Modeling approach for Ti3C2 MXene-based fluorescent aptasensor for amoxicillin biosensing in water matrices.

Amoxicillin, a member of the penicillin family, is primarily utilized for the treatment of various bacterial infections affecting ears, nose, throat, urinary tract, and skin. Given its widespread application in medicine, agriculture, environment, and food industry, the precise and sensitive detection of amoxicillin is important. This study introduces a novel approach to developing a sensitive and selective fluorescent aptasensor relying on fluorescence resonance energy transfer (FRET) for the specific detection of amoxicillin. The carboxyfluorescein-labeled aptamer serves as a energy donor, while MXene functions as an energy acceptor, and acting as a quencher. To achieve optimal detection efficiency, a dual optimization strategy utilizing RSM-CCD and ANN-GA was used to fine-tune experimental conditions. The fluorescence measurements revealed an expansive linear range extending from 100 to 2400 ng mL-1, accompanied by an exceptionally low detection limit of 1.53 ng mL-1. Additionally, it shows an excellent selectivity towards amoxicillin over other antibiotics commonly found in water matrices. The aptasensor demonstrates good stability and reproducibility; effectiveness of the aptasensor was validated by testing in real water samples. This remarkable sensitivity and broad dynamic range affirm the efficacy aptasensor in accurately detecting varying concentrations of amoxicillin in wastewater bodies.

Design of a label-free aptasensor for electrochemical determination of hemoglobin: investigation of the peroxidase-like activity of hemoglobin for the sensing of different substrates.

The unbalanced hemoglobin level in biological fluids can cause several diseases; hence it can be used as a biomarker for diagnosis. We aim, in the present study, to construct a label-free electrochemical aptasensor for the quantification of hemoglobin. For that, a conjugate of L-cysteine and gold nanoparticles was used for the aptamer immobilization on screen printed carbon electrodes. Using square wave voltammetry, the calibration plot was obtained and it was linear in the range of 50 ng ml-1 to 36 000 ng ml-1 while the detection limit was 1.2 ng ml-1. After the binding of Hb on the modified screen-printed carbon electrode surface, the peroxidase-like activity of the bound hemoglobin was explored in the quantification of different substrates. Hydrogen peroxide and nitrite were chosen as model analytes. Amperometric measurements showed wide linear ranges: 0.2 µM-7.7 mM and 3.6 nM-1.3 mM for H2O2 and nitrite, respectively, with detection limits of 0.044 µM and 0.55 nM. In the proposed strategy, the aptamer provides excellent orientation and a biocompatible environment for hemoglobin whose catalytic activity plays a key role in H2O2 and nitrite analysis.

Acetylcholinesterase- and Butyrylcholinesterase-Based Biosensors for the Detection of Quaternary Ammonium Biocides in Food Industry.

A sensitive and robust electrochemical cholinesterase-based sensor was developed to detect the quaternary ammonium (QAs) biocides most frequently found in agri-food industry wash waters: benzalkonium chloride (BAC) and didecyldimethylammonium chloride (DDAC). To reach the maximum residue limit of 28 nM imposed by the European Union (EU), two types of cholinesterases were tested, acetylcholinesterase (AChE, from Drosophila melanogaster) and butyrylcholinesterase (BChE, from horse serum). The sensors were designed by entrapping AChE or BChE on cobalt phthalocyanine-modified screen-printed carbon electrodes. The limits of detection (LOD) of the resulting biosensors were 38 nM for DDAC and 320 nM for BAC, using, respectively, AChE and BChE. A simple solid-phase extraction step was used to concentrate the samples before biosensor analysis, allowing for the accurate determination of DDAC and BAC in tap water with limits of quantification (LOQ) as low as 2.7 nM and 5.3 nM, respectively. Additional assays demonstrated that the use of a phosphotriesterase enzyme allows for the total removal of interferences due to the possible presence of organophosphate insecticides in the sample. The developed biosensors were shown to be stable during 3 months storage at 4 °C.

Screen-printed electrochemical immunosensor based on a novel nanobody for analyzing aflatoxin M1 in milk.

This study aimed to devise a nontoxic electrochemical immunosensor to quantitatively determine aflatoxin M1 by chronoamperometry with novel anti-idiotypic nanobody-functionalized screen-printed carbon electrodes (SPCEs). Anti-idiotype nanobodies (AIdnb) were developed to replace the high toxic chemically synthesized antigen. AIdnb was immobilized on the surface of SPCE via covalent coupling as capture reagent. The functionalized SPCEs were followed by characterization using electrochemical impedance spectroscopy, fourier-transform infrared spectroscopy, transmission electron microscopy mapping, and atomic force microscopy. After optimizing experimental parameters, the assembled immunosensor exhibited a good linearity range of 0.25-5.0 ng/mL, with the limit of detection of 0.09 ng/mL. The immunosensor showed a satisfactory selectivity to AFM1, without interference from analogs, including zearalenone, ochratoxin, and fumonisin B1. For practical application, the developed immunosensor was validated using real spiked samples with the recovery range 82.0%-108.0% and relative standard deviation (RSD) 10.1%-13.0%, indicating that it could be used in milk samples.

A Sensitive Aptasensor Using Biotin-Streptavidin System for Patulin Detection in Apple Juice.

Patulin contamination in fruits, vegetables, and their products is considered a serious health risk factor for food safety and human health. Thus, a rapid, simple detection method for patulin is becoming important, which could provide a tool for routine screening and food surveys. The objective of this study was to develop a sensitive aptamer-based lateral flow assay (FLA) using Streptavidin functionalized gold nanoparticles for sensitive patulin detection. An excellent dynamic range for patulin detection was obtained (2.7~139.8 ng/mL in the buffer and 7.07~359.5 ng/mL in the sample) with no affinity for other mycotoxins such as zearalenone (ZEN), ochratoxin A (OTA), aflatoxin B1 (AFB1), citrinin or tenuazonic acid (TEA). The limit of detection was 0.19 ng/mL in the buffer and 0.36 ng/mL in the real sample. The recoveries were 83.3% to 107.1%, with a satisfactory RSD value from 6.5% to 7.5%. Hence the established LFA could be used as a rapid, simple, on-site screening tool for PAT determination in apple juice.

A Simple Fluorescent Aptasensing Platform Based on Graphene Oxide for Dopamine Determination.

Dopamine (DA) is a catecholamine neurotransmitter playing an important role in different biological functions including central nervous, renal, cardiovascular, and hormonal systems. The sensitive and selective detection of this neurotransmitter plays a key role in the early diagnosis of various diseases related to abnormal levels of dopamine. Therefore, it is of great importance to explore rapid, simple, and accurate methods for detection of dopamine with high sensitivity and specificity. We propose in this work a fluorescent aptasensor based on graphene oxide (GO) as a quencher, for the rapid determination of dopamine. The principle of this aptasensor is based on fluorescence resonance energy transfer (FRET), where GO was used as energy donor, and a carboxy fluorescein (FAM)-labeled aptamer as acceptor. In the absence of DA, FAM-aptamer was adsorbed on the surface of GO through π-π stacking interactions between nucleotide bases and the carbon network, leading to a weak FRET and a quenching of the FAM fluorescence. However, by adding the target, the aptamer undergoes a conformational change to bind to DA with high affinity, resulting in a fluorescence recovery. Under the optimal experimental conditions, the fluorescence recovery was linearly proportional to the concentration of DA in the range of 3-1680 nM, with a limit of detection of 0.031 nM and a limit of quantification of 0.1 nM. Moreover, the developed assay exhibited minor response in the presence of various interferents and it revealed a satisfactory applicability in human serum samples.

Development of a label-free electrochemical aptasensor based on diazonium electrodeposition: Application to cadmium detection in water.

In this study we have developed a new aptasensor for cadmium (Cd2+) detection in water. Gold electrode surface has been chemically modified by electrochemical reduction of diazonium salt (CMA) with carboxylic acid outward from the surface. This was used for amino-modified cadmium aptamer immobilization through carbodiimide reaction. Chemical surface modification was characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). This latter was also used for Cd2+ detection. The aptasensor has exhibited a good linear relationship between the logarithm of the Cd2+ concentration and the impedance changes in the range from 10-3 to 10-9 M with a correlation R2 of 0.9954. A high sensitivity was obtained with a low limit of detection (LOD) of 2.75*10-10 M. Moreover, the developed aptasensor showed a high selectivity towards Cd2+ when compared to other interferences such as Hg2+, Pb2+ and Zn2+. The developed aptasensor presents a simple and sensitive approach for Cd2+detection in aqueous solutions with application for trace Cd2+ detection in spring water samples.

Fabrication of AuNPs/MWCNTS/Chitosan Nanocomposite for the Electrochemical Aptasensing of Cadmium in Water.

Cadmium (Cd2+) is one of the most toxic heavy metals causing serious health problems; thus, designing accurate analytical methods for monitoring such pollutants is highly urgent. Herein, we report a label-free electrochemical aptasensor for cadmium detection in water. For this, a nanocomposite combining the advantages of gold nanoparticles (AuNPs), carbon nanotubes (CNTs) and chitosan (Cs) was constructed and used as immobilization support for the cadmium aptamer. First, the surface of a glassy carbon electrode (GCE) was modified with CNTs-CS. Then, AuNPs were deposited on CNTs-CS/GCE using chrono-amperometry. Finally, the immobilization of the amino-modified Cd-aptamer was achieved via glutaraldehyde cross-linking. The different synthesis steps of the AuNPs/CNTs/CS nano assembly were characterized by cyclic voltammetry (CV). Electrochemical impedance spectroscopy (EIS) was employed for cadmium determination. The proposed biosensor exhibited excellent performances for cadmium detection at a low applied potential (-0.5 V) with a high sensitivity (1.2 KΩ·M-1), a detection limit of 0.02 pM and a wide linear range (10-13-10-4 M). Moreover, the aptasensor showed a good selectivity against the interfering ions: Pb2+; Hg2+ and Zn2+. Our electrochemical biosensor provides a simple and sensitive approach for Cd2+ detection in aqueous solutions, with promising applications in the monitoring of trace amounts of heavy metals in real samples.

Investigation of a Truncated Aptamer for Ofloxacin Detection Using a Rapid FRET-Based Apta-Assay.

In this work, we describe the use of a new truncated aptamer for the determination of ofloxacin (OFL), being a principal quinolone commonly used in both human and animal healthcare. Since the affinity of a 72-mer ssDNA sequence has been previously described without further investigations, this paper demonstrates the first computational prediction of the binding motif between this aptamer and OFL through in silico molecular docking studies. Besides, we suggest the application of the characterized recognition mechanism in a simple FRET (Förster Resonance Energy Transfer) pattern for the rapid aptasensing of the quinolone of interest. Accordingly, our approach harnesses the fluorescence quenching of the fluorescein-tagged aptamer (FAM-APT) induced by its partial hybridization to a tetramethyl rhodamine-labelled complementary ssDNA (TAMRA-cDNA). In such a structure, dye labels brought into close proximity act as a FRET pair. Upon ofloxacin addition, an affinity competition occurs to form a more stable FAM-APT/OFL complex, thus unquenching the FAM-APT signal. Interestingly, the recovered fluorescence intensity was found to correlate well with the antibiotic's concentrations in the range of 0.2-200 µM in HEPES buffer, with a linear response that ranged between 0.2 and 20 µM. The rapid apta-assay achieved limits of detection and quantification of 0.12 and 0.40 µM, respectively. The truncated aptamer has also shown an improved specificity toward OFL than other quinolones, compared to the original full-length aptamer described in previous works. Finally, the practical application of the developed apta-assay was successfully confirmed to detect OFL quinolone in spiked milk samples, with satisfactory recoveries ranging between 97.4% and 111.4%.

Switchable fluorescence sensor toward PAT via CA-MWCNTs quenched aptamer-tagged carboxyfluorescein.

A quenching based apta-sensing platform was developed for the detection of Patulin. Three different aptamer sequences were studied to screen the aptamer with the maximum affinity towards Patulin. Carboxyfluorescein (CFL) was used as a fluorescent dye while -COOH functionalized multiwall carbon nanotubes (MWCNTs) were applied as novel nanoquenchers. Aptamer tagged at the 3' end with 40 nucleotide bases exhibited the maximum affinity towards Patulin and caused substantial fluorescence recovery. Interestingly, the limit of detection (LOD) and limit of quantification (LOQ) were calculated as 0.13 µg L-1and 0.41 µg L-1 respectively. Commonly occurring mycotoxins in food were also tested to confirm the selectivity of apta-assay. The developed apta-assay was applied to a spiked apple juice sample and toxin recoveries were observed ranging from 96% to 98% (n = 3). These results demonstrated the potential of the developed apta-assay for the selective detection and quantification of Patulin in food samples.

Polymer scaffold layers of screen-printed electrodes for homogeneous deposition of silver nanoparticles: application to the amperometric detection of hydrogen peroxide.

A method is described for electrochemical oxidation of polymers on the surface of screen-printed electrodes (SPCE). These act as scaffold layers for homogeneous deposition of silver nanoparticles (AgNPs). Hexamethylenediamine (HMDA) and poly(ethylene glycol) were immobilized on the SPCE surface via electrochemical oxidation. AgNPs were then electrodeposited on the scaffolds on the SPCE. This type of different carbon chain containing materials like PEG and HMDA act as big tunnels for electron mobility and are useful for the homogenous deposition of AgNPs on the SPCE surface without agglomeration. The resulting sensor was applied to the determination of hydrogen peroxide (H2O2) as a model analyte. It is found to display favorable catalytic and conductive properties towards the reduction of H2O2. Cyclic voltammetry and amperometry revealed that the modified electrode performs better than other modified SPCEs. Best operated at a potential of around -0.61 V (vs Ag|AgCl), the amperometric response is linear in the 10-180 µM H2O2 concentration range and the detection limit is 1.5 µM. The sensor is stable and reproducible. The resultant sensor was appplied to toothpaste analysis, and good recovery values were gained. Graphical abstractSchematic representation of electropolymerization of poly(ethylene glycol) and hexamethylenediamine scaffold layers on screen-printed electrodes for homogeneous electrodeposition of silver nanoparticles. This electrode was applied for the amperometric determination of hydrogen peroxide.

Highly sensitive label-free in vitro detection of aflatoxin B1 in an aptamer assay using optical planar waveguide operating as a polarization interferometer.

This work reports on further development of an optical biosensor for the in vitro detection of mycotoxins (in particular, aflatoxin B1) using a highly sensitive planar waveguide transducer in combination with a highly specific aptamer bioreceptor. This sensor is built on a SiO2-Si3N4-SiO2 optical planar waveguide (OPW) operating as a polarization interferometer (PI), which detects a phase shift between p- and s-components of polarized light propagating through the waveguide caused by the molecular adsorption. The refractive index sensitivity (RIS) of the recently upgraded PI experimental setup has been improved and reached values of around 9600 rad per refractive index unity (RIU), the highest RIS values reported, which enables the detection of low molecular weight analytes such as mycotoxins in very low concentrations. The biosensing tests yielded remarkable results for the detection of aflatoxin B1 in a wide range of concentrations from 1 pg/mL to 1 µg/mL in direct assay with specific DNA-based aptamers. Graphical abstract Optical planar waveguide polarization interferometry biosensor for detection of aflatoxin B1 using specific aptamer.

Development of a highly sensitive xanthine oxidase-based biosensor for the determination of antioxidant capacity in Amazonian fruit samples.

The paper describes the development of an amperometric biosensor using Prussian Blue (PB) modified electrodes containing xanthine oxidase (XOD). The enzyme is immobilized by photo-polymerization into an azide-unit pendant water-soluble photopolymer (PVA-AWP). The parameters of the fabrication of the biosensor, XOD:PVA/AWP ratio, crosslinking irradiation time, and XOD charge, were optimized. Operational conditions for electrode preparation were defined as 1:2 ratio of XOD:PVA/AWP; exposure time to neon light of 30  min; pH = 7.5  at room temperature and enzymatic charge of 8 mU per electrode. The biosensors showed stable, fast, simple, selective, cost-effective and sensitive (-2.72E-8  A mol L-1), with a good linear range (1.0-75  µmol L-1), and respectively detection and quantification limits for antioxidants of 2.17, and 7.15  µmol L-1. The applicability of this biosensor was demonstrated by in vitro analysis of gallic acid as standard antioxidant and Amazonian fruits as natural sources.

Ultrasensitive ciprofloxacin assay based on the use of a fluorescently labeled aptamer and a nanocomposite prepared from carbon nanotubes and MoSe2.

A nanocomposite was prepared from carbon nanotubes and MoSe2 (CNT-MoSe2). This nanomaterial quenches the fluorescence of fluorescein-labeled aptamers. When ciprofloxacin (CIP) binds to the aptamer, an aptamer/G-quadruplex complex will be formed and the interaction between labeled aptamer and CNT-MoSe2 nanostructures is weakened. This leads to significant fluorescence recovery. Under optimized experimental conditions, the limit of detection is 0.63 ng mL-1 with a good linearity in the range from 0.63 to 80 ng mL-1. The assay was applied to the determination of CIP in spiked milk, and the recoveries range between 94.3 and 97.0% (n = 3). Conceivably, the method is a generic approach that can be extended to the determination of other analyte for which adequate aptamers are available. Graphical abstract Schematic presentation of CNT-MoSe2 quenching based aptamer assay for the detection of ciprofloxacin. The assay exhibits good selectivity, stability and reproducibility, and low limit of detection.

Development of an Impedimetric Aptasensor for Label Free Detection of Patulin in Apple Juice.

In the present work, an aptasensing platform was developed for the detection of a carcinogenic mycotoxin termed patulin (PAT) using a label-free approach. The detection was mainly based on a specific interaction of an aptamer immobilized on carbon-based electrode. A long linear spacer of carboxy-amine polyethylene glycol chain (PEG) was chemically grafted on screen-printed carbon electrodes (SPCEs) via diazonium salt in the aptasensor design. The NH2-modified aptamer was then attached covalently to carboxylic acid groups of previously immobilized bifunctional PEG to build a diblock macromolecule. The immobilized diblocked molecules resulted in the formation of long tunnels on a carbon interface, while the aptamer was assumed as the gate of these tunnels. Upon target analyte binding, the gates were assumed to be closed due to conformational changes in the structure of the aptamer, increasing the resistance to the charge transfer. This increase in resistance was measured by electrochemical impedance spectroscopy, the main analytical technique for the quantitative detection of PAT. Encouragingly, a good linear range between 1 and 25 ng was obtained. The limit of detection and limit of quantification was 2.8 ng L-1 and 4.0 ng L-1, respectively. Selectivity of the aptasensor was confirmed with mycotoxins commonly occurring in food. The developed apta-assay was also applied to a real sample, i.e., fresh apple juice spiked with PAT, and toxin recovery up to 99% was observed. The results obtained validated the suitability and selectivity of the developed apta-assay for the identification and quantification of PAT in real food samples.

Design of a redox-active surface for ultrasensitive redox capacitive aptasensing of aflatoxin M1 in milk.

Herein, we report the design of a novel label-free aptasensor based on ferrocene and silicon nanoparticles (SiNPs) for ultrasensitive detection of aflatoxin M1 (AFM1) in milk. Given that silicon nanomaterials stand out by their high capacitive power, we used them to develop a novel capacitive transduction system based on electrochemical capacitance spectroscopy (ECS). This strategy relies on the changes of the redox capacitance signal owed to the surface-tethered ferrocene film, by performing electrochemical impedance spectroscopy (EIS) measurements without using an external redox probe. The redox capacitance variation was found to correlate well with the increasing concentrations of AFM1 in the linear range from 10 to 500 fmol⋅L-1 with a sensitivity of 0.46 µF*fM-1*cm - 2. Furthermore, the aptasensor allowed to reach very low limits of detection and quantification equal to 4.53 fM and 14.95 fM, respectively. The platform revealed a high selectivity toward the target analyte, and it was applied to quantify very low concentrations of AFM1 in commercial pasteurized milk. Finally, the results of real sample analysis were successfully gauged against those obtained using commercially available enzyme-linked immunoassay (ELISA) kits.

Label-Free Optical Detection of Mycotoxins Using Specific Aptamers Immobilized on Gold Nanostructures.

This work focuses on the development of the novel label-free optical apta-sensors for detection of mycotoxins. A highly sensitive analytical method of total internal reflection ellipsometry (TIRE) combined with Localized Surface Plasmon Resonance (LSPR) phenomenon in nano-structured gold films was exploited here for the first time for detection of aflatoxin B1 and M1 in direct assay with specific aptamers immobilized on the surface of gold. The achieved detection of low molecular weight molecules, such as aflatoxin B1 and M1, in a wide range of concentrations from 100 ng/mL down to 0.01 ng/mL is remarkable for the LSPR method. The study of binding kinetics of aflatoxin molecules to their respective aptamers using dynamic TIRE measurements yielded the values of affinity constants in the range of 10-8⁻10-7 mol, which is characteristic for highly specific aptamer/target interactions similar to that for monoclonal antibodies. The effect of aptamers' DNA chain length on their binding characteristics was analyzed.

Development of a portable and disposable NS1 based electrochemical immunosensor for early diagnosis of dengue virus.

The present study represents fabrication of nonstructural antibody (NS1) based immunosensor coupled with bovine serum albumin (BSA) modified screen printed carbon electrodes (SPCE) as transducing substrate for the early diagnosis of dengue virus. The anti-NS1 monoclonal antibody was immobilized on electro grafted BSA surface of working electrode. The electrons transfer resistance before and after NS1 attachment was monitored as a function of its concentration to perform the qualitative and quantitative analysis. The as prepared impedimetric immunosensor successfully detected the dengue virus protein with enhanced limit of detection (0.3 ng/mL) and linear range (1-200 ng/mL). The selectivity of the designed device was further elaborated with several interfering analytes and was finally demonstrated with human serum samples. The extravagant selectivity, sensitivity and easier fabrication protocol corroborate the potential applications of such immunosensor for practical diagnosis of dengue virus.

Designed Strategies for Fluorescence-Based Biosensors for the Detection of Mycotoxins.

Small molecule toxins such as mycotoxins with low molecular weight are the most widely studied biological toxins. These biological toxins are responsible for food poisoning and have the potential to be used as biological warfare agents at the toxic dose. Due to the poisonous nature of mycotoxins, effective analysis techniques for quantifying their toxicity are indispensable. In this context, biosensors have been emerged as a powerful tool to monitors toxins at extremely low level. Recently, biosensors based on fluorescence detection have attained special interest with the incorporation of nanomaterials. This review paper will focus on the development of fluorescence-based biosensors for mycotoxin detection, with particular emphasis on their design as well as properties such as sensitivity and specificity. A number of these fluorescent biosensors have shown promising results in food samples for the detection of mycotoxins, suggesting their future potential for food applications.

An Overview on Recent Progress in Electrochemical Biosensors for Antimicrobial Drug Residues in Animal-Derived Food.

Anti-microbial drugs are widely employed for the treatment and cure of diseases in animals, promotion of animal growth, and feed efficiency. However, the scientific literature has indicated the possible presence of antimicrobial drug residues in animal-derived food, making it one of the key public concerns for food safety. Therefore, it is highly desirable to design fast and accurate methodologies to monitor antimicrobial drug residues in animal-derived food. Legislation is in place in many countries to ensure antimicrobial drug residue quantities are less than the maximum residue limits (MRL) defined on the basis of food safety. In this context, the recent years have witnessed a special interest in the field of electrochemical biosensors for food safety, based on their unique analytical features. This review article is focused on the recent progress in the domain of electrochemical biosensors to monitor antimicrobial drug residues in animal-derived food.