Smartphone-based device for rapid and single-step detection of piRNA-651 using anti-DNA:RNA hybrid antibody and enzymatic signal amplification.

P-element-induced wimpy testis (PIWI)-interacting RNAs (piRNAs/piRs) are a class of small noncoding RNAs that play a crucial role in regulating various biological processes, including carcinogenesis. One specific piRNA, piR-651, has been reported to be overexpressed in both human blood serum and solid cancer tissues, that can be used a viable biomarker in cancer diagnosis. Early diagnosis of cancer can help reduce the burden of the disease and improve survival rates. In the present work, we report for the first time a smartphone-based colorimetric biosensor for highly sensitive and specific detection of piR-651 thanks to an enzymatic signal amplification, which yielded high colorimetric intensities. Indeed, a heteroduplex DNA:RNA was formed in the presence of piR-651 with the capture DNA probe immobilized on the magnetic beads for easy magnetic separation. Then, a HRP tethered to anti-DNA:RNA (S9.6) was used to reveal the DNA-RNA heteroduplex formed by catalyzing the oxidation of TMB substrate into colorimetric TMBox, which absorbs at 630 nm. The absorbance is positively proportional to the piR-651 concentrations. On the other hand, the colorimetric product of the assay can be photographed with a smartphone camera and analyzed using ImageJ software. Using a smartphone and under optimal conditions, the biosensor responded linearly to the logarithm of piRNA-651 from 8 fM to 100 pM with a detection limit of 2.3 fM and discriminates against other piRNAs. It was also successfully applied to the determination of piRNA-651 levels in spiked human serum.

Untreated vs. Treated Carbon Felt Anodes: Impacts on Power Generation in Microbial Fuel Cells.

This research sought to enhance the efficiency and biocompatibility of anodes in bioelectrochemical systems (BESs) such as microbial fuel cells (MFCs), with an aim toward large-scale, real-world applications. The study focused on the effects of acid-heat treatment and chemical modification of three-dimensional porous pristine carbon felt (CF) on power generation. Different treatments were applied to the pristine CF, including coating with carbon nanofibers (CNFs) dispersed using dodecylbenzene sulfonate (SDBS) surfactant and biopolymer chitosan (CS). These processes were expected to improve the hydrophilicity, reduce the internal resistance, and increase the electrochemically active surface area of CF anodes. A high-resolution scanning electron microscopy (HR-SEM) analysis confirmed successful CNF coating. An electrochemical analysis showed improved conductivity and charge transfer toward [Fe(CN)6]3-/4- redox probe with treated anodes. When used in an air cathode single-chamber MFC system, the untreated CF facilitated quicker electroactive biofilm growth and reached a maximum power output density of 3.4 W m-2, with an open-circuit potential of 550 mV. Despite a reduction in charge transfer resistance (Rct) with the treated CF anodes, the power densities remained unchanged. These results suggest that untreated CF anodes could be most promising for enhancing power output in BESs, offering a cost-effective solution for large-scale MFC applications.

A highly sensitive colorimetric DNA sensor for MicroRNA-155 detection: leveraging the peroxidase-like activity of copper nanoparticles in a double amplification strategy.

A novel and highly sensitive colorimetric DNA sensor for determination of miRNA-155 at attomolar levelsis presented that combines the peroxidase-like activity of copper nanoparticles (CuNPs) with the hybridization chain reaction (HCR) . The utilization of CuNPs offers advantages such as strong interaction with double-stranded DNA, excellent molecular recognition, and mimic catalytic activity. Herein, a capture probe DNA (P1) was immobilized on carboxylated magnetic beads (MBs), allowing for amplified immobilization due to the 3D surface. Subsequently, the presence of the target microRNA-155 led to the formation of a sandwich structure (P2/microRNA-155/P1/MBs) when P2 was introduced to the modified P1/MBs. The HCR reaction was then triggered by adding H1 and H2 to create a super sandwich (H1/H2)n. Following this, Cu2+ ions were attracted to the negatively charged phosphate groups of the (H1/H2)n and reduced by ascorbic acid, resulting in the formation of CuNPs, which were embedded into the grooves of the (H1/H2)n. The peroxidase-like activity of CuNPs catalyzed the oxidation reaction of 3,3',5,5'-Tetramethylbenzidine (TMB), resulting in a distinct blue color measured at 630 nm. Under optimal conditions, the colorimetric biosensor exhibited a linear response to microRNA-155 concentrations ranging from 80 to 500 aM, with a detection limit of 22 aM, and discriminate against other microRNAs. It was also successfully applied to the determination of microRNA-155 levels in spiked human serum.

Selective Sensing in Microbial Fuel Cell Biosensors: Insights from Toxicity-Adapted and Non-Adapted Biofilms for Pb(II) and Neomycin Sulfate Detection.

This study introduces the utilization of self-powered microbial fuel cell (MFC)-based biosensors for the detection of biotoxicity in wastewater. Current MFC-based biosensors lack specificity in distinguishing between different pollutants. To address this limitation, a novel approach is introduced, capitalizing on the adaptive capabilities of anodic biofilms. By acclimating these biofilms to specific pollutants, an enhancement in the selectivity of MFC biosensors is achieved. Notably, electrochemically active bacteria (EAB) were cultivated on 3D porous carbon felt with and without a model toxicant (target analyte), resulting in the development of toxicant-resistant anodic biofilms. The model toxicants, Pb2+ ions and the antibiotic neomycin sulfate (NS), were deployed at a concentration of 1 mg L-1 during MFC operation. The influence of toxicity on biofilm growth and power production was investigated through polarization and power density curves. Concurrently, the electrochemical activity of both non-adapted and toxicity-adapted biofilms was investigated using cyclic voltammetry. Upon maturation and attainment of peak powers, the MFC reactors were evaluated individually as self-powered biosensors for pollutant detection in fresh wastewater, employing the external resistor (ER) mode. The selected ER, corresponding to the maximum power output, was positioned between the cathode and anode of each MFC, enabling output signal tracking through a data logging system. Subsequent exposure of mature biofilm-based MFC biosensors to various concentrations of the targeted toxicants revealed that non-adapted mature biofilms generated similar current-time profiles for both toxicity models, whereas toxicity-adapted biofilms produced distinctive current-time profiles. Accordingly, these results suggested that merely by adapting the anodic biofilm to the targeted toxicity, distinct and identifiable current-time profiles can be created. Furthermore, these toxicity-adapted and non-adapted biofilms can be employed to selectively detect the pollutant via the differential measurement of electrical signals. This differentiation offers a promising avenue for selective pollutant detection. To the best of our current knowledge, this approach, which harnesses the natural adaptability of biofilms for enhanced sensor selectivity, represents a pioneering effort in the realm of MFC-based biosensing.

Fast impedimetric immunosensing of IgGs associated with peanut and hazelnut allergens.

Food allergies trigger a variety of clinical adverse symptoms and clinical evidence suggests that the presence of food allergy-related IgG can be helpful in the diagnosis when analyzed at the peptide-epitope level. To validate and select the peptides based on their specificity toward hazelnut or peanut epitopes, the authors of this study developed a silicon-based microchip coupled with click-chemistry bound peptides identified by the Fraunhofer Institute for Cell Therapy and Immunology. Peptides related to hazelnut and peanut allergies were identified and used to develop a silicon-based microchip. Peptides were coupled with click-chemistry to the sensor surface. The immunosensor was developed by electrografting diazotized amino phenylacetic acid and subsequently, dibenzocyclooctyne-amine (DBCO-NH2) was used as click-chemistry to allow coupling of the peptides with a C-terminal linker and azide structure. Energy-dispersive X-ray spectroscopy, electrochemical impedance spectroscopy (EIS), and fluorescence microscopy techniques have been used to analyze the bio-functionalization of the developed electrode. The peptide-epitope recognition was studied for seven allergen-derived peptides. The electrochemical responses were studied with sera from rabbits immunized with hazelnut and peanut powder. The microchips functionalized with the chosen peptides (peanut peptides T12 and EO13 and hazelnut peptides S4 and EO14 with an RSD of 4%, 3%, 9%, and 1% respectively) demonstrated their ability to specifically detect prevalent anti-nut related IgGs in rabbit sera in a range of dilutions from 1:500000 (0.0002%) until 1:50000 (0.002%). In addition, the other peptides showed promising differentiation abilities which can be further studied to perform multivariable detection fingerprint of anti-allergens in blood sera.

Development of an Innovative Colorimetric DNA Biosensor Based on Sugar Measurement.

The development of biosensors for target detection plays a crucial role in advancing various fields of bioscience. This work presents the development of a genosensor that exploits the colorimetric phenol-sulfuric acid sugar reaction for the detection of DNA, and RNA as specific targets, and DNA intercalator molecules. The biosensor combines simplicity and reliability to create a novel bioassay for accurate and rapid analysis. A 96-well microplate based on a polystyrene polymer was used as the platform for an unmodified capture DNA immobilization via a silanization process and with (3-Aminopropyl) triethoxysilane (APTES). After that, a hybridization step was carried out to catch the target molecule, followed by adding phenol and sulfuric acid to quantify the amount of DNA or RNA sugar backbone. This reaction generated a yellow-orange color on the wells measured at 490 nm, which was proportional to the target concentration. Under the optimum conditions, a calibration curve was obtained for each target. The developed biosensor demonstrated high sensitivity, good selectivity, and linear response over a wide concentration range for DNA and RNA targets. Additionally, the biosensor was successfully employed for the detection of DNA intercalator agents that inhibited the hybridization of DNA complementary to the immobilized capture DNA. The developed biosensor offers a potential tool for sensitive and selective detection in various applications, including virus diagnosis, genetic analysis, pathogenic bacteria monitoring, and drug discovery.

Development of a New Route for the Immobilization of Unmodified Single-Stranded DNA on Chitosan Beads and Detection of Released Guanine after Hydrolysis.

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.

Paper-Based Colorimetric Detection of miRNA-21 Using Pre-Activated Nylon Membrane and Peroxidase-Mimetic Activity of Cysteamine-Capped Gold Nanoparticles.

Irregular expression of MicroRNA-21 (miRNA-21) is considered as a promising biomarker for early cancer diagnosis. In this paper, a new genosensor based on paper and nanozyme activity of cysteamine-capped gold nanoparticles (Cys/AuNPs) was developed to detect picomolar concentrations of miRNA-21. Such nanozyme catalyzes the colorimetric reaction of hydrogen peroxide (H2O2) and 3,3',5,5' tetramethylbenzidine (TMB), to produce a blue color measurable by a smartphone. Due to their positive charge, Cys/AuNPs were attached to the negative phosphate groups of the DNA strand backbone via electrostatic interactions, leading to the quantitative determination of miRNA-21 concentration by the peroxidase-like activity of Cys/AuNPs. Furthermore, a paper-based assay was carried out on nylon disk devices to allow fast immobilization of DNAprobe. After performing the paper-based assay, a good linear range was observed between 1 pM and 1 nM (Y = 0.080 [MiRNA-21]/pM + 13.846, R2 = 0.993) with a detection limit of 0.5 pM. The developed method was effective, selective, and sensitive for the miRNA-21 detection. The application of the proposed method for miRNA-21 detection was examined in a human serum sample, and a recovery rate of 90.0-97.6% was obtained showing the acceptable accuracy of the developed approach.

Electrochemical DNA Biosensor Based on Immobilization of a Non-Modified ssDNA Using Phosphoramidate-Bonding Strategy and Pencil Graphite Electrode Modified with AuNPs/CB and Self-Assembled Cysteamine Monolayer.

The present paper describes an alternative approach to the traditionally used covalent immobilization methods that require cost-intensive and complicated chemistry modification of a single-stranded DNA (ssDNA) capture probe. The low-cost pencil graphite electrode (PGE) modified with carbon black (CB) and gold nanoparticles (AuNPs) was used as an electrochemical platform and the non-modified ssDNA was immobilized on a self-assembled cysteamine modified AuNPs/CB-PGE through a phosphoramidate bond between the 5'-terminal phosphate group of ssDNA and the primary amine group of cysteamine. The microRNA-21 was used as a target model in the fabrication of this electrochemical DNA biosensor and the hybridization process with the complementary probe was monitored by differential pulse voltammetry using methylene blue (MB) as an electrochemical hybridization indicator. The decreased reduction peak current of MB shows a good linear correlation with the increased concentration of microRNA-21 target sequences because the MB signal is determined by the amount of exposed guanine bases. The linear range of the fabricated DNA biosensor was from 1.0 × 10-8 to 5.0 × 10-7 M with a detection limit of 1.0 × 10-9 M. These results show that the covalent immobilization of a non-modified ssDNA capture probe through a phosphoramidate-bonding strategy could serve as a cost-effective and versatile approach for the fabrication of DNA biosensors related to a wide range of applications that cover the fields of medical diagnostic and environmental monitoring. The fabricated electrochemical DNA biosensor was used to analyze microRNA-21 in a (spiked) human serum sample and it showed satisfactory and encouraging results as an electrochemical DNA biosensor platform.

A membrane-less Glucose/O2 non-enzymatic fuel cell based on bimetallic Pd-Au nanostructure anode and air-breathing cathode: Towards micro-power applications at neutral pH.

Herein, the authors propose a miniaturized glucose/O2 n-EFC based on a new direct electron transfer. The anode is a screen-printed carbon electrode (SPCE) modified with functionalized carbon nanotubes (f-CNTs) and cauliflower-like PdAu nanostructures (PdAuNS). The PdAuNS/f-CNT biomimetic nanocatalyst was prepared using a cost-effective and straightforward method, which consisted of drop-casting well-dispersed f-CNTs over the SPCE surface before PdAuNS electrodeposition. This enzyme-free interface was used for glucose electrooxidation at neutral medium (pH 7.4). The electrochemical behaviour of the PdAuNS/f-CNT/SPCE was investigated using cyclic voltammetry, linear sweep voltammetry, and amperometry. Several parameters were optimized and discussed, including the metal precursor concentration (HAuCl4, PdCl2) and the electrodeposition conditions. The cathode for oxygen electroreduction is an air-cathode which is composed of Pt-coated carbon cloth. The electrochemical performances of the anode and the cathode were evaluated separately for glucose oxidation and oxygen reduction, respectively. Both electrodes were then assembled in a membrane-less single chamber n-EFC with an innovative architecture. Electrical characterization of the n-EFC supplied with a neutral buffered solution containing 20 mM glucose showed a maximal power output of 129 ± 11 µW cm-2, a current density of 600 ± 39 µA cm-2 with a cell voltage of 0.35 V, and an open circuit potential of 0.56 V. The proposed electrocatalyst possesses several advantages such as fast response, low cost, reusability, poison-free characteristics, and good stability. Hence, glucose/O2 n-EFC could be of great interest in direct glucose fuel cell applications (e.g., powering mountable/implantable biomedical micro-devices running at low electrical power supply) or in self-powered biosensing.

Nanoporous Cauliflower-like Pd-Loaded Functionalized Carbon Nanotubes as an Enzyme-Free Electrocatalyst for Glucose Sensing at Neutral pH: Mechanism Study.

In this work, we propose a novel functionalized carbon nanotube (f-CNT) supporting nanoporous cauliflower-like Pd nanostructures (PdNS) as an enzyme-free interface for glucose electrooxidation reaction (GOR) in a neutral medium (pH 7.4). The novelty resides in preparing the PdNS/f-CNT biomimetic nanocatalyst using a cost-effective and straightforward method, which consists of drop-casting well-dispersed f-CNTs over the Screen-printed carbon electrode (SPCE) surface, followed by the electrodeposition of PdNS. Several parameters affecting the morphology, structure, and catalytic properties toward the GOR of the PdNS catalyst, such as the PdCl2 precursor concentration and electrodeposition conditions, were investigated during this work. The electrochemical behavior of the PdNS/f-CNT/SPCE toward GOR was investigated through Cyclic Voltammetry (CV), Linear Sweep Voltammetry (LSV), and amperometry. There was also a good correlation between the morphology, structure, and electrocatalytic activity of the PdNS electrocatalyst. Furthermore, the LSV response and potential-pH diagram for the palladium-water system have enabled the proposal for a mechanism of this GOR. The proposed mechanism would be beneficial, as the basis, to achieve the highest catalytic activity by selecting the suitable potential range. Under the optimal conditions, the PdNS/f-CNT/SPCE-based biomimetic sensor presented a wide linear range (1-41 mM) with a sensitivity of 9.3 µA cm-2 mM-1 and a detection limit of 95 µM (S/N = 3) toward glucose at a detection potential of +300 mV vs. a saturated calomel electrode. Furthermore, because of the fascinating features such as fast response, low cost, reusability, and poison-free characteristics, the as-proposed electrocatalyst could be of great interest in both detection systems (glucose sensors) and direct glucose fuel cells.

Electrochemical Biosensors for Detection of MicroRNA as a Cancer Biomarker: Pros and Cons.

Cancer is the second most fatal disease in the world and an early diagnosis is important for a successful treatment. Thus, it is necessary to develop fast, sensitive, simple, and inexpensive analytical tools for cancer biomarker detection. MicroRNA (miRNA) is an RNA cancer biomarker where the expression level in body fluid is strongly correlated to cancer. Various biosensors involving the detection of miRNA for cancer diagnosis were developed. The present review offers a comprehensive overview of the recent developments in electrochemical biosensor for miRNA cancer marker detection from 2015 to 2020. The review focuses on the approaches to direct miRNA detection based on the electrochemical signal. It includes a RedOx-labeled probe with different designs, RedOx DNA-intercalating agents, various kinds of RedOx catalysts used to produce a signal response, and finally a free RedOx indicator. Furthermore, the advantages and drawbacks of these approaches are highlighted.

Recent Advances in Electrochemical Monitoring of Chromium.

The extensive use of chromium by several industries conducts to the discharge of an immense quantity of its various forms in the environment which affects drastically the ecological and biological lives especially in the case of hexavalent chromium. Electrochemical sensors and biosensors are useful devices for chromium determination. In the last five years, several sensors based on the modification of electrode surface by different nanomaterials (fluorine tin oxide, titanium dioxide, carbon nanomaterials, metallic nanoparticles and nanocomposite) and biosensors with different biorecognition elements (microbial fuel cell, bacteria, enzyme, DNA) were employed for chromium monitoring. Herein, recent advances related to the use of electrochemical approaches for measurement of trivalent and hexavalent chromium from 2015 to 2020 are reported. A discussion of both chromium species detections and speciation studies is provided.

Recent Advances in Electrochemical Biosensors Based on Enzyme Inhibition for Clinical and Pharmaceutical Applications.

A large number of enzyme inhibitors are used as drugs to treat several diseases such as gout, diabetes, AIDS, depression, Parkinson's and Alzheimer's diseases. Electrochemical biosensors based on enzyme inhibition are useful devices for an easy, fast and environment friendly monitoring of inhibitors like drugs. In the last decades, electrochemical biosensors have shown great potentials in the detection of different drugs like neostigmine, ketoconazole, donepezil, allopurinol and many others. They attracted increasing attention due to the advantage of being high sensitive and accurate analytical tools, able to reach low detection limits and the possibility to be performed on real samples. This review will spotlight the research conducted in the past 10 years (2007-2017) on inhibition based enzymatic electrochemical biosensors for the analysis of different drugs. New assays based on novel bio-devices will be debated. Moreover, the exploration of the recent graphical approach in diagnosis of reversible and irreversible inhibition mechanism will be discussed. The accurate and the fast diagnosis of inhibition type will help researchers in further drug design improvements and the identification of new molecules that will serve as new enzyme targets.

Electrochemical DNA sandwich biosensor based on enzyme amplified microRNA-21 detection and gold nanoparticles.

In this work, a novel electrochemical biosensor for miRNA-21 determination, involving a sandwich hybridization assay onto gold nanoparticles modified pencil graphite electrode (PGE) and enzyme signal amplification was reported. The thiol terminated capture probe 1 (SH-P1) was immobilized on the electrode through AuS interaction. In the presence of target miRNA-21, SH-P1 hybridized with the first part of the target, however, the second part hybridizes with a biotinylated probe P2 (B-P2). Then, a streptavidin-conjugated alkaline phosphatase was immobilized by a specific binding of avidin-B-P2. The enzyme catalyzed the electro-inactive α-naphtyl phosphate to an electro-active α-naphtol. The miRNA-21 detection was achieved through the changes of α-naphtol oxidation signals observed at +0.12V vs Ag/AgCl with Differential Pulse Voltammetry. Under the optimal detection conditions, the biosensor exhibited selective and sensitive detection with a linear range from 200pM to 388nM and the detection limit was 100pM (10fmol in 100µL).

Investigation of sulfonamides inhibition of carbonic anhydrase enzyme using multiphotometric and electrochemical techniques.

Two approaches for sulfonamides (SA's) determination, based on carbonic anhydrase enzyme inhibition have been investigated in this work. In the first method, nine different SA's have been screened using simultaneous multiphotometric measurement in multiwell plates. The sulfanilamide (SAD) showed significant inhibition compared to other sulfonamides. The carbonic anhydrase (CA) kinetic interactions reveal noncompetitive binding of SAD. Interferences from other inhibitors with enzyme were studied and the results showed very good selectivity toward SAD. In the second approach, an electrochemical enzyme inhibition biosensor, based on CA entrapped in a carbon paste electrode using carbon black nanoparticles and solid paraffin, was successfully applied to SAD measurements. Results from the quantitative analysis of SAD are discussed in terms of detection limit, linear range and sensitivity using multiphotometric and biosensor-based methods The biosensor developed was successfully applied to the determination of SAD at submicromolar levels and it is recommended for application for in situ analysis.

Enzyme inhibition-based biosensors for food safety and environmental monitoring.

Analytical technology based on sensors is an extremely broad field which impacts on many major industrial sectors such as the pharmaceutical, healthcare, food, and agriculture industries as well as environmental monitoring. This review will highlight the research carried out during the last 5 years on biosensors that are based on enzyme inhibition for determination of pollutants and toxic compounds in a wide range of samples. Here the different enzymes implicated in the inhibition, different transducers forming the sensing devices, and the different contaminants analyzed are considered. The general application of the various biosensors developed, with emphasis on food and environmental applications, is reviewed as well as the general approaches that have been used for enzyme immobilization, the enzyme catalysis, and the inhibition mechanism.

Determination of mercury(II) by invertase enzyme inhibition coupled with batch injection analysis.

The determination of mercury(II) ions at the trace level by inhibition of the invertase enzyme-catalysed hydrolysis of sucrose into glucose and fructose coupled to electrochemical batch injection analysis was investigated using two approaches. In the first, the glucose produced was detected by injection of 100 microliters samples into the batch injection cell containing a platinum electrode modified by immobilised glucose oxidase. In the second, the glucose and fructose present in injected samples were oxidised directly at a copper-modified glassy carbon electrode. The experimental parameters were optimised and the degree of enzyme inhibition by mercury(II) ions under both conditions was measured. Mercury concentrations in the ng ml-1 range were determined by these two techniques with low sample and reagent consumption. Comparison is made between the two methods and perspectives as a screening test for field application are indicated.