Search Results (27)

Anticoagulants, including warfarin, are often administered to patients who are exhibiting early symptoms of thromboembolic episodes or who have already experienced such episodes. However, warfarin has a limited therapeutic index and might cause bleeding and other clinical problems. Warfarin inhibits the vitamin K epoxide reductase complex subunit 1 (VKORC1), an enzyme essential for activating vitamin K, in the coagulation cascade. Genetic factors, such as polymorphisms, can change the natural function of VKORC1, causing variations in the medication reaction among individuals. Hence, before prescribing warfarin, the patient’s genetic profile should also be considered. In this study, an electrochemical genosensor capable of detecting the VKORC1 1639 G>A polymorphism was designed and optimized. This analytical approach detects the electric current obtained during the hybridization reaction between two 52 base pair complementary oligonucleotide sequences. Investigating public bioinformatic platforms, two DNA sequences with the A and G single-nucleotide variants were selected and designed. The experimental protocol of the genosensor implied the formation of a bilayer composed of a thiolate DNA and an alkanethiol immobilized onto gold electrodes, as well as the formation of a DNA duplex using a sandwich-format hybridization reaction through a fluorescein labelled DNA signalling probe and the enzymatic amplification of the electrochemical signal, detected by chronoamperometry. A detection limit of 20 pM and a linear range of 0.05–1.00 nM was obtained. A clear differentiation between A/A, G/A and G/G genotypes in biological samples was successfully identified by his novel device. Full article

Background/Objectives: The CYP2C9 enzyme is involved in the metabolism of warfarin. The CYP2C9 gene harbors several single-nucleotide polymorphisms (SNPs), including CYP2C9*2 (rs1799853), which is known to affect warfarin’s therapeutic response. So, it is important to develop analytical tools capable of genotyping these SNPs to adjust warfarin’s therapeutic outcomes. In this work, an electrochemical DNA-based sensor was constructed and optimized for the detection of the CYP2C9*2 polymorphism. Methods: Using bioinformatic database platforms, two 71 base pair DNA target probes with the polymorphic variants A and G were chosen and designed. A DNA-based sensor was composed by mercaptohexanol and the CYP2C9*2 DNA capture probe in a self-assembled monolayer connected to screen-printed gold electrodes. Two independent hybridization events of the CYP2C9*2 allele were designed using complementary fluorescein-labeled DNA signaling to improve selectivity and avoid secondary structures. Three human samples with the homozygous variant (G/G) and non-variant (A/A) and heterozygous (G/A) genotypes were amplified by PCR and then applied to the developed genosensor. Results: Chronoamperometry measurements were performed for both polymorphic probes. A calibration curve in the 0.25 to 2.50 nM (LOD of 13 pM) and another in the 0.15 to 5.00 nM range (LOD of 22.6 pM) were obtained for the homozygous non-variant and variant probes, respectively. This innovative tool was capable of identifying the hybridization reaction between two complementary strands of immobilized DNA, representing a genotyping alternative to the classical PCR methodology. Conclusions: The developed electrochemical DNA-based sensor was able to discriminate two synthetic SNP target sequences (Target-A and Target-G) and detect, with specificity, the three patients’ genotypes (G/G, G/A, and A/A). This tool is therefore a promising, sensitive, and cost-effective analytical way to determine and discriminate an individual’s genotype and predict the appropriate warfarin dose. Full article

Honey is a natural sweet food product with multiple nutritional and medicinal properties, making it a healthy alternative to processed sugars. With the consumers’ recent interest and purchase of dietary products, the global honey market has greatly increased. To keep up with production or simply for financial gain, some producers/companies are now blending pure honey with cheaper substances that possess similar physical characteristics. As there are no notable visible differences between pure and adulterated honey, it is extremely difficult to determine the purity of the available honeys. In this study, an electrochemical genosensor based on the sandwich format DNA hybridization reaction between two complementary probes was developed for the detection and quantification of Erica arborea pollen DNA in real samples. Analyzing public database platforms, a 98 base-pair DNA-target probe capable of unequivocally detecting the pollen from E. arborea was selected and designed. The complementary probe to the DNA-target oligonucleotide sequence was then cut into a 28-base-pair thiolated DNA-capture probe and a 70-base-pair fluorescein isothiocyanate-labelled DNA-signaling probe. To increase the hybridization reaction, a self-assembled monolayer formed from mixing the DNA-capture probe with mercaptohexanol was employed. Using chronoamperometry, the enzymatic amplification of the electrochemical signal was achieved with a concentration range of 0.03 to 2.00 nM. The DNA from certified E. arborea leaves was extracted using liquid nitrogen and mechanical grinding, and the targeted region was amplified by PCR. The developed genosensor was successfully applied for the detection and quantification of the DNA concentration of the extracted E. arborea plant leaves. Therefore, the developed genosensor is a promising, cost-effective, and innovative analytical method to detect and quantify the DNA concentration of plant DNA in real honey samples. Full article

This study investigates the feasibility of a simple electrochemical detection of Prostate Cancer Antigen 3 (PCA3) fragments extracted from patients’ urine, using a thiolated single-strand DNA probe immobilized on a gold surface without using a redox probe. To enhance the PCA3 recognition process, we conducted a comparative analysis of the hybridization location using two thiolated DNA probes: Probe 1 targets the first 40 bases, while Probe 2 targets the fragment from bases 47 to 86. Hybridization with PCA3 followed, using square wave voltammetry. The limit of detection of the designed genosenors were of the order of (2.2 ng/mL), and (1.6 ng/mL) for Probes 1 and 2, respectively, and the subsequent sensitivities were of the order of (0.09 ± 0.01) µA⁻¹ · µg⁻¹ · mL and (0.10 ± 0.01) µA⁻¹ · µg⁻¹ · mL. Specificity tests were then conducted with the sensor functionalized with Probe 2, as it presents better analytical performances. The electrochemical results indicate that the designed sensor can clearly discriminate a complementary target from a non-complementary one. A further modeling of the calibration curves with the Power Law/Hill model indicates that the dissociation constant increases by one order of magnitude, confirming the ability of the designed sensor to perfectly discriminate complementary targets from non-complementary ones. Full article

The BDNF gene is associated with high degrees of variability in antidepressant treatments. The Val66Met polymorphism is widely known as a source of this variability, warranting growing interest in genotyping patients that undergo antidepressant treatment to better suit their needs. This paper reports on an electrochemical genosensing platform, based on gold electrodes, capable of detecting this polymorphism, through the use of synthetic enzymatically labeled DNA probes for two different BDNF alleles. The sensor showed promising results, and its applicability to real samples is currently being tested. Full article

The rapid and sensitive detection of food contaminants is becoming increasingly important for timely prevention and treatment of foodborne disease. In this review, we discuss recent developments of electrochemical biosensors as facile, rapid, sensitive, and user-friendly analytical devices and their applications in food safety analysis, owing to the analytical characteristics of electrochemical detection and to advances in the design and production of bioreceptors (antibodies, DNA, aptamers, peptides, molecular imprinted polymers, enzymes, bacteriophages, etc.). They can offer a low limit of detection required for food contaminants such as allergens, pesticides, antibiotic traces, toxins, bacteria, etc. We provide an overview of a broad range of electrochemical biosensing designs and consider future opportunities for this technology in food control. Full article

In view of the presence of pathogenic Vibrio cholerae (V. cholerae) bacteria in environmental waters, including drinking water, which may pose a potential health risk to humans, an ultrasensitive electrochemical DNA biosensor for rapid detection of V. cholerae DNA in the environmental sample was developed. Silica nanospheres were functionalized with 3-aminopropyltriethoxysilane (APTS) for effective immobilization of the capture probe, and gold nanoparticles were used for acceleration of electron transfer to the electrode surface. The aminated capture probe was immobilized onto the Si-Au nanocomposite-modified carbon screen printed electrode (Si-Au-SPE) via an imine covalent bond with glutaraldehyde (GA), which served as the bifunctional cross-linking agent. The targeted DNA sequence of V. cholerae was monitored via a sandwich DNA hybridization strategy with a pair of DNA probes, which included the capture probe and reporter probe that flanked the complementary DNA (cDNA), and evaluated by differential pulse voltammetry (DPV) in the presence of an anthraquninone redox label. Under optimum sandwich hybridization conditions, the voltammetric genosensor could detect the targeted V. cholerae gene from 1.0 × 10⁻¹⁷–1.0 × 10⁻⁷ M cDNA with a limit of detection (LOD) of 1.25 × 10⁻¹⁸ M (i.e., 1.1513 × 10⁻¹³ µg/µL) and long-term stability of the DNA biosensor up to 55 days. The electrochemical DNA biosensor was capable of giving a reproducible DPV signal with a relative standard deviation (RSD) of <5.0% (n = 5). Satisfactory recoveries of V. cholerae cDNA concentration from different bacterial strains, river water, and cabbage samples were obtained between 96.5% and 101.6% with the proposed DNA sandwich biosensing procedure. The V. cholerae DNA concentrations determined by the sandwich-type electrochemical genosensor in the environmental samples were correlated to the number of bacterial colonies obtained from standard microbiological procedures (bacterial colony count reference method). Full article

The present work developed an electrochemical genosensor for the detection of virulence outer membrane protein A (ompA, tDNA) gene of Cronobacter sakazakii (C. sakazakii) by exploiting the excellent glucose-oxidase-mimicking activity of copper Metal-organic frameworks (Cu-MOF) doped with gold nanoparticle (AuNPs). The signal nanotags of signal probes (sDNA) that biofunctionalized AuNPs@Cu-MOF (sDNA-AuNPs@Cu-MOF) were designed using an Au-S bond. The biosensor was prepared by immobilization capture probes (cDNA) onto an electrodeposited AuNPs-modified glassy carbon electrode (GCE). AuNPs@Cu-MOF was introduced onto the surface of the GCE via a hybridization reaction between cDNA and tDNA, as well as tDNA and sDNA. Due to the enhanced oxidase-mimicking activity of AuNPs@Cu-MOF to glucose, the biosensor gave a linear range of 1.0 × 10⁻¹⁵ to 1.0 × 10⁻⁹ mol L⁻¹ to tDNA with a detection limit (LOD) of 0.42 fmol L⁻¹ under optimized conditions using differential pulse voltammetry measurement (DPV). It can be applied in the direct detection of ompA gene segments in total DNA extracts from C. sakazakii with a broad linear range of 5.4−5.4 × 10⁵ CFU mL⁻¹ and a LOD of 0.35 CFU mL⁻¹. The biosensor showed good selectivity, fabricating reproducibility and storage stability, and can be used for the detection of ompA gene segments in real samples with recovery between 87.5% and 107.3%. Full article

Electrochemical biosensors are known as analytical tools, guaranteeing rapid and on-site results in medical diagnostics, food safety, environmental protection, and life sciences research. Current research focuses on developing sensors for specific targets and addresses challenges to be solved before their commercialization. These challenges typically include the lowering of the limit of detection, the widening of the linear concentration range, the analysis of real samples in a real environment and the comparison with a standard validation method. Nowadays, functional nanomaterials are designed and applied in electrochemical biosensing to support all these challenges. This review will address the integration of functional nanomaterials in the development of electrochemical biosensors for the rapid diagnosis of viral infections, such as COVID-19, middle east respiratory syndrome (MERS), influenza, hepatitis, human immunodeficiency virus (HIV), and dengue, among others. The role and relevance of the nanomaterial, the type of biosensor, and the electrochemical technique adopted will be discussed. Finally, the critical issues in applying laboratory research to the analysis of real samples, future perspectives, and commercialization aspects of electrochemical biosensors for virus detection will be analyzed. Full article

Tuberculosis (TB) has been a devastating human illness for thousands of years. According to the WHO, around 10.4 million new cases of tuberculosis are identified every year, with 1.8 million deaths. To reduce these statistics and the mortality rate, an early and accurate TB diagnosis is essential. This study offers a highly sensitive and selective electrochemical biosensor for Mycobacterium tuberculosis (MTB) detection based on a ternary nanocomposite of reduced graphene oxide, polydopamine, and gold nanoparticles (rGO-PDA-AuNP). Avidin-biotin coupling was used to bind the MTB probe DNA onto the rGO-PDA-AuNP modified glassy carbon electrode (ssDNA/avidin/rGO-PDA-AuNP). UV-Visible, Raman, XRD, and TEM were used to evaluate the structural and morphological characteristics of rGO-PDA-AuNP. Furthermore, DNA immobilization is validated using FESEM and FT-IR techniques. The modified electrodes were electrochemically analyzed using cyclic voltammetry (CV) and linear sweep voltammetry (LSV), and the results indicate that the produced electrode can detect target DNA up to 0.1 × 10⁻⁷ mM with 2.12 × 10⁻³ mA µM⁻¹ sensitivity and a response time of 5 s. The constructed genosensor displayed high sensitivity and stability, and it also provides a unique strategy for diagnosing MTB at an early stage. Furthermore, our rGO-PDA-AuNP/GCE-based electrochemical platform has broad potential for creating biosensor systems for detecting various infectious pathogens and therapeutically significant biomarkers. Full article

In this work, chitosan beads were used as a cost-effective platform for the covalent immobilization of unmodified single-stranded DNA, using glutaraldehyde as a cross-linking agent. The immobilized DNA capture probe was hybridized in the presence of miRNA-222 as a complementary sequence. The target was evaluated based on the electrochemical response of the released guanine, using hydrochloride acid as a hydrolysis agent. Differential pulse voltammetry technique and screen-printed electrodes modified with COOH-functionalized carbon black were used to monitor the released guanine response before and after hybridization. The functionalized carbon black provided an important signal amplification of guanine compared to the other studied nanomaterials. Under optimal conditions (6 M HCl at 65 °C for 90 min), an electrochemical-based label-free genosensor assay exhibited a linear range between 1 nM and 1 µM of miRNA-222, with a detection limit of 0.2 nM of miRNA-222. The developed sensor was successfully used to quantify miRNA-222 in a human serum sample. Full article

Viruses, including influenza viruses, MERS-CoV (Middle East respiratory syndrome coronavirus), SARS-CoV (severe acute respiratory syndrome coronavirus), HAV (Hepatitis A virus), HBV (Hepatitis B virus), HCV (Hepatitis C virus), HIV (human immunodeficiency virus), EBOV (Ebola virus), ZIKV (Zika virus), and most recently SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2), are responsible for many diseases that result in hundreds of thousands of deaths yearly. The ongoing outbreak of the COVID-19 disease has raised a global concern and intensified research on the detection of viruses and virus-related diseases. Novel methods for the sensitive, rapid, and on-site detection of pathogens, such as the recent SARS-CoV-2, are critical for diagnosing and treating infectious diseases before they spread and affect human health worldwide. In this sense, electrochemical impedimetric biosensors could be applied for virus detection on a large scale. This review focuses on the recent developments in electrochemical-impedimetric biosensors for the detection of viruses. Full article

Acinetobacter baumannii (A. baumannii) are phenotypically indistinguishable from the Acinetobacter calcoaceticus–A. baumannii (ACB) complex members using routine laboratory methods. Early diagnosis plays an important role in controlling A. baumannii infections and this could be assisted by the development of a rapid, yet sensitive diagnostic test. In this study, we developed an enzyme-based electrochemical genosensor for asymmetric PCR (aPCR) amplicon detection of the bla_OXA-51-like gene in A. baumannii. A. baumanniibla_OXA-51-like gene PCR primers were designed, having the reverse primer modified at the 5′ end with FAM. A bla_OXA-51-like gene sequence-specific biotin labelled capture probe was designed and immobilized using a synthetic oligomer (FAM-labelled) deposited on the working electrode of a streptavidin-modified, screen-printed carbon electrode (SPCE). The zot gene was used as an internal control with biotin and FAM labelled as forward and reverse primers, respectively. The bla_OXA-51-like gene was amplified using asymmetric PCR (aPCR) to generate single-stranded amplicons that were detected using the designed SPCE. The amperometric current response was detected with a peroxidase-conjugated, anti-fluorescein antibody. The assay was tested using reference and clinical A. baumannii strains and other nosocomial bacteria. The analytical sensitivity of the assay at the genomic level and bacterial cell level was 0.5 pg/mL (1.443 µA) and 10³ CFU/mL, respectively. The assay was 100% specific and sensitive for A. baumannii. Based on accelerated stability performance, the developed genosensor was stable for 1.6 years when stored at 4 °C and up to 28 days at >25 °C. The developed electrochemical genosensor is specific and sensitive and could be useful for rapid, accurate diagnosis of A. baumannii infections even in temperate regions. Full article

Food allergy has been indicated as the most frequent adverse reaction to food ingredients over the past few years. Since the only way to avoid the occurrence of allergic phenomena is to eliminate allergenic foods, it is essential to have complete and accurate information on the components of foodstuff. In this framework, it is mandatory and crucial to provide fast, cost-effective, affordable, and reliable analysis methods for the screening of specific allergen content in food products. This review reports the research advancements concerning food allergen detection, involving electrochemical biosensors. It focuses on the sensing strategies evidencing different types of recognition elements such as antibodies, nucleic acids, and cells, among others, the nanomaterial role, the several electrochemical techniques involved and last, but not least, the ad hoc electrodic surface modification approaches. Moreover, a selection of the most recent electrochemical sensors for allergen detection are reported and critically analyzed in terms of the sensors’ analytical performances. Finally, advantages, limitations, and potentialities for practical applications of electrochemical biosensors for allergens are discussed. Full article

Nanoengineering biosensors have become more precise and sophisticated, raising the demand for highly sensitive architectures to monitor target analytes at extremely low concentrations often required, for example, for biomedical applications. We review recent advances in functional nanomaterials, mainly based on novel organic-inorganic hybrids with enhanced electro-physicochemical properties toward fulfilling this need. In this context, this review classifies some recently engineered organic-inorganic metallic-, silicon-, carbonaceous-, and polymeric-nanomaterials and describes their structural properties and features when incorporated into biosensing systems. It further shows the latest advances in ultrasensitive electrochemical biosensors engineered from such innovative nanomaterials highlighting their advantages concerning the concomitant constituents acting alone, fulfilling the gap from other reviews in the literature. Finally, it mentioned the limitations and opportunities of hybrid nanomaterials from the point of view of current nanotechnology and future considerations for advancing their use in enhanced electrochemical platforms. Full article