Enhanced enzymatic electrochemical detection of an organophosphate Pesticide: Achieving Wide linearity and femtomolar detection via gold nanoparticles growth within polypyrrole films.

The indiscriminate use of pesticides in agriculture demands the development of devices capable of monitoring contaminations in food supplies, in the environment and biological fluids. Simplicity, easy handling, high sensitivities, and low limits-of-detection (LOD) and quantification are some of the required properties for these devices. In this work, we evaluated the effect of incorporating gold nanoparticles into indigo carmine-doped polypyrrole during the electropolymerization of films for use as an acetylcholinesterase (AChE) enzyme-based biosensor. As proof of concept, the pesticide methyl parathion was tested towards the inhibition of AChE. The enzyme was immobilized simply by drop-casting a solution, eliminating the need for any prior surface modification. The biosensors were characterized with cyclic voltammetry, scanning electron microscopy, transmission electron microscopy, and Raman spectroscopy. The assays for the detection of methyl parathion with films containing polypyrrole, indigo carmine and AChE (PPy-IC-AChE) presented a sensitivity of 5.7 µA cm-2 g-1 mL and a LOD of 12 nmol L-1 (3.0 ng L-1) with a linear range from 1.3 x 10-7 mol L-1 to 1.0 x 10-5 mol L-1. The introduction of gold nanoparticles (AuNP) into the film (PPy-IC-AuNP-AChE) led to remarkable improvements on the overall performance, such as a lower redox potential for the enzymatic reaction, a 145 % increase in sensitivity (14 µA cm-2 g-1 mL), a wider detection dynamic range (from 1.3x10-7 to 1.0x10-3 mol L-1), and a very low LOD of 24 fmol L-1 (64 ag mL-1). These findings underscore the potential of using AuNPs to improve the enzymatic performance of biosensor devices.

A disposable, portable electrochemical immunosensor for rapid in situ detection of bovine tuberculosis.

This contribution describes the development of a simple, fast, cost-effective, and sensitive impedimetric immunosensor for quantifying bovine tuberculosis (TB) in bovine serum samples. The construction of the immunosensor involved immobilizing the purified protein derivative (PPD) of M. bovis onto a screen-printed electrode that was modified with gold nanoparticles (AuNPs) and a polypyrrole (pPy) film synthesized electrochemically. The immunosensor exhibited a linear range from 0.5 µg mL-1 to 100 µg mL-1 and achieved a limit of detection (LD) of 100 ng mL-1 for the detection of anti-M. bovis antibody. The recovery percentages obtained in bovine serum samples were excellent, ranging between 98 % and 103 %. This device presents several advantages over alternative methods for determining TB in bovine serum samples. These include direct, in situ measurement without the need for pre-treatment, utilization of small volumes, thus avoiding harmful solvents and expensive reagents, and portability. In addition, the immunosensor exhibits both physical and chemical stability, retaining effectiveness even after 30 days of modification. This allows simultaneous incubations and facilitates large-scale detection. Hence, this immunosensor presents itself as a promising diagnostic tool for detecting anti-M. bovis antibodies in bovine serum. It serves as a viable alternative to tuberculin and ELISA tests.

Parkinson biomarker determination with an Au microflower-enhanced electrochemical immunosensor using non-Faradaic capacitance measurements.

This work comprehends the development and characterization of a carbon black-based electrode modified with Au microflowers to increase its effect as a capacitance biosensor for the determination of PARK7/DJ-1. Due to its high surface-to-volume ratio and biocompatibility, Au particles are suitable for antibody binding, and by monitoring surface capacitance, it is possible to identify the immune-pair interaction. Au microflowers allowed the adequate immobilization of Parkinsonian-related proteins: PARK7/DJ-1 and its antibody. The protein is associated with several antioxidant mechanisms, but its abnormal concentrations or mutations can be the cause of the loss of dopaminergic neurons, leading to Parkinson's disease. The device was characterized by scanning electron microscopy and cyclic voltammetry, revealing the flower-like structures and the electrochemically-interest enhancements they provide, such as increased heterogeneous electron transfer rate coefficient and electroactive area. The self-assembled monolayers of different molecules were optimized with the aid of 22 central composite experiments and a linear calibration curve was obtained between 0.700 and 120 ng mL-1 of PARK7/DJ-1, with a limit of detection of 0.207 ng mL-1. The data confirms that the addition of Au microflowers enhanced the electrochemical signal of the device, as well as allowed for the determination of an early stage Parkinson's disease biomarker with appreciable analytical performance.

Disentangling the Regulatory Response of Agrobacterium tumefaciens CHLDO to Glyphosate for Engineering Whole-Cell Phosphonate Biosensors.

Phosphonates (PHTs), organic compounds with a stable C-P bond, are widely distributed in nature. Glyphosate (GP), a synthetic PHT, is extensively used in agriculture and has been linked to various human health issues and environmental damage. Given the prevalence of GP, developing cost-effective, on-site methods for GP detection is key for assessing pollution and reducing exposure risks. We adopted Agrobacterium tumefaciens CHLDO, a natural GP degrader, as a host and the source of genetic parts for constructing PHT biosensors. In this bacterial species, the phn gene cluster, encoding the C-P lyase pathway, is regulated by the PhnF transcriptional repressor. We selected the phnG promoter, which displays a dose-dependent response to GP, to build a set of whole-cell biosensors. Through stepwise genetic optimization of the transcriptional cascade, we created a whole-cell biosensor capable of detecting GP in the 0.25-50 µM range in various samples, including soil and water.

Diagnosis of viral infectious diseases through sensitive detection of human serum antibodies using a modified label-free electrochemical biosensor.

Rapid virus identification is crucial for preventing outbreaks. The COVID-19 pandemic has highlighted the critical nature of rapid virus detection. Here, we designed a label-free electrochemical biosensor modified with gold nanoparticles (AuNPs) to detect IgG antibodies from human serum, enabling rapid point-of-care diagnostics. AuNPs were synthesized and characterized. A multivariate optimization was carried out to determine the optimal condition for functionalizing AuNPs with anti-IgG. Subsequently, using a glassy carbon electrode (GCE), a modified AuNPs/GCE electrochemical biosensor was developed for IgG detection. The results indicated that AuNPs displayed a spherical morphology with a size distribution of 19.54 nm. Additionally, the zeta potential was recorded at -7.84 mV. Central composite design (CCD) analysis determined the optimal conditions for functionalizing AuNPs to be an anti-IgG concentration of 320 µg mL-1, a temperature of 25 °C, and pH of 7.4. The characterization study confirmed the successful synthesis and functionalization of AuNPs. Through electrochemical impedance spectroscopy measurement, the biosensor demonstrated a limit of detection (LOD) of 0.2 ng mL-1 and limit of quantification (LOQ) of 0.8 ng mL-1. Furthermore, tests in real samples showed the interaction between IgG antibodies in serum samples and AuNPs/GCE, confirming the biosensor's ability to detect and quantify IgG in clinical samples.

Multi-walled carbon nanotubes functionalized with a new Schiff base containing phenylboronic acid residues: application to the development of a bienzymatic glucose biosensor using a response surface methodology approach.

An innovative supramolecular architecture is reported for bienzymatic glucose biosensing based on the use of a nanohybrid made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with a Schiff base modified with two phenylboronic acid residues (SB-dBA) as platform for the site-specific immobilization of the glycoproteins glucose oxidase (GOx) and horseradish peroxidase (HRP). The analytical signal was obtained from amperometric experiments at - 0.050 V in the presence of 5.0 × 10-4 M hydroquinone as redox mediator. The concentration of GOx and HRP and the interaction time between the enzymes and the nanohybrid MWCNT-SB-dBA deposited at glassy carbon electrodes (GCEs) were optimized through a central composite design (CCD)/response surface methodology (RSM). The optimal concentrations of GOx and HRP were 3.0 mg mL-1 and 1.50 mg mL-1, respectively, while the optimum interaction time was 3.0 min. The bienzymatic biosensor presented a sensitivity of (24 ± 2) × 102 µA dL mg-1 ((44 ± 4) × 102 µA M-1), a linear range between 0.06 mg dL-1 and 21.6 mg dL-1 (3.1 µM-1.2 mM) (R2 = 0.9991), and detection and quantification limits of 0.02 mg dL-1 (1.0 µM) and 0.06 mg dL-1 (3.1 µM), respectively. The reproducibility for five sensors prepared with the same MWCNT-SB-dBA nanohybrid was 6.3%, while the reproducibility for sensors prepared with five different nanohybrids and five electrodes each was 7.9%. The GCE/MWCNT-SB-dBA/GOx-HRP was successfully used for the quantification of glucose in artificial human urine and commercial human serum samples.

Red and near-infrared light-activated photoelectrochemical nanobiosensors for biomedical target detection.

Photoelectrochemical (PEC) nanobiosensors integrate molecular (bio)recognition elements with semiconductor/plasmonic photoactive nanomaterials to produce measurable signals after light-induced reactions. Recent advancements in PEC nanobiosensors, using light-matter interactions, have significantly improved sensitivity, specificity, and signal-to-noise ratio in detecting (bio)analytes. Tunable nanomaterials activated by a wide spectral radiation window coupled to electrochemical transduction platforms have further improved detection by stabilizing and amplifying electrical signals. This work reviews PEC biosensors based on nanomaterials like metal oxides, carbon nitrides, quantum dots, and transition metal chalcogenides (TMCs), showing their superior optoelectronic properties and analytical performance for the detection of clinically relevant biomarkers. Furthermore, it highlights the innovative role of red light and NIR-activated PEC nanobiosensors in enhancing charge transfer processes, protecting them from biomolecule photodamage in vitro and in vivo applications. Overall, advances in PEC detection systems have the potential to revolutionize rapid and accurate measurements in clinical diagnostic applications. Their integration into miniaturized devices also supports the development of portable, easy-to-use diagnostic tools, facilitating point-of-care (POC) testing solutions and real-time monitoring.

Microfluid biosensor for detection of hpv in patient scraping samples: Determining E6 and E7 oncogenes.

E6 and E7 oncogenes are pivotal in the carcinogenic transformation in HPV infections and efficient diagnostic methods can ensure the detection and differentiation of HPV genotype. This study describes the development and validation of an electrochemical, label-free genosensor coupled with a microfluidic system for detecting the E6 and E7 oncogenes in cervical scraping samples. The nanostructuring employed was based on a cysteine and graphene quantum dots layer that provides functional groups, surface area, and interesting electrochemical properties. Biorecognition tests with cervical scraping samples showed differentiation in the voltammetric response. Low-risk HPV exhibited a lower biorecognition response, reflected in ΔI% values of 82.33 % ± 0.29 for HPV06 and 80.65 % ± 0.68 for HPV11 at a dilution of 1:100. Meanwhile, high-risk, HPV16 and HPV18, demonstrated ΔI% values of 96.65 % ± 1.27 and 93 % ± 0.026, respectively, at the same dilution. Therefore, the biorecognition intensity followed the order: HPV16 >HPV18 >HPV06 >HPV11. The limit of detection and the limit of quantification of E6E7 microfluidic LOC-Genosensor was 26 fM, and 79.6 fM. Consequently, the E6E7 biosensor is a valuable alternative for clinical HPV diagnosis, capable of detecting the potential for oncogenic progression even in the early stages of infection.

The Tunable Parameters of Graphene-Based Biosensors.

Graphene-based surface plasmon resonance (SPR) biosensors have emerged as a promising technology for the highly sensitive and accurate detection of biomolecules. This study presents a comprehensive theoretical analysis of graphene-based SPR biosensors, focusing on configurations with single and bimetallic metallic layers. In this study, we investigated the impact of various metallic substrates, including gold and silver, and the number of graphene layers on key performance metrics: sensitivity of detection, detection accuracy, and quality factor. Our findings reveal that configurations with graphene first supported on gold exhibit superior performance, with sensitivity of detection enhancements up to 30% for ten graphene layers. In contrast, silver-supported configurations, while demonstrating high sensitivity, face challenges in maintaining detection accuracy. Additionally, reducing the thickness of metallic layers by 30% optimizes light coupling and enhances sensor performance. These insights highlight the significant potential of graphene-based SPR biosensors in achieving high sensitivity of detection and reliability, paving the way for their application in diverse biosensing technologies. Our findings pretend to motivate future research focusing on optimizing metallic layer thickness, improving the stability of silver-supported configurations, and experimentally validating the theoretical findings to further advance the development of high-performance SPR biosensors.

Digitalization of Enzyme-Linked Immunosorbent Assay with Graphene Field-Effect Transistors (G-ELISA) for Portable Ferritin Determination.

Herein, we present a novel approach to quantify ferritin based on the integration of an Enzyme-Linked Immunosorbent Assay (ELISA) protocol on a Graphene Field-Effect Transistor (gFET) for bioelectronic immunosensing. The G-ELISA strategy takes advantage of the gFET inherent capability of detecting pH changes for the amplification of ferritin detection using urease as a reporter enzyme, which catalyzes the hydrolysis of urea generating a local pH increment. A portable field-effect transistor reader and electrolyte-gated gFET arrangement are employed, enabling their operation in aqueous conditions at low potentials, which is crucial for effective biological sample detection. The graphene surface is functionalized with monoclonal anti-ferritin antibodies, along with an antifouling agent, to enhance the assay specificity and sensitivity. Markedly, G-ELISA exhibits outstanding sensing performance, reaching a lower limit of detection (LOD) and higher sensitivity in ferritin quantification than unamplified gFETs. Additionally, they offer rapid detection, capable of measuring ferritin concentrations in approximately 50 min. Because of the capacity of transistor miniaturization, our innovative G-ELISA approach holds promise for the portable bioelectronic detection of multiple biomarkers using a small amount of the sample, which would be a great advancement in point-of-care testing.

Conceptually innovative fluorophores for functional bioimaging.

Fluorophore chemistry is at the forefront of bioimaging, revolutionizing the visualization of biological processes with unparalleled precision. From the serendipitous discovery of mauveine in 1856 to cutting-edge fluorophore engineering, this field has undergone transformative evolution. Today, the synergy of chemistry, biology, and imaging technologies has produced diverse, specialized fluorophores that enhance brightness, photostability, and targeting capabilities. This review delves into the history and innovation of fluorescent probes, showcasing their pivotal role in advancing our understanding of cellular dynamics and disease mechanisms. We highlight groundbreaking molecules and their applications, envisioning future breakthroughs that promise to redefine biomedical research and diagnostics.

Metal Nanoparticle-Based Biosensors for the Early Diagnosis of Infectious Diseases Caused by ESKAPE Pathogens in the Fight against the Antimicrobial-Resistance Crisis.

The species included in the ESKAPE group (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and the genus Enterobacter) have a high capacity to develop antimicrobial resistance (AMR), a health problem that is already among the leading causes of death and could kill 10 million people a year by 2050. The generation of new potentially therapeutic molecules has been insufficient to combat the AMR "crisis", and the World Health Organization (WHO) has stated that it will seek to promote the development of rapid diagnostic strategies. The physicochemical properties of metallic nanoparticles (MNPs) have made it possible to design biosensors capable of identifying low concentrations of ESKAPE bacteria in the short term; other systems identify antimicrobial susceptibility, and some have been designed with dual activity in situ (bacterial detection and antimicrobial activity), which suggests that, in the near future, multifunctional biosensors could exist based on MNPs capable of quickly identifying bacterial pathogens in clinical niches might become commercially available. This review focuses on the use of MNP-based systems for the rapid and accurate identification of clinically important bacterial pathogens, exhibiting the necessity for exhaustive research to achieve these objectives. This review focuses on the use of metal nanoparticle-based systems for the rapid and accurate identification of clinically important bacterial pathogens.

Ultrasonic Sensor: A Fast and Non-Destructive System to Measure the Viscosity and Density of Molecular Fluids.

This study presents the design and development of an ultrasonic sensor as a fundamental tool for characterizing the properties of fluids and biofluids. The analysis primarily focuses on measuring the electrical parameters of the system, which correlate with the density and viscosity of the solutions, in sample volumes of microliters and with high temporal resolution (up to 1 data point per second). The use of this sensor allows the fast and non-destructive evaluation of the viscosity and density of fluids deposited on its free surface. The measurements are based on obtaining the impedance versus frequency curve and the phase difference curve (between current and voltage) versus frequency. In this way, characteristic parameters of the transducer, such as the resonance frequency, phase, minimum impedance, and the quality factor of the resonant system, can characterize variations in density and viscosity in the fluid under study. The results obtained revealed the sensor's ability to identify two parameters sensitive to viscosity and two parameters sensitive to density. As a proof of concept, the unfolding of the bovine albumin protein was studied, resulting in a curve that reflects its unfolding kinetics in the presence of urea.

Upconverting Nanoparticle-based Enhanced Luminescence Lateral-Flow Assay for Urinary Biomarker Monitoring.

Development of efficient portable sensors for accurately detecting biomarkers is crucial for early disease diagnosis, yet remains a significant challenge. To address this need, we introduce the enhanced luminescence lateral-flow assay, which leverages highly luminescent upconverting nanoparticles (UCNPs) alongside a portable reader and a smartphone app. The sensor's efficiency and versatility were shown for kidney health monitoring as a proof of concept. We engineered Er3+- and Tm3+-doped UCNPs coated with multiple layers, including an undoped inert matrix shell, a mesoporous silica shell, and an outer layer of gold (UCNP@mSiO2@Au). These coatings synergistically enhance emission by over 40-fold and facilitate biomolecule conjugation, rendering UCNP@mSiO2@Au easy to use and suitable for a broad range of bioapplications. Employing these optimized nanoparticles in lateral-flow assays, we successfully detected two acute kidney injury-related biomarkers─kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL)─in urine samples. Using our sensor platform, KIM-1 and NGAL can be accurately detected and quantified within the range of 0.1 to 20 ng/mL, boasting impressively low limits of detection at 0.28 and 0.23 ng/mL, respectively. Validating our approach, we analyzed clinical urine samples, achieving biomarker concentrations that closely correlated with results obtained via ELISA. Importantly, our system enables biomarker quantification in less than 15 min, underscoring the performance of our novel UCNP-based approach and its potential as reliable, rapid, and user-friendly diagnostics.

CRISPR-based electrochemical biosensors: an alternative for point-of-care diagnostics?

The combination of CRISPR technology and electrochemical sensors has sparked a paradigm shift in the landscape of point-of-care (POC) diagnostics. This review explores the dynamic convergence between CRISPR and electrochemical sensing, elucidating their roles in rapid and precise biosensing platforms. CRISPR, renowned for its remarkable precision in genome editing and programmability capability, has found a novel application in conjunction with electrochemical sensors, promising highly sensitive and specific detection of nucleic acids and biomarkers associated with diverse diseases. This article navigates through fundamental principles, research developments, and applications of CRISPR-based electrochemical sensors, highlighting their potential to revolutionize healthcare accessibility and patient outcomes. In addition, some key points and challenges regarding applying CRISPR-powered electrochemical sensors in real POC settings are presented. By discussing recent advancements and challenges in this interdisciplinary field, this review evaluates the potential of these innovative sensors as an alternative for decentralized, rapid, and accurate POC testing, offering some insights into their applications across clinical scenarios and their impact on the future of diagnostics.

Detection of foodborne pathogens in contaminated food using nanomaterial-based electrochemical biosensors.

Foodborne pathogens are a grave concern for the for food, medical, environmental, and economic sectors. Their ease of transmission and resistance to treatments, such as antimicrobial agents, make them an important challenge. Food tainted with these pathogens is swiftly rejected, and if ingested, can result in severe illnesses and even fatalities. This review provides and overview of the current status of various pathogens and their metabolites transmitted through food. Despite a plethora of studies on treatments to eradicate and inhibit these pathogens, their indiscriminate use can compromise the sensory properties of food and lead to contamination. Therefore, the study of detection methods such as electrochemical biosensors has been proposed, which are devices with advantages such as simplicity, fast response, and sensitivity. However, these biosensors may also present some limitations. In this regard, it has been reported that nanomaterials with high conductivity, surface-to-volume ratio, and robustness have been observed to improve the detection of foodborne pathogens or their metabolites. Therefore, in this work, we analyze the detection of pathogens transmitted through food and their metabolites using electrochemical biosensors based on nanomaterials.

Role of Graphene in Surface Plasmon Resonance-Based Biosensors.

This work explores the transformative role of graphene in enhancing the performance of surface plasmon resonance (SPR)-based biosensors. The motivation for this review stems from the growing interest in the unique properties of graphene, such as high surface area, excellent electrical conductivity, and versatile functionalization capabilities, which offer significant potential to improve the sensitivity, specificity, and stability of SPR biosensors. This review systematically analyzes studies published between 2010 and 2023, covering key metrics of biosensor performance. The findings reveal that the integration of graphene consistently enhances sensitivity. Specificity, although less frequently reported numerically, showed promising results, with high specificity achieved at sub-nanomolar concentrations. Stability enhancements are also significant, attributed to the protective properties of graphene and improved biomolecule adsorption. Future research should focus on mechanistic insights, optimization of integration techniques, practical application testing, scalable fabrication methods, and comprehensive comparative studies. Our findings provide a foundation for future research, aiming to further optimize and harness the unique physical properties of graphene to meet the demands of sensitive, specific, stable, and rapid biosensing in various practical applications.

Ultrafast and highly sensitive detection of SARS-CoV-2 spike protein by field-effect transistor graphene-based biosensors.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), etiological agent for the coronavirus disease 2019 (COVID-19), has resulted in over 775 million global infections. Early diagnosis remains pivotal for effective epidemiological surveillance despite the availability of vaccines. Antigen-based assays are advantageous for early COVID-19 detection due to their simplicity, cost-effectiveness, and suitability for point-of-care testing (PoCT). This study introduces a graphene field-effect transistor-based biosensor designed for high sensitivity and rapid response to the SARS-CoV-2 spike protein. By functionalizing graphene with monoclonal antibodies and applying short-duration gate voltage pulses, we achieve selective detection of the viral spike protein in human serum within 100 µs and at concentrations as low as 1 fg ml-1, equivalent to 8 antigen molecules perµl of blood. Furthermore, the biosensor estimates spike protein concentrations in serum from COVID-19 patients. Our platform demonstrates potential for next-generation PoCT antigen assays, promising fast and sensitive diagnostics for COVID-19 and other infectious diseases.

Revolutionizing food safety with electrochemical biosensors for rapid and portable pathogen detection.

Foodborne diseases remain a worldwide concern, despite the advances made in sanitation, pathogen surveillance and food safety management systems. The methods routinely applied for detecting pathogens in foods are time consuming, labor intensive and usually require trained and qualified individuals. The objective of this review was to highlight the use of biosensors, with a focus on the electrochemical devices, as promising alternatives for detecting foodborne pathogens. These biosensors present high speed for obtaining results, with the possibility of evaluating foods in real time, at low cost, ease of use, in addition to being compact and portable. These aspects are considered advantageous and suitable for use in food safety management systems. This work also shows some limitations for the application of biosensors, and we present perspectives with the development and use of nanomaterials.

Wearable, battery-free, wireless multiplexed printed sensors for heat stroke prevention with mussel-inspired bio-adhesive membranes.

Wearable technologies are becoming pervasive in our society, and their development continues to accelerate the untapped potential of continuous and ubiquitous sensing, coupled with big data analysis and interpretation, has only just begun to unfold. However, existing wearable devices are still bulky (mainly due to batteries and electronics) and have suboptimal skin contact. In this work, we propose a novel approach based on a sensor network produced through inkjet printing of nanofunctional inks onto a semipermeable substrate. This network enables real-time monitoring of critical physiological parameters, including temperature, humidity, and muscle contraction. Remarkably, our system operates under battery-free and wireless near-field communication (NFC) technology for data readout via smartphones. Moreover, two of the three sensors were integrated onto a naturally adhesive bioinspired membrane. This membrane, developed using an eco-friendly, high-throughput process, draws inspiration from the remarkable adhesive properties of mussel-inspired molecules. The resulting ultra-conformable membrane adheres effortlessly to the skin, ensuring reliable and continuous data collection. The urgency of effective monitoring systems cannot be overstated, especially in the context of rising heat stroke incidents attributed to climate change and high-risk occupations. Heat stroke manifests as elevated skin temperature, lack of sweating, and seizures. Swift intervention is crucial to prevent progression to coma or fatality. Therefore, our proposed system holds immense promise for the monitoring of these parameters on the field, benefiting both the general population and high-risk workers, such as firefighters.