phenylenediamine) film

New three-enzyme lactose amperometric biosensor based on platinum disc electrode modified by nanosized semipermeable poly (meta-phenylenediamine) film was developed in this work. It was studied the influence of different conditions of meta-phenylenediamine electropolymerization on the parameters (sensitivity and selectivity) of the amperometric transducers. The most optimal parameters were achieved after performing the electropolymerization of 5 mM meta-phenylenediamine applying 10–13 cyclic voltammograms. The bioselective element of lactose biosensor consisted of three enzymes (β-galactosidase, mutarotase, glucose oxidase) that were immobilized on the surface of modified working electrode. The analytical characteristics of the developed biosensor were studied. The linear range of lactose determination extended from 0.01 mM to 1.25 mM. The response time of the biosensor was 30 s. The biosensor showed high signal reproducibility (RSD = 1.16%), high sensitivity (LOD = 0.005 mM) and high selectivity. The proposed biosensor was tested for measurement of lactose concentration in milk samples. The high correlation between results obtained with lactose biosensor and HPLC was shown (R = 0.96%). Thus, the developed biosensor is suited for rapid, inexpensive, sensitive, selective and simple determination of lactose in milk samples.