We report on the fabrication and characterisation of Graphene field-effect transistor (GFET) Biosensors for detecting clusterin, a prominent protein biomarker of Alzheimer’s disease (AD). There are approximately 54 million people currently living with dementia worldwide and this is expected to rise to 130 million by 2050. Although there are over 400 different types of dementia, AD is the most common type, affecting between 50–75% of those diagnosed with dementia. Diagnosis of AD can take up to 2 years currently using MRI, PET, CT scans and memory tests. There is, therefore, an urgent need to develop low-cost, accurate, non-invasive and point-of-care (PoC) sensors for early diagnosis of AD. The GFET sensors we are developing to address this challenge were fabricated on Si/SiO₂ substrate through processes of photolithographic patterning and metal lift-off techniques with evaporated chromium and sputtered gold contacts. Raman Spectroscopy was performed on the devices to determine the quality of the graphene. The GFETs were annealed to improve their performance before the channels were functionalized by immobilising the graphene surface with a linker molecule and anti-clusterin antibody. The detection was achieved through the binding reaction between the antibody and varying concentrations of clusterin antigen from 1 pg/mL to 1 ng/mL. The GFETs were characterized using 4-probe direct current (DC) electrical measurements which demonstrated a limit of detection of the biosensors to be below 1 pg/mL. Full article

Lactate is a metabolite biomarker of tissue oxygenation and it can be used in medicine to evaluate a pathology or in sport activities to evaluate physical performance. Lactate level assessment is also important for food industry. This acid is found in food and beverages and the concentration level can be correlated with the freshness, stability and quality of several products. In this work, we present a smartphone-based enzymatic biosensor utilizing the unique colorimetric properties of the poly(aniline-co-anthranilic acid) (p(ANI-co-AA)) composite film coupled with lactate oxidase–horseradish peroxidase (LOx–HRP) enzymes. The enzymes are immobilized on the composite polymer film by adsorption and they catalyze a reversible redox color change of the host polymer from green to blue in the presence of l-lactate as the substrate. A smartphone was applied as color detector, for image acquisition and data handling. The free-of-charge Color Grab® application for Android OS was used to enable an easy and clear display of the sensor’s response, indicating remarkable changes in the optical features. The results were confirmed by spectrophotometric measurements. The developed colorimetric enzymatic biosensors were studied and optimized in relation to different experimental parameters. Moreover, the colorimetric enzymatic biosensor was applied to real matrices analysis. It has been shown by these studies that the colorimetric biosensors are promising as quick and simple tests for handheld analysis in various fields. Full article

The outstanding properties of metal nanoclusters, stabilized with different scaffolds, i.e., proteins, nucleic acids, polymers and dendrimers, enable their application in a wide range of fields. The recent advances in the fabrication and synthesis of nanoclusters have revolutionized the design of biosensors, leading to significant improvements in the selective and sensitive determination of several targets. In particular, in recent years, copper nanoclusters (CuNCs) have attracted more attention mainly for their unique fluorescent properties, as well as their large Stokes shifts, low toxicity, and high biocompatibility. The high-photoluminescent features of CuNCs facilitate highly sensitive target detection even in complex biological matrices. For these reasons, in this work, we exploited the specific template-targeted CuNCs’ growth for the sensitive and accurate determination of human serum albumin (HSA) in urine and human serum. HSA is the most abundant protein in plasma, acting as a carrier for many key biological molecules such as hormones, fatty acids and steroids, and it contributes to the maintenance of the oncotic blood pressure. The concentration of HSA in body fluids greatly influences the state of health of the patients. Taking into account these considerations, the quantitative detection of human serum albumin plays a key role in the early diagnosis of serious pathological conditions such as albuminuria and albuminemia. Here, we present a CuNCs-based assay in which copper nanoclusters were used as fluorescent signal indicators to detect serum albumin in a complex biological matrix. Full article

This article demonstrates a model of a near-field sensor, which is a combined slot antenna based on a flexible substrate RO3003. It is shown that the sensor has a high penetration of electromagnetic waves into highly absorbing media due to the length of the near field. In addition, it has a small size (diameter the sensor is 25 mm, thickness 0.76 mm). The simplified model of a hand simplified with a shallow vein depth was constructed. This model based on the experimentally obtained data of dielectric constant for glucose concentrations of 1.2–10 mmol/l. The simulation results, carried out in the frequency range 0.5–5 GHz, showed the possibility of determining a small change in blood sugar level from the reflected signal data. Full article

So far, in some of our previous works, we have managed to rapidly (within minutes) identify and discriminate pathogens by using surface-enhanced Raman scattering (SERS) spectroscopy with a single cell sensitivity. Having a more user friendly and robust system, which could be used not only by experts, would be the next step. In order to meet our goal, we developed an experimental setup, including an in-house built microfluidic device and we optimized the SERS detection of common bacterial pathogens by using the developed device. The main components of the system are a microfluidic flow-cell coupled to a syringe pump mediated flow system and a portable Raman spectrometer for detecting the bacteria immobilized in the flow cell. Inside the microfluidic channel of the flow cell, a silver spot was generated under laser irradiation for further use as SERS active substrate for detection. The silver spot can be washed and reused for a different pathogen from one experiment to another. No specific capturing receptors are used. The total analysis time was reduced to less than 15 min. Considering the fit-for-purposes experimental parameters for detection and its easy-to-use dedicated software, this portable microfluidic device has been tested in our lab and is ready to be transferred in the research/clinical premises for further use. Full article

Silver nanoclusters (AgNCs) generated on DNA templates belong to a new class of fluorescent tags showing excellent brightness, photostability and biocompatibility. Thus, AgNCs-DNA has been applied in various applications, from the detection of DNA/RNA and environmental monitoring to bioimaging and cancer therapy. In this work, we report fluorescent AgNCs synthesized using two 1,3-diaza-2-oxophenothiazine (tC)-modified oligonucleotides that contain RET-related sequence CCCCGCCCCGCCCCGCCCCA. The communication compares the absorption and emission properties of the obtained systems with silver nanoclusters synthesized on the unmodified oligonucleotide. First, we showed the optimal conditions for AgNCs-DNA synthesis on three DNA templates: (1) RET20 with the sequence 5′-CCC CGC CCC GCC CCG CCC CA-3′; (2) RET19tC with the sequence 5′-CCC CGC CCC GCC CCG CCC tCA-3′; and (3) RET14tC with the sequence 5′-CCC CGC CCC GCC CtCG CCC CA-3′. Next, the silver nanoclusters were characterized by UV/Vis absorption, fluorescence and circular dichroism spectroscopy. Silver nanoclusters RET19tC-AgNCs and RET14tC-AgNCs indicated several times higher fluorescence intensities in the long-wave emission spectra as compared to RET20-AgNCs. Moreover, silver nanoclusters on tC-modified oligonucleotides showed higher stability over time. The possibility of using the silver nanoclusters RET19tC-AgNCs for monitoring pH changes will be also tested. Full article

Mono-and multi-metal nanoparticles (MNPs), thanks to their unique and tunable features, still fascinate the analytical sciences, from their widespread use in sensing and biosensing as nanoplasmonic tags or catalysts up to MNPs-decorated surfaces. Here, a lab µ-Tip decorated with plasmonic-active polymeric films embodying gold/silver nanostructures is presented. The proposed lab-on-a-tip device speed-up the 4-nitrophenol conversion in 4-aminophenol, retaining the performances for more than 10 consecutive measures, acting as an enzyme-like catalyst. Full article

Oncological diseases belong to the most serious illnesses with high mortality. The most common cancer in children is acute lymphoblastic leukemia (ALL). It is important to develop diagnostic methods that will be able to detect this disease in early stage. One of the possible options can be non-invasive diagnostics using the biosensors based on nucleic acid aptamers. Aptamer recognizes the surface markers on the membrane of cancer cells with the high binding affinity. Biosensors based on aptamers with redox markers are among the most sensitive experimental tools of this type. We developed and optimized the redox-labeled electrochemical aptasensors for the detection of Jurkat leukemia cells. The aptamers specific to the protein tyrosine kinase 7 (PTK7), which is important membrane protein cancer marker that is overexpressed in Jurkat cells were used. We compared the sensitivity of aptasensors for aptamers modified either by methylene blue (MB) and ferrocene carboxylic acid (Fc), respectively. Both aptasensors were tested in the presence of Jurkat cells at concentration range 50–5000 cells/mL using differential pulse voltammetry. In both cases the comparable sensitivity was obtained with limit of detection: 37 ± 6 cells/mL for Fc-labeled aptamers and 38 ± 8 cells/mL for MB-labeled aptamers based on 3.3S/N (signal to noise) rule. The interaction of the sensing surface with control U266 cells was less significant. Full article

The determination of protease activity is very important for disease diagnosis, drug development, and quality and safety assurance for dairy products. Therefore, the development of low-cost methods for assessing protease activity is critical. Here, we demonstrate that an acoustic wave-based biosensor operated in the thickness-shear mode (TSM) enables the low-cost detection of protease activity in real time. The TSM sensor was based on a protein substrate (PS) β-casein immobilized on a piezoelectric quartz crystal electrode. The β-casein layer was immobilized onto a gold surface by a carboxylate terminated self-assembled monolayer (SAM) of 11-mercaptoundecanoic acid (MUA). The carboxylic acid terminal was activated by the reaction of a mixture of water- soluble N-(3-Dimethylaminopropyl)-N0-ethylcarbodiimide (EDC) and N–Hydroxysuccinimide (NHS) on the electrode surfaces. We demonstrated that β-casein can form a stable assembly on a piezoelectric quartz crystal electrode. After an enzymatic reaction with trypsin, it cleaved the surface-bound β-casein substrate, which increased the frequency of the crystal in a sigmoidal manner. Trypsin was detected in the range of concentrations from 1 to 50 nM. The limit of detection was 0.2 nM. Initial reaction rates measured at different enzyme concentrations have been used to construct a calibration curve. Considering the results obtained, we believe that the TSM biosensor is a useful tool for protease analysis. Full article

In this work, we investigate the catalytic effects of gold (Au) and platinum (Pt) nanoparticle layer deposition on highly sensitive zinc oxide (ZnO) nanowires (NWs) used for selective H₂S detection in the sub-ppm region. Optimum quality pristine ZnO NWs were grown by high temperature chemical vapor deposition (CVD) in the vapor liquid solid growth (VLS) mode on silicon with a thin Au layer acting as a growth catalyst. The surface of pristine ZnO NWs was modified by systematic magnetron sputtering of discontinuous Au and Pt layers of 0–5 nm thickness. Resistive gas sensors based on the gas sensing mechanism of a chemical field effect transistor (ChemFET) with open gate, which is formed by hundreds of parallel aligned pristine Au-modified or Pt-modified ZnO NWs, were measured toward H₂S diluted in dry nitrogen (N₂) or in dry synthetic air at room temperature. Gas sensing results showed a largely improved response due to the catalytic effects of metal deposition on the ZnO NW surface. Controlled application of ZnO NW growth under optimized conditions and metal catalyst deposition showed a clear response enhancement toward 1 ppm H₂S from the initial 20% achieved with pristine ZnO to over 5000% with ZnO NWs covered by 5 nm of Au, and, hence, significantly lower than the limit of detection. Full article

Acute cardiovascular conditions require prompt assistance, which depends on a timely and accurate diagnosis. This could be achieved by using biosensor systems based on peptide aptamers capable of selectively binding protein markers of diseases. In this work, a label-free biosensor system based on fluorometric registration of the formation of a “peptide aptamer—target protein” complex is considered. It comprises a microfluidic subsystem integrated with arrays of sites with immobilized peptide aptamers, coupled with an optical detection system. The clinical sample of the whole blood is loaded into the inlet basin, where the cells are separated and plasma flows into the microfluidic channel for analysis. Peptide aptamers were created using the molecular complement search technique based on the search for systems of conjugated ion-hydrogen bonds in the three-dimensional structures of target proteins. The technology for manufacturing a microfluidic chip is a combination of thick-film and photolithography technologies based on the SU-8 photoresist, for which the relief and surface morphology have been studied. The composition of the biochip layers is selected in such a way that ultraviolet light with a wavelength of 280 nm passes through an inlet window, excites fluorescence inside the channel, which passes through the glass window, which absorbs UV-light. This wavelength accounts for the maximum absorption of aromatic amino acids—tyrosine and tryptophan. In this case, one of the last layers is a luminophore layer for re-emission of protein fluorescence as a visible light. The reading platform includes a 280 nm LED, a video sensor, 3D-printed PLA tooling, and software for processing and analyzing the received signal. Full article

A long-period fiber grating (LPFG) with maximum enhancement of evanescent field has been designed and fabricated, along with theoretical modeling, by working near the turn-around point (TAP) of the lowest order symmetric cladding mode (LP_0.2 cladding mode). The LPFG was fabricated using a point-by-point inscription technique and it was characterized in terms of surrounding refractive index (SRI) within the range of 1.333 to 1.3335 using a thermostated flow-cell. This closed cell, made of poly(methyl methacrylate) (PMMA), was designed for better handling of the sensor, because during the fabrication process, the diameter of the LPFG was reduced up to ~20 µm by chemical etching, for the maximum enhancement of the evanescent field. The sensitivity of dual peak resonance of the LP_0.2 cladding mode near TAP was measured and resulted to be ~8751 nm/SRIU with a resolution of the order of 10⁻⁵ RIU. The sensor was further used for the label-free immunosensing application by the implementation of Immunoglobulin G (IgG)/anti-Immunoglobulin G (anti-IgG) bioassay in human serum on the grating region inside the thermostated closed flow cell. Full article

Stroke is the leading cause of mortality worldwide. Differentiating patients with intracerebral hemorrhage (ICH) or ischemic stroke in the first hours of symptoms onset is of paramount importance to the optimal management of patients. Current diagnosis of acute stroke relies on neuroimaging techniques that provide valuable information but not always are readily available. In this context, the development of analytical tools capable of a rapid and on-site differentiation between the types of stroke is an important challenge with great socio-economic benefits. Glial fibrillary acidic protein (GFAP) is considered one of the ICH biomarkers in patients with symptoms of acute stroke. In this work, a simple electroanalytical device for the analysis of GFAP was developed combining stainless-steel pins and a microcentrifuge tube. The sandwich immunoassay for the determination of GFAP was carried out inside the microcentrifuge tube immobilizing the capture antibody on the bottom of the tube. The three stainless-steel pins acting as electrodes were inserted in the cap in such a way that, when the immunoassay is finished, the tube is turned bottom up allowing the electrochemical detection in the same tube. Full article

Fabrication of ion-selective sensors for continuous measurement in fluids depends on understanding the electrochemical and morphological properties of transducers. Electropolymerized nanomaterials essentially offer stable transducers that can reduce measurement drifts. This study aims to elucidate the electrochemical and morphological characteristics of electropolymerized reduced graphene oxide stabilized in polystyrenesulfonate and poly(3,4-ethylenedioxythiophene):polystyrenesulfonate composites on screen-printed carbon electrodes (rGO:PSS-PEDOT:PSS/SPCEs) using scanning electron microscopy (SEM) and cyclic voltammetry (CV) in 0.1 M potassium ferricyanide (K₃Fe(CN)₆) solution. We fabricated the rGO:PSS-PEDOT:PSS/SPCEs by two different techniques: electropolymerization deposition (EPD) and drop-casting (DC). Results revealed smaller peak-to-peak potential separation (∆E_p) of 360 mV for EPD rGO:PSS-PEDOT:PSS/SPCEs, compared to 510 mV for the DC rGO:PSS-PEDOT:PSS/SPCEs. A smaller ∆E_p indicates higher reversibility and faster electron-transfer rate at the electrode-analyte interface. SEM results showed EPD rGO:PSS-PEDOT:PSS/SPCEs have the roughest surface among electrodes; homogeneous globular structures with diameter range of 1.4–5.3 µm covered the electrode surface. In terms of electrode integrity in fluids, cracks can be seen on the surface of DC PEDOT:PSS/SPCEs after undergoing CV in 0.1 M K₃Fe(CN)₆, whereas rGO:PSS-PEDOT:PSS/SPCEs for both deposition methods maintained their integrity. Globular structures of rGO:PSS-PEDOT:PSS using EPD methods remained after undergoing CV. The results suggest that EPD serves as a potential method to fabricate a stable transducer for ion-selective sensing. This study aims to elucidate performance of nanocomposites via EPD methods, to develop stable ion-selective sensors for physiological and environmental applications. Full article

Ammonium ion (NH4+) is one of the indicators of water quality. High ammonium concentration [NH4+] in water can cause eutrophication, affect aquatic biota, and cause cell death in the central nervous system of human beings. However, current ion-selective electrodes used for water-quality monitoring are bulky, require frequent calibration owing to membrane fouling, and cannot be integrated into mobile sensor platforms. We fabricated an all-solid-state ion-selective electrode (AS-NISE) for ammonium ion using a two-step process. The first step is electropolymerization deposition on the electrode using a solution of 3,4-ethylenedioxythiophene (EDOT), sodium polystyrene sulfonate (NaPSS), and lithium perchlorate (LiCLO4), resulting in a solid-state transducer on screen-printed carbon electrodes (SPCEs), and the second is electropolymerization deposition of EDOT, NaPSS, and o-phenylenediamine (o-PD) as an ammonium ion–selective membrane (NISM) on top of the transducer. The electropolymerization deposition of the transducer and the NISM were simply achieved by cyclic voltammetry (CV) with potential from 0.0 V to 0.8 V and 50 mVs-1 scan rates. The fabricated AS-NISEcan detect [NH4+] as low as 5.7×10-5 M with a slope of 61.9 mV/decade (R2>0.99) and a linear range from 10-3 M to 1 M. These preliminary results provide an initial insight into the applicability of the simple two-steps fabrication process of NH4+ ISEs for scaling-up purposes with the ability for miniaturization and integration into a mobile sensor platform. Full article

Mixture of drinking-water supplies with sewage discharges poses disease threats in flood-stricken areas. In such exigent conditions, on-site testing of water samples is the only option, as water samples cannot be transported to laboratories owing to severely impacted transportation services. Hence, we developed a low-cost electrochemical biosensor fabricated from a screen-printed carbon electrode (SPCE) to detect E. coli O157:H7, a virulent pathogen often found in sewage discharges. We focused on understanding antigen-antibody interaction when the antibody used is not specific for E. coli O157:H7. We found that antibody immobilized on a reduced graphene oxide (rGO)–modified SPCEs distinguished between E. coli O157:H7 concentrations of 4 × 10⁸ and 4 CFU/ml, with lowest current reported for 4 × 10⁸ CFU/ml. In contrast, a reduced graphene oxide–modified SPCEs without antibody immobilization does not produce a prominent peak that distinguishes the highest and lowest E. coli concentrations. However, a few E. coli cells were still attached to the rGO/SPCEs in the absence of antibody, as shown in FESEM images. A processing step of differential readings from reference and active electrodes needs to be programmed into an Arduino^® microprocessor to realize a prototype of a bacteria sensor for field use. Full article

Liquid biopsies are becoming increasingly important as a potential replacement for existing biopsy procedures which can be invasive, painful and compromised by tumour heterogeneity. This paper reports a simple electrochemical approach tailored towards point-of-care cancer detection and treatment monitoring from biofluids using a label-free detection strategy. The mutations under test were the KRAS G12D and G13D mutations, which are both important in the development and progression of many human cancers and which have a presence that correlates with poor outcomes. These common circulating tumour markers were investigated in clinical samples and amplified by standard and specialist PCR methodologies for subsequent electrochemical detection. Following pre-treatment of the sensor to present a clean surface, DNA probes developed specifically for detection of the KRAS G12D and G13D mutations were immobilized onto low-cost carbon electrodes using diazonium chemistry and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide coupling. Following the functionalisation of the sensor, it was possible to sensitively and specifically detect a mutant KRAS G13D PCR product against a background of wild-type KRAS DNA from the representative cancer sample. Our findings give rise to the basis of a simple and very low-cost system for measuring ctDNA biomarkers in patient samples. The current time to result of the system was 3.5 h with considerable scope for optimisation, and it already compares favourably to the UK National Health Service biopsy service where patients can wait weeks for their result. This paper reports the technical developments we made in the production of consistent carbon surfaces for functionalisation, assay performance data for KRAS G13D and detection of PCR amplicons under ambient conditions. Full article

The measurement of temperature is of fundamental importance in a huge scale of applications, from nanomedicine, where the early detection of tumorous cells is an essential requirement, to microelectronics and microcircuits. Optical sensors with a micro/nano-spatial resolution can be used for temperature determination within a biological frame. Within this context, Raman spectroscopy is particularly interesting: the inelastic scattering of light has the advantage of a contactless measurement and exploits the temperature-dependence of intensities in the spectrum by observing the intensity ratio of anti-Stokes and Stokes signals. Titanium dioxide can be regarded as a potential optical material for temperature detection in biological samples, thanks to its high biocompatibility, already demonstrated in literature, and to its strong Raman scattering signal. The aim of the present work is the realization of biocompatible optical thermometers, with a sub-micrometric spatial resolution, made of titanium dioxide. Raman measurements have been performed on anatase powder using 514.5, 568.2 and 647.1 nm excitation lines of the CW Ar/Kr ion laser. The laser beam was focalized through a microscope on the sample, kept at defined temperature using a temperature controller. The Stokes and anti-Stokes scattered light was analyzed through a triple monochromator and detected by a liquid nitrogen-cooled CCD camera. Raw data were analyzed with Matlab and Raman spectrum parameters—such as area, intensity, frequency position and width of the peak—were calculated using a Lorentz fitting curve. Preliminary results showed that good reliable temperatures can be obtained. Full article

Breast cancer is the leading cancer type for women with two million new yearly infections and more than half a million dead worldwide. Human Epidermal Receptor 2 (HER2) is a prominent breast cancer biomarker that indicates aggressive cancer and is often associated with a bad prognosis and low survival rates. However, current detection methods for HER2 are often time-consuming, expensive, and require a high level of expertise. Biosensors are devices that turn biological interaction into a readable electronic signal; they are known for their high specificity and selectivity for low concentration, as well as their low cost and ease of use, thus making them a better alternative to traditional methods. Also, saliva is becoming a better alternative to blood for the detection of biomarkers due to its non-invasive collection in large quantities with simple collection methods with a richness in disease biomarkers including HER2. Thus, this project aims to develop a label-free, low cost, electrochemical biosensor for the detection of HER2 in saliva. This was done by first depositing diazonium salt onto a screen-printed electrode (SPE) through cyclic voltammetry, then immobilizing anti-HER2 antibodies on the activated SPE using 1-ethyl-3-(3-dimethylamino) propyl carbodiimide/N-hydroxysuccinimide. HER2 biomarker concentrations were detected using electrochemical impedance spectroscopy inside a microfluidic system. The biosensor showed a higher linear detection of HER2 (Y = 0.0062X + 0.1066/R² = 0.9909) in its physiological concentration range of 5 and 40 pg/mL when compared to other interference proteins: Epidermal Growth Factor Receptor (Y = 0.0016X + 0.0188/R² = 0.8072) and Human Epidermal Receptor 3 (Y = (0.0035X + 0.0225/R² = 0.1302). The biosensor was then used to detect 10 pg/mL of HER2 concentration in real saliva using the standard addition methods (Y = 0.0118X + 0.1282/R² = 0.9876). Full article

Early detection of foodborne pathogens is significant for ensuring food safety. Nowadays, the detection of pathogens found in food can take up to 72 h and it might take a week to confirm a positive sample. While standardized methods give test results in a shorter period, the reoccurring costs for each measurement are high. Therefore, it is necessary to develop technology that will be low-cost, fast, simple and accurate enough. Biosensors in combination with nucleic acid aptamers offer such possibilities. This work is focused on the development and testing of a biosensor based on DNA aptamers for detection of pathogenic bacteria Listeria innocua using the method of multi-harmonic quartz crystal microbalances (QCM). The aptasensor was prepared on the surface of a piezo crystal, whose frequency was affected by deposited mass. An aptamer specific to the genus Listeria spp. was used for the detection of this pathogen, which includes 16 subspecies, out of which 3 are excluded as their antigen structure differs from other species (L. murrayi, L. grayi, L. ivanovii). We found that addition of the pathogens at the surface of QCM transducer modified by aptamers resulted in the decrease of the resonant frequency in concentration depending manner. We also confirmed the specificity of the aptamer used for Listeria innocua, as neglected response of the sensor took place for E. coli for which Listeria spp. has some partial antigens identical and thus can cause cross-reactions in serological tests. The developed aptasensor showed promising sensitivity and specificity for real-time detection of Listeria innocua, with a detection time of 30 min. The achieved limit of detection was approximately 1.6 × 10³ CFU/mL. Full article

Fumonisin B1 (FB1), a mycotoxin commonly produced by Fusarium verticillioides and classified as a group 2B hazard, has been identified in various food products; hence, sensitive and rapid analytical detection methods are needed. Since the first reported aptamer (96 nt ssDNA) for the highly specific molecular recognition of FB1, only 30 aptamer-based biosensors have been published. A critical point, yet commonly overlooked during the design of aptasensors, is the selection of the binding buffer. In this work, a colorimetric assay was designed by incubating a folded aptamer with FB1 and the subsequent addition of gold nanoparticles (AuNPs). The changes in the aggregation profile of AuNPs by a 40 nt aptamer and a 96 nt aptamer were tested after the addition of FB1 under different buffer conditions, where the incubation with Tris-HCl and MgCl₂ exhibited the most favorable performances. The assay with the longest aptamer was specific to FB1 and comparable to other aptasensors with a limit of detection (LOD) of 3 ng/mL (A_650/520 ratio). Additionally, the application of asymmetric-flow field-flow fractionation (AF4) with multidetection allowed for the analysis of the peak area (λ) and multi-angle light scattering (MALS) with LODs of up to the fg/mL level. Full article

Epilepsy is represented by a set of neurological disorders that result in recurring seizures and convulsions. Although several types of the condition have been characterized, the underlying cause for these remains largely unknown. A number of molecular biomarkers for epilepsy have been identified including glutamate, γ-aminobutyric acid, and miRNAs. In addition, a special role appears to be played by the potassium cation. Detection of these species is anticipated to assist in both diagnosis and fundamental understanding of the condition. This review details the application of a number of biosensor devices that have been designed specifically for the detection of both molecular biomarkers and the K⁺ cation in proximity to an animal cortex. These devices offer considerable potential not only for diagnostic goals, but also for study of the cause and spread of the epileptic seizure, especially if such biosensors can detect analytes in a multiplexed, real-time manner. Full article

Magnetic microparticles (MMPs) have been notably used as platforms in biosensing. Due to their magnetic behavior, they simplify purification and separation procedures, reducing time of analysis. They also allow sample preconcentration, minimizing matrix effects, which is of key relevance for applications using real samples. Even though there is a great number of commercially available MMPs, their performance is not always reliable. In this work, we propose the synthesis of novel polymeric MMPs for their use as electrochemical immunosensing platforms. Initially, magnetic nanoparticles of a diameter of 12 ± 2 nm and a saturation magnetization of 70 emu/g were synthesized and characterized. Then, they were encapsulated in a polymeric matrix of poly (lactic-co-glycolic) acid (PLGA), generating MMPs with a diameter of 90 ± 18 nm. Later, MMPs were functionalized with polyethyleneimine (PEI) as an intermediate step for the immobilization of affinity proteins or antibodies, necessary in electrochemical immunosensing. This would allow the obtaining of MMPs comparable to the commercially available ones but possessing higher saturation magnetization. The use of such MMPs could facilitate the detection of analytes of interest in diagnostics, among other applications. Full article

A surface plasmon resonance (SPR) platform, based on a D-shaped plastic optical fiber (POF), combined with a biomimetic receptor, i.e., a molecularly imprinted polymer (MIP), is proposed to detect 2-furaldheide (2-FAL) in fermented beverages such as wine. The determination of 2-FAL in food samples is becoming a very crucial task, on the one hand for its role in the flavor and on the other in relation to its toxic and carcinogenic effects on human beings. The proposed sensing device is easy to use and cheap; it has been tested successfully for the detection and quantification of substances of interest in different fields, such as health, the environment and industry. The possibility of performing single-drop measurements is a further favorable aspect for practical applications. As an example, the use of an SPR-MIP sensor for the analysis of 2-FAL in wine, in a concentration range useful for practical applications, is here described. Full article

In this work, two sensing approaches for the specific quantification of the Salmonella genome are comparatively evaluated. These devices successfully integrate solid-phase isothermal HDA or RPA amplification in electrochemical platforms and support efficient detection of small amounts of the bacterial genome without thermal cycling, while using simple equipment. The detectability of the RPA-based sensing platform (10⁵ genomes) was surpassed by that of the HDA-based one (10 genomes), though it could be enhanced by primer design optimization. This, together with a low operation temperature compatible with conventional surface chemistry and faster amplification, makes the RPA-based electrochemical platform a promising device for point-of-need analysis. Full article

A sandwich genoassay for the detection of PCA3, a nucleic acid biomarker overexpressed in the urine of prostate cancer patients, has been developed by using the enzyme alkaline phosphatase (ALP) as a tracer of the hybrid generated onto the surface of magnetic particles. ALP converts D-glucose-1-phosphate into D-glucose, which is quantified with a personal glucose meter. The resulting methodology allows the reliable detection of PCA3 at low picomolar levels, thus fostering massive screening of prostate cancer. Full article

Surface-enhanced Raman scattering (SERS) nanoprobes have shown tremendous potential in in vivo imaging. The development of single oligomer resolution in the SERS promotes experiments on DNA and protein identification using SERS as a nanobiosensor. As Raman scanners rely on a multiple spectrum acquisition, the faster imaging in real-time is required. SERS weak signal requires averaging of the acquired spectra that erases information on conformation and interaction. To build spectral libraries, the simulation of measurement conditions and conformational variations for the nucleotides relative to enhancer nanostructures would be desirable. In the molecular dynamic (MD) model of a sensing system, we simulate vibrational spectra of the cytosine nucleotide in FF2/FF3 potential in the dynamic interaction with the Au₂₀ nanoparticles (NP) (EAM potential). Fourier transfer of the density of states (DOS) was performed to obtain the spectra of bonds in reaction coordinates for nucleotides at a resolution 20 to 40 cm⁻¹. The Au₂₀ was optimized by ab initio DFT GGA and relaxed by MD. The optimal localization of nucleotide vs. NP was defined and spectral modes of both components vs. interaction studied. Bond-dependent spectral maps of nucleotide and NP have shown response to interaction. The marker frequencies of the Au_20—nucleotide interaction have been evaluated. Full article

Affinity characterization is an essential but time-consuming task to develop reliable aptamers for tumor biomarker detection and is not always thoroughly addressed. For neutrophil gelatinase-associated lipocalin (NGAL), a potential biomarker of pancreatic cancer, two DNA aptamers were described with very different affinity. Likewise, another pair of DNA aptamers was developed with very different affinity for alpha-fetoprotein (AFP), a biomarker of hepatocellular carcinoma. In this work, we estimated the dissociation constant of these aptamers by means of a direct assay on magnetic beads modified with biomarker and electrochemical detection on screen-printed carbon electrodes. In order to improve the performance of these aptamers, we proposed the isothermal amplification of the aptamers for both biomarkers by rolling circle amplification (RCA). In the case of AFP aptamers, we also tried terminal deoxynucleotidyl transferase (TdT), a template-independent amplification. Both DNA amplifications improved the sensitivity and the apparent binding constants of the aptamers for the two cancer biomarkers. Nevertheless, this improvement depends on the true affinity of the binding pair, which ultimately limits their analytical usefulness. Full article

Breast cancer is a leading cause of death in women worldwide, and yet its pathophysiology is poorly understood. Although single-cell studies have highlighted the contribution of membrane depolarisation to the proliferation of breast cancer, dynamic signalling at a network level has not been extensively researched. It is urgent therefore to decode the intercellular signalling patterns linked to metastasis, particularly at a cell cohort level. This paper introduces a novel strategy for conducting such recordings on highly metastatic MDA-MB-231 cells, via an ultra-low noise biosensor based on a large electrode area which maximises the Helmholtz double-layer capacitance. The extracellular sensitivity of our biosensor allows the detection of pA-level random telegraph signal (RTS) noise superimposed with an omnipresent 1/f noise. The RTS noise is validated and modelled using a Markov chain. The analysis of slow cooperative potentials across the large area electrode suggests the involvement of cohort calcium signalling, and the 1/f noise analysis suggests a strong contribution of resting membrane noise. Overall, this work shows the potential of the new recording platform and statistical analysis for better understanding and predicting the underlying signalling mechanisms of metastatic breast cancer cells. In future, this platform could highlight the effects of compounds, or drugs, on the underlying activity of cancer cell cohorts in a clinical setting. Full article

In light of an upcoming series of missions beyond low Earth orbit (LEO) through NASA’s Artemis program and the potential establishment of bases on the Moon and Mars, the effects of the deep space environment on biology need to be examined and protective countermeasures need to be developed. Even though many biological experiments have been performed in space since the 1960s, most of them have occurred in LEO and for only short periods of time. These LEO missions have studied many biological phenomena in a variety of model organisms, as well as utilized a broad range of technologies. Given the constraints of the deep space environment, however, future deep space biological missions will be limited to microbial organisms using miniaturized technologies. Small satellites like CubeSats are capable of querying relevant space environments using novel instruments and biosensors. CubeSats also provide a low-cost alternative to more complex and larger missions, and require minimal crew support, if any. Several have been deployed in LEO, but the next iteration of biological CubeSats will go farther. BioSentinel will be the first interplanetary CubeSat and the first biological study NASA has sent beyond Earth’s magnetosphere in 50 years. BioSentinel is an autonomous free-flyer platform able to support biology and to investigate the effects of radiation on a model organism in interplanetary deep space. The BioSensor payload contained within the free-flyer is also an adaptable instrument that can perform biologically relevant measurements with different microorganisms and in multiple space environments, including the ISS, lunar gateway, and on the surface of the Moon. Nanosatellites like BioSentinel can be used to study the effects of both reduced gravity and space radiation and can house different organisms or biosensors to answer specific scientific questions. Utilizing these biosensors will allow us to better understand the effects of the space environment on biology so humanity may return safely to deep space and venture farther than ever before. Full article

Major advances in cancer control can be greatly aided by early diagnosis and effective treatment in its pre-invasive state. Lung cancer (small cell and non-small cell) is a leading cause of cancer-related death among both men and women around the world. A lot of research attention has been attracted to diagnosing and treating lung cancer. A common method of lung cancer treatment is based on COX-2 (Cyclooxygenase-2) inhibitors. This is because COX-2 is commonly over expressed in lung cancer and also the abundance of its enzymatic product Prostaglandin E2 (PGE₂). Instead of using traditional COX-2 inhibitors to treat lung cancer, here, we report a new anti-cancer strategy recently developed for lung cancer treatment. It adopts more abundant omega-6 (ω-6)fatty acids such as dihomo-γ-linolenic acid (DGLA) in the daily diet and the commonly high levels of COX expressed in lung cancer to promote the formation of 8-hydroxyoctanoic acid (8-HOA) through a new delta-5-desaturase (D5Di) inhibitor. The D5Di will not only limit the metabolic product, PGE₂ but also promote the COX-2 catalyzed DGLA peroxidation to form 8-HOA, a novel anti-cancer free radical byproduct. Therefore, the measurement of the PGE₂ and 8-HOA levels in cancer cells can be an effective method to treat lung cancer by providing in-time guidance. A novel sensor based on a newly developed functionalized nanomaterial, 2-dimensional nanosheets, Ti₃C₂ MXene, has proved to sensitively, selectively, precisely and effectively detect PGE₂ and 8-HOA in A549 lung cancer cells. Due to the multilayered structure and extremely large surface area, metallic conductivity and easy and versatile in surface modification, Ti₃C₂ MXene-based sensor will be able to selectively adsorb different molecules through physical adsorption or electrostatic attraction, and lead to a measurable change in the conductivity of the material with high signal-to-noise ratio and excellent sensitivity. Full article

Energy recovery methods are currently receiving a very great deal of attention from the research community. Especially, in the case of implantable biosensors where wireless energy transfer has become the main technique in these applications. An implant is a medical device manufactured to replace a missing biological structure, support a damaged biological structure, or enhance an existing biological structure. Biosensors are man-made devices, in contrast to a transplant, which is a transplanted biomedical tissue. The method of energy transfer eliminates the risk of skin infection, as well as the need for invasive surgery to change the battery. In this paper, we present the efficient approach to design an optimized octagonal spiral inductor operating at a frequency of 2.4 GHz with an inductance L value of 4 nH and a maximum factor of quality Q. The principle part of this work is based on the use of a collection of methods called metaheuristics, which are approaches used to solve a wide range of optimization problems, in order to achieve a high-performance optimized design. The problem is represented by an objective function that will be implemented using a MATLAB script and then the validation of the results obtained will be performed using the advanced design system (ADS) microwave circuit simulation software. Full article

Organic electrochemical transistors (OECTs) are now well-known, robust and efficient as amplification devices for redox reactions, typically biologically ones. In contrast, electrolyte-gated organic field-effect transistors (EGOFETs) have never been described for that kind of application because field-effect transistors are known as capacitive coupled devices, i.e., driven by changes in capacitance at the electrolyte/gate or electrolyte/semiconductor interface. For such a kind of transistors, any current flowing at the gate electrode is seen as a drawback. However, we demonstrate in this paper that not only the gate potential can trigger the source-drain current of EGOFETs, which is the generally accepted mode of operation, but that the current flowing at the gate can also be used. Because EGOFETs can work directly in water, and as an example of application, we demonstrate the possibility to monitor microalgae photosynthesis through the direct measurement of photosynthetic O₂ production within the transistor’s electrolyte, thanks to its electroreduction on the EGOFET’s gate. This paves the way for the use of EGOFETs for environmental monitoring. Full article

MicroRNAs are widely studied as circulating biomarkers for early stage diagnosis of several diseases, but the procedures for their detection and quantification are currently complex and time consuming. We demonstrate a rapid, multiplex, one-pot detection method based on two-step amplification of the signal measured by a recent label-free optical biosensor, Reflective Phantom Interface (RPI). The specific capture with surface DNA probes is combined with mass amplification by an antibody targeting DNA–RNA hybrids and polyclonal secondary antibody, all performed without washing steps. Through this method, we achieved linear, sub-pM quantification of different miRNAs in 1.5 h. The RPI enabled the characterization of equilibrium and kinetics of each individual interaction involved in this multi-step process, which allowed us to model and optimize the relative concentrations and the time intervals of the assay. Full article

Paper-based microfluidics have gained widespread attention for use as low-cost microfluidic diagnostic devices in low-resource settings. However, variability in fluid transport due to evaporation and lack of reproducibility with processing real-world samples limits their commercial potential and widespread adoption. We have developed a novel fabrication method to address these challenges. This approach, known as “Microfluidic Pressure in Paper” (μPiP), combines thin laminating polydimethylsiloxane (PDMS) membranes and precision laser-cut paper microfluidic structures to produce devices that are low-cost, scalable, and exhibit controllable and reproducible fluid flow dynamics similar to conventional microfluidic devices. We present a new μPiP DNA sample preparation and processing device that reduces the number of sample preparation steps and improves sensitivity of the quantitative polymerase chain reaction (qPCR) by electrophoretically separating and concentrating nucleic acids (NAs) continuously on paper. Our device was assembled using two different microfluidic paper channels: one with a larger pore (25 microns) size for bulk fluid transport and another with a smaller pore size (11 microns) for electrophoretic sample concentration. These two paper types were aligned and laminated within PDMS sheets, and integrated with adhesive copper tape electrodes. A solution containing a custom DNA sequence was introduced into the large pore size paper channel using a low-cost pressure system and a DC voltage was applied to the copper tape to electrophoretically deflect the solution containing NAs into the paper channel with the smaller pore size. Samples were collected from both DNA enriched and depleted channels and analyzed using qPCR. Our results demonstrate the ability to use these paper devices to process and concentrate nucleic acids. Our concentration device has the potential to reduce the number of sample preparation steps and to improve qPCR sensitivity, which has immediate applications in disease diagnostics, microbial contamination, and public health monitoring. Full article

Presently, long-lasting health disorders represent a significant health problem in developing countries. Further, epidemiological trends associated with lifestyle habits suggest that chronic conditions tend not to slow down all over the world. As such, reliable analytical techniques to manage chronic health conditions such as diabetes-mellitus, cardiovascular diseases and neurodegenerative diseases, among other non-communicable diseases (NCD), are of paramount importance. Full article

Porous silicon (PSi) has been widely used as a biosensor over the last years due to its large surface area and its optical properties. Most PSi biosensors consist in close-ended porous layers, and, because of the diffusion-limited infiltration of the analyte, they lack sensitivity and speed of response. In order to overcome these shortcomings, PSi membranes (PSiMs) have been fabricated using electrochemical etching and standard microfabrication techniques. In this work, PSiMs have been used for the optical detection of Bacillus cereus lysate. Before detection, the bacteria are selectively lysed by PlyB221, an endolysin encoded by the bacteriophage Deep-Blue targeting B. cereus. The detection relies on the infiltration of bacterial lysate inside the membrane, which induces a shift of the effective optical thickness. The biosensor was able to detect a B. cereus bacterial lysate, with an initial bacteria concentration of 10⁶ colony forming units per mL (CFU/mL), in less than 10 min. This work also demonstrates the selectivity of the lysis before detection. Not only does this detection platform enable the fast detection of bacteria, but the same technique can be extended to other bacteria using selective lysis. Full article

Investigating the binding kinetics of small molecule analytes to larger ligands, such as proteins and antibodies, is a compelling task for the field of drug and biomarker development, as well as the food industry and agro-biotechnology. Here, we improve the limit of detection of the Interferometric Reflectance Imaging Sensor (IRIS), a label-free, highly multiplexed biosensor, to perform real-time affinity measurement of small molecules binding to immobilized antibodies in a microarray format. As the analytes bind to the surface probes, the biomass accumulation on the surface is quantified by measuring the optical reflectance from the layered Si/SiO2 chip through the solution, in a common-path interferometer configuration. As a proof of concept, label-free detection of biotin molecules binding to immobilized streptavidin probes is performed, achieving 1 pg/mm2 sensitivity through signal averaging in a shot noise limited operation. Furthermore, we apply the optimized sensor to the screening of a 20-multiplexed antibody chip (MW~150 kDa ligands) against Fumonisin B1 (MW = 721.8 Da), one of the most prevalent mycotoxins found in many cereal grains such as corn and wheat. The simultaneously recorded binding curves of the toxin to the multiplexed sensor yield a signal-to-noise ratio of ≈8 when noise reduction methods of spatial and temporal averaging are utilized. Full article

A microfluidic chip for electrochemical impedance spectroscopy (EIS) is presented as biosensor for the detection of cardiac troponin I (cTnI). Troponin I is one of the most specific diagnostic serum biomarkers for myocardial infarction. As impedimetric biosensors allow direct and label-free analyte detection, they are particularly suitable for fast biomarker detection. This is essential in the diagnosis of cardiac infarctions to enable an early treatment promoting a positive outcome. The microfluidic impedance biosensor chip presented here consists of a microscope glass slide serving as base plate, sputtered electrodes, and a polydimethylsiloxane (PDMS) microchannel. Electrode functionalization protocols were developed considering a low initial impedance in addition to analyte-specific binding by corresponding antibodies and reduction of non-specific protein adsorption to prevent false-positive signals. Reagents tested for self-assembled monolayers (SAMs) on gold electrodes included thiolated hydrocarbons and thiolated oligonucleotides, where SAMs based on the latter showed a better performance. The corresponding antibody (anti-cTnI) was covalently coupled on the SAM using carbodiimide chemistry. The PDMS microchannel was bonded on the glass slide with the functionalized electrodes, and the completed microfluidic impedance biosensor chip was connected to the readout system. Sampling and measurement took only a few minutes. Application of a human serum albumin (HSA) sample, 1000 ng/mL, led to negligible signal changes, while application of a troponin I sample, 1 ng/mL, led to a significant signal shift in the Nyquist plot. The results are promising regarding specific detection of clinically relevant concentrations of cardiac markers with the newly developed impedance biosensor chip. Full article

Preliminary results on an electrosynthesized ion imprinted polymeric film (IIP-film) for Cd(II) ions determination in sensor development are here reported. The sensor was prepared by electropolymerization of 4-aminophenylacetic acid (4-APA) monomer in presence of Cd(II) ions, which acts as the template. The screen-printed carbon electrodes (SPCE) were used as transducer during sensor development, whereas the cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were selected as the electrochemical methods for the synthesis and Cd(II) ions sensing, respectively. The incubation of the developed sensor in NaOH 250 mM involved into remove the template and the formation of specific recognition cavities into the polymer. A multivariate optimization based on central composite design (CCD) was employed to study the effect of three independent parameters on electrochemical performances of the sensor. The electrochemical characterization of sensors was performed in ferrocyanide-ferricyanide redox couple and in KCl 0.1 M, the latter revealing redox properties from the polymeric film. The performances of sensors and the control (non-imprinted film, NIP) was observed in sodium acetate buffer (100 mM, pH = 5) over the Cd(II) concentration range 0.1–10 µM. Full article

The technological advantages that biosensors have over conventional technical sensors for odor detection have not yet been comprehensively analyzed. However, this is necessary for assessing their suitability for specific fields of application as well as their improvement and development goals. In this paper specific market potentials of biosensors for odor detection are identified by applying a tailored methodology that enables the derivation and systematic comparison of both the performance profiles of biosensors as well as the requirement profiles for various application fields. Therefore, the fulfillment of defined requirements is evaluated for biosensors by means of 16 selected technical criteria in order to determine a specific performance profile. Further, a selection of application fields for odor detection sensors is derived to compare the importance of the criteria for each of the fields, leading to market-specific requirement profiles. The analysis reveals that the requirement criteria considered to be the most important ones across all application fields are high specificity, high selectivity, high repeat accuracy, high resolution, high accuracy, and high sensitivity. All these criteria, except for the repeat accuracy, can potentially be better met by biosensors than by technical sensors, according to the results obtained. Therefore, biosensor technology in general has a high application potential for all the areas of application under consideration. Health and safety applications especially are considered to have high potential for biosensors due to their correspondence between requirement and performance profiles. Full article

Improving the sensitivity of the competitive lateral flow immunoassay (LFIA) is important, given the increasing demands for the monitoring of chemical contaminants in food. The choice of nanosized marker is an essential task for improving the LFIA sensitivity. In this study, a CdSe/ZnS quantum dot (QD)-based LFIA combined with a portable reader was developed for rapid and quantitative detection of an antibiotic lincomycin (LIN). The performance of the proposed fluorescence LFIA was compared to the conventional gold nanoparticle (AuNP)-based LFIA realized with the same immunoreagents. The visual cutoff values were 10 ng/mL for AuNP-based LFIA and 20 ng/mL for QD-based LFIA. Furthermore, the instrumental limits of detection have been shown to be comparable for both nanosized markers and amounted to 0.4 ng/mL for AuNPs and 0.2 ng/mL for QDs, respectively. According to the results obtained, both LFIAs may be used for rapid, cost-effective, on-site testing of antibiotics, in particular LIN. However, the QD-based LFIA exhibits lowest limit of detection with the least immunoreagent consumption, which makes it economically beneficial. Full article

Proteases play an important role in various biological as well as dairy industrial processes. In this work, we focused on chymotrypsin, which is an important protease in human digestion. Chymotrypsin also cleaves milk proteins, which makes it useful in the study of activity of milk proteases. In this work, we focused on detection of chymotrypsin using optical and gravimetric techniques, their peculiarities, and the limit of detection (LOD) (0.15 ± 0.005 nM and 0.64 ± 0.31 nM for optical and gravimetric methods, respectively). While the optical detection is easier to implement, the gravimetric method is more robust for sample preparation. Both methods could represent a novel assay for detection of chymotrypsin and other milk proteases. Full article

The method of Salmonella detection recommended is cultural, but it is laborious, presents a high consumption of material, and requires about five days for presumptive results. Immunosensor is an alternative tool that has shown promising and rapid results, although many devices have their performance evaluated only under buffering conditions and few achieve the validation stage. The objective was to perform a pre-collaborative validation of an electrochemical immunosensor assembled on screen-printed electrodes for the detection of Salmonella sp. in milk. The antibodies were immobilized by cysteamine self-assembled monolayer. The sandwich-type amperometric immunosensor was evaluated for contaminated raw and whole UHT milk and compared to performance with a gold standard reference method (BAM) according to AOAC recommendations for a single laboratory. A binary response (positive/negative) of the immunosensor was used based on a cut off established from current electric obtained for the absence of the pathogen. There was no significant difference for the results of the biosensor and the reference method, in the absence and the levels from 10¹ to 10³ CFU mL⁻¹ of Salmonella Typhimurium for the two types of milk. This result indicates the efficiency of the biosensor in detecting the pathogen into a complex matrix. Full article

The present work reports the potential of a bio-inspired system based on spectrometry, also known as Electronic Eye (EE), capable of detecting different Tequila samples. The reported system analyzes small volumes of Tequila Reposado and Blanco by calculating samples’ absorbances, using a low cost and portable instrumentation employing a CCD camera. The absorbance imaging method consisted of exciting samples with light passes through an 8MP camera connected to a Raspberry Pi Card. The camera’s image data are analyzed using MATLAB 2018b to be represented in Red, Green and Blue (RGB) components for each pixel, in order to get an approximation of the absorbance and the Surface Color Index (Isc) associated with sample concentration. Using the developed EE, it was possible to identify seven different kinds and brands of Tequila. From the obtained results, it was observed that the average absorbance of the Tequila Reposado was greater than the absorbance of the Tequila Blanco. Otherwise, with the Isc, the Tequila Blanco color index is lower concerning the Tequila Reposado’s. Finally, the EE allowed the identification of Tequila samples with reproducibility and repeatability. Full article

Carbendazim is a broad-spectrum benzimidazole-type fungicide effective against fungi that compromise the safety/quality of food products. Despite its potential usefulness, carbendazim constitutes a major environmental pollutant, being hazardous for humans and animals; therefore, reliable determination of carbendazim levels in water, soil, and food samples remains a highly desirable analytical goal. Herein, an optical (white light reflectance spectroscopy, WLRS) label-free biosensor for fast and sensitive determination of carbendazim is presented. The transducer is a SiO₂/Si chip, on which a suitable benzimidazole-conjugate has been immobilized; determination is based on the competitive immunoassay format: A mixture of an in-house developed anti-carbendazim antibody with the calibrators/samples is pumped over the chip, followed by biotinylated secondary antibody and unlabeled streptavidin. The WLRS platform allows for real-time monitoring of biomolecular interactions carried out onto the SiO₂/Si chip by transforming the shift in the reflected interference spectrum caused by the immunoreaction to effective biomolecular adlayer thickness. The sensor is capable of detecting carbendazim levels within 28 min (LoD: 20 ng/mL; intra- and inter-assay CVs: ≤6.9% and ≤9.4%, respectively). Excellent analytical characteristics and short analysis time combined with its small size render the proposed WLRS biosensor ideal for future point-of-need determination of carbendazim in food and environmental samples. Full article

Reactive oxygen/nitrogen species (ROS/RNS) have a great impact on cellular response to stress, cell proliferation, cell death, cancer, aging, or male infertility. Additionally, in the food industry and for consumers as well, it is very important to monitor quality and freshness of raw meat. Different factors are a sign of meat alteration (e.g., discoloration, rancidity, alteration of flavor). One pathway of alteration is the scavenging activity of myoglobin towards RNS (such as peroxynitrite, PON). This paper presents the development of an electrochemical PON sensor using cobalt phthalocyanine (CoPc) as a simple, cost effective, highly thermally stable, biomimetic catalyst, and the application of this electrochemical screen-printed carbon electrode (SPCE)-based sensor to meat extract samples, using flow injection analysis (FIA). The reduction of peroxynitrite, mediated by CoPc, occurs at a very low potential (around 0.1 V vs. Ag/AgCl pseudoreference); as for higher potentials, the mechanism of mediation changes, and the electro-oxidation of PON is observed. The surface of the modified electrode was characterized using SEM, FTIR and Cyclic Voltametry. The interaction of PON with myoglobin was studied using both UV-Vis and chronoamperometry (at 0.1 V, using the FIA system). The calibration of the electrode was performed: I_red (nA) = 6.313·C_PON (µM) + 17.469; (R² = 0.9938). The calculated LOD was equal to 2.37 µM and the linear range was 3–180 µM. The performance of the electrode can be further improved using a pre-treatment (electro-reduction of the CoPc deposited film, at −0.3 V, during 60s). This could help us monitor and quantify how much PON was decomposed and when meat extracts are spiked with different PON concentrations, in a highly selective, sensitive, and reproducible way. Full article

Prostate-specific Antigen (PSA) is the biomarker that is used for prostate cancer (PCa) detection, although its lack of specificity results in a high rate of false-positives and many unnecessary biopsies. Therefore, there is a need for more specific cancer biomarkers for PCa. Recent studies have shown that the aberrant glycosylation of proteins is a common feature of the presence of cancer. In the case of prostate cancer, there are changes in core-fucose and sialic acids in the glycan structure of PSA. In this work, we describe two different strategies to direct the selection of aptamers toward the glycans of PSA. From these strategies, we identified two aptamers (PSA-1 and PSAG-1) that bind to the glycan structure of PSA with high affinity. Both aptamers were applied in the design of electrochemical aptasensors, in sandwich and direct formats, in order to detect the changes in the glycosylation of PSA. The sensors responded to different levels of PSA in serum, and they showed higher potential to discriminate clinically-meaningful PCa than the ELISA (Enzyme-linked immunosorbent assay) test used in hospitals (reducing the number of false positives), although validation on more samples is needed. Full article

Veterinary drugs could contaminate animal-derived food products for human consumption. Some antibiotic residues (e.g., chloramphenicol (CAP), nitrofuran metabolites) are banned in foodstuffs of animal origin (e.g., milk, honey, etc.) in the European Union because of toxicological risks for the consumer. Screening methods applied for food safety monitoring should be sensitive, specific, cheap, quick, and portable for field testing (e.g., self-control). Electrochemical biosensors make it possible to develop a promising and economically interesting approach. An innovative and cheap electrochemical method based on disposable screen-printed carbon electrodes (SPCE), coupled to magnetic beads (MB), that allows the simultaneous detection of three families of antibiotics in milk was published by a Spanish academic team. When the biosensor method was applied to detect CAP residues in honey, two major issues were identified: firstly, the very low levels of residues to reach (i.e., regulatory limits below 1 µg/kg), and secondly, the complexity of the honey matrix; there is not a single honey matrix. Honey composition and color vary considerably depending on the botanical origin. Moreover, some honey ingredients can interfere with the electrochemical detection, especially substances with antioxidant activities (e.g., polyphenols). Therefore, in parallel with the optimization of the electrochemical method, the reduction of matrix effects was a big challenge. Full article

Listeria monocytogenes is an intracellular bacterium that causes serious epidemic and sporadic food-borne illnesses in humans. Rapid and trustworthy methods are necessary for the detection of the pathogen to prevent potential food contamination. The aim of this study was to test a newly developed L. monocytogenes biosensor on actual food samples and validate its ability to detect the presence of pathogens robustly and accurately. The newly developed method uses a cell-based biosensor technology (BERA) and a portable device developed by EMBIO Diagnostics called B.EL.D, and provides results within 3 min. Tests were conducted on ready-to-eat lettuce salads, milk and halloumi cheese and the results indicate that the novel system was able to identify inoculated samples with 98%, 90%, and 91% accuracy, respectively. Furthermore, the limit of detection was determined to be as low as 0.6 log CFU mL⁻¹ or g⁻¹ in all food types. Classification of the samples Above or Below the detection limit was accessed through a newly developed algorithm for each food substrate. Samples were also analyzed with the ISO 11290-1:2017 and 11290-2:2017, in parallel. Thus, it was concluded that the newly developed biosensor can be a useful tool in the food supply chain, decreasing the required time for the detection of pathogens and increasing the number of tested samples before they reach the market. Full article

The presented results indicate virus-like particles of the coronavirus (CVP) using a nanowire (NW) biosensor based on silicon-on-insulator technology. In the experiment, we used suspensions of CVP and of specific antibodies to the virus. Measurements of the current value of the field-effect transistor before and after the introduction of the CVP on the surface of the nanowire were performed. Results showed antibody + CVP complexes on the phase section with the surface of the nanowire modulate the current of the field-effect transistor; CVP has an electrically positive charge on the phase section “nanowire surface-viral suspension»; antibody + CVP complexes have an electrically negative charge on the phase section “nanowire surface-viral suspension”; the sensitivity of the biosensor is made up of 10⁻¹⁸ M; the time display was 200–300 s. Full article

In this paper, a fabrication method for two-dimensional multi-electrode arrays (MEAs) using inexpensive material and method is proposed. The focus in this work is on the design and fabrication of 2D Microelectrode arrays using metallic electrodes on a silica substrate. Titanium/gold multi-electrode arrays containing 60 electrodes with optimized metal thicknesses and 30 μm diameter, covered with thin modified SU-8 insulator layer as biocompatible material have been designed and manufactured using the standard photolithography-based microfabrication method. The utilization of affordable and more accessible materials and simpler techniques can be mentioned as the distinct point of the proposed fabrication method. Using these multi-electrode arrays, it is possible to either record or stimulate cells by accessing multiple sites of cell tissues and collect signals from the sources around each electrode simultaneously. Precisely adjusting the size, distance, and number of microelectrodes causes high measurement selectivity and reliability which has been taken into account in the design of the microelectrodes. In this study, we manufactured a preliminary representative MEA and the bio-compatibility of the manufactured MEA is going to be evaluated by neural cells, obtained from rat cortices. The main aim of this study is to compare our inexpensive strategy with other approaches. Full article

At the end of year 2019 the first reports appeared of a new coronavirus and on 31st December 2019 WHO declared a public health emergency of international concern. To date (as of 6:08 pm CET, 24th November 2020) according to WHO the new coronavirus, now called severe acute respiratory syndrome (SARS)-CoV-2, has infected 58,900,547 people and killed 1,393,305 people worldwide. It is extremely important to develop means for express diagnostics to ensure prompt action to limit the spread of infection. One of the diagnostic approaches, is the detection of viral particles in swabs. This approach can be realized using a biosensor with specific ligands, based on peptide molecules complementary to surface viral proteins. The concept of the so-called Systems of Conjugated Ionic-Hydrogen Bonds (abbreviated—SSIVS, CIHBS) implemented in the Protein-3D computer program, was applied to analyze the spatial structures of the bonds between the SARS-CoV-2 spike protein and the ACE-2 (Angiotensin converting enzyme 2) receptor, in order to reveal the perspective peptide sequences. There are two clearly marked areas of contact of the spike with the cell receptor—upper and lower, which are visualized in the SSIVS form, and the complex formed at this site is strong enough to ensure its attachment to the coronavirus spike and can compete for binding with the ACE-2 receptor. Two peptides were developed that form a spatial structure complementary to the coronavirus spike: of eight (No. one) and of 15 (No. two) amino acid residues. The peptides were covalently bound to biochip platforms via neutral linkers to form sites with peptides No. one and No. two. The third site has a neutral hydrophilic surface to serve as a reference. The platform was integrated with a microfluidic channel and was used as a flow through device. The detection of bound viral particles was carried out using UV excitation and direct registration of viral proteins fluorescence. The preliminary laboratory tests demonstrated the efficiency of the biosensor. Full article

Micro-optical resonators have been introduced as sensors in many applications for a wide number of variable types of stimuli due to their very high resolution, high sensitivity, and high-quality factor. In this paper, a novel micro-optical sensor was designed and tested as a concentration meter for chemical composition of a solution. The micro-optical resonator used is based on whispering gallery mode (WGM). This phenomenon appears when a tapered, single-mode laser carrying micro-optical fiber is evanescently coupled with a polymeric or silica micro-optical resonator. The presented sensor shows the change in concentration by experiencing a change in its morphology due to the varied viscosity of its environment. The variation of concentrations or fluid contents results in a change between the radii of the micro-optical resonator. With varied chemical composition and concentration in the tested sample varied infinitesimally small morphological changes are detected. The change in the resonators shape is read as a WGM shift in the resonance transmission spectrum, which is interpreted using a technique called cross-correlation, which compares the output across time to display the shift, which is later translated into distinct concentration levels. The proposed, exceptionally low-cost sensors were able to detect change at very high resolutions allowing better sensitivity along with wider range of variation. Experimental work for detection of ranges of concentrations of variable type of contaminants is presented. Full article

Early detection of pollutants in wastewater, water coming out of treatment facilities, drinking water, and water for agricultural needs is a challenging problem. Effective water quality monitoring requires development of new methods for express detection of pollutants. Enzymes from bioluminescent bacteria can be used for the development of new express enzyme-based bioassay systems. This work demonstrates, for the first time, a microfluidic chip to generate emulsion droplets containing two enzymes of the bacterial bioluminescent system (luciferase and NAD(P)H:FMN-oxidoreductase) with reaction substrates. The developed chip generated “water-in-oil” emulsion droplets with a volume of 0.1 $\mathsf{\mu }$L and a frequency of up to 12 droplets per second. A portable photomultiplier tube (PMT) was used to measure the bioluminescent signal in each individual droplet; the signal-to-noise ratio was 3000/1. The intensity of luminescence in droplets depended on the concentration of copper ions. The limit of detection (LOD) for copper sulfate was 1 mg/L. We showed that bioluminescent enzymatic reactions can be carried out in droplet reactors that can be applied for online monitoring of water quality. Thus, the suggested method of biological measurements has a good perspective for biosensing in general. Full article

The microacoustic methods of biomedical analysis, implemented on piezoelectric crystals and ceramics, are becoming increasingly popular due to the fact of their potential for integration into laboratories-on-a-chip, biochips, and biosensors as functional elements of biosensors. An important stage in diagnostics of infectious diseases is the identification of pathogens. One possible applications of such a sensor is an alternative to the time- and labor-consuming Gram method of discriminating bacteria according to the composition of their cell walls. Thus, bacteria, which in a Gram staining procedure do not decolor after application of the dye solution, are classified as Gram-positive (G(+)). They are surrounded with a thick peptidoglycan layer that is pulpy and dampens acoustic waves. While Gram-negative (G(–)) bacteria, which acquire a red color in a Gram procedure, are covered with a thin and springy layer, demonstrating resonance effects when interacting with acoustic fields. Thus, G(+) and G(–), which are differently colored in Gram procedures, also react differently to an external acoustic field: for G(–) bacteria, this was a sharp decrease in the Q-factor of the “resonator–suspension” system and a shift of the resonance curve to lower frequencies. While for G(+) bacteria, although a certain shift of the resonance curve was also observed, the bandwidth of the resonance curve practically did not change. This effect was studied for L. acidophilus (G(+)) and Escherichia coli (G(–)) bacilli with quarts resonators of 4 MHz, 5 MHz, and 10 MHz. The biosensor was tested using Lactobacillus fermentum, E. coli M-17, Bifidobacterium bifidum, Burkholderia cepacian, and Staphylococcus aureus. At this stage, it has been demonstrated that the method is particularly effective for discriminating bacteria of a similar shape, such as, for example, cocci. The discrimination of the Gram factor for cocci and bacilli was less accurate and needs further studies for selection of precise resonance frequencies. Full article

Nanozymes (NZs) are catalytically active nanomaterials that have enzyme-like activity but possess increased stability and greater availability due to the fact of their simpler preparation technologies. Nanozymes as nanoscale artificial enzymes demonstrate various catalytic specificities as oxidoreductases, such as peroxidase, catalase, laccase, and others as well as hydrolases, proteases, endonucleases, DNA-ases, NO synthases, etc. A broad variety of NZs exhibits dual- or multienzyme mimetic activity. Nanozymes as stable, low-cost mimetics of natural enzymes have a high potential for application in different branches of biotechnology including scientific investigations, industry, and ecology. Nanozymes can be applied in medicine as diagnostic tools and components of therapeutic drugs. Since NZs have high catalytic activity and chemical and biological stability, they are very promising in the construction of biosensors and biofuel cells. For these reasons, the search for simple methods of synthesis and characterization of different NZs is a very important and real problem. The “green” synthesis of Prussian blue analogous as peroxidase-like NZs using oxido-reductases is described in this study. The obtained green-synthesized hexacyanoferrates (gHCFs) of transition metals were characterized by structure, size, composition, catalytic properties, electro-mediator activities, and substrate specificity. Copper hexacyanoferrate (gCuHCF) was studied in more detail. When immobilized on a graphite electrode (GE), gCuHCF under special conditions of pH and tension gave amperometric signals on hydrogen peroxide and can be used as a peroxidase mimetic in oxidase-based biosensors. Under other conditions, gCuHCF/GE reacts to other analytes. We propose that gHCFs of transition metals synthesized via enzymes may become prospect platforms for the construction of multi-functional amperometric (bio)sensors. Full article

Pathogenic bacterial contamination in food is a public health concern. It represents a health and safety consumer risk that could cause several diseases and even death. Currently, the food industry uses culture-based assays to determine the presence of pathogens as a gold standard method. Although this method is highly accurate, it is often time-consuming and expensive. In this regard, the development of biosensing platforms results as an alternative for the reduction of time and cost of pathogenic bacteria detection in food. In this work, we report the development of a single-step bacterial detection platform based on graphene oxide. Non-radiative energy transfer between graphene oxide coated microplates (GOMs) and photoluminescence bioprobes (PLBs) is presented in absence of the target analyte, but in presence of analyte, PLBs exhibit strong photoluminescence due to the distance between GOMs and PLBs. These PLBs are based on quantum dot (Qds)-antibody (Ab) complexes, thereby resulting as a biorecognition and interrogation element. Escherichia coli was used as model analyte. In optimal conditions, the bacterial detection platform reached a limit of detection around 2 CFU mL⁻¹ in 30 min, enabling a fast and sensitive alternative for bacterial detection. The biosensing platform was also used to test food industry samples achieving a qualitative response, that allows determining the presence of E. coli during the first 30 min of the assay. This biosensing strategy potentially offers a low-cost and quick option for the food industry to assure the quality of the product and consumer safety. Full article

Immunoassays are, at present, an important tool for diagnostics, drug development, and environmental monitoring. However, most immunoassays involve procedures that require many elements for their development. We introduce a novel biosensing platform based on fluorescence quenching caused by graphene oxide (GO) for the detection of Human-IgG and Prostate-Specific Antigen (PSA). We employ a single antibody for the capture and detection processes, avoiding washing steps. FITC fluorophore was conjugated with antibodies for H-IgG detection, whereas quantum dots were conjugated with antibodies for PSA detection. The simple biosensing platform consists of covering a 96-well microplate (with a polystyrene bottom) with GO. The graphene oxide adhesion is possible by way of electrostatic interactions between the plate surface modified with amino groups (positively charged) and the graphene oxide (negatively charged). This proposal showed an excellent response for the detection of Human-IgG, with acceptable precision (from 0.27% to 5%). The limit of detection reached for H-IgG was 3.35 ng mL-1. In the same manner, for PSA detection, the limit of detection reached was 0.02 ng mL-1 and the precision range was from 0.7% to 15.2%. Furthermore, this biosensing platform was demonstrated to operate with real samples of human urine doped with different concentrations of prostate-specific antigen. Full article

Paper substrates have shown a high potential for development of cost-effective and efficient point-of-care biosensors, essential for public healthcare and environmental diagnostics. Most paper-based biosensors rely on qualitative colorimetric detection schemes with high limits of detection. To overcome this limitation, technologies that combine paper-based substrates and electrochemical detection are being developed to allow for quantification and achieve better performances. In this work, we explore the potential of dielectric measurements towards electrical detection of whole-cell bacteria in nitrocellulose membranes, a paper-derivative. Impedance spectroscopy was considered to characterize the membranes with and without Bacillus thuringiensis cells, used as model microorganism. To specifically target this bacterial strain, phage endolysin cell-wall binding domain (CBD) encoded by a bacteriophage targeting B. thuringiensis were prepared and integrated into the membranes as recognition biointerface. The fluid sample containing the bacteria is conducted in the membrane through passive capillarity, and the bacteria are specifically immobilized in the test zone. Resulting changes of the dielectric properties of the membrane are sensed through impedance changes, highlighting the contribution of ions in the bacterial detection mechanism. This experimental proof-of-concept illustrates the electrical detection of 10⁸ CFU/mL bacteria in low-salinity buffers within 5 min. Full article

Electric Cell-substrate Impedance Sensing (ECIS), xCELLigence and cellZscope are commercially available instruments which are able to measure the impedance of cellular monolayers continuously and with high precision. The small currents used allow the label-free, real-time monitoring of the cells in a non-invasive manner. Despite the widespread use of these systems individually, direct comparisons between the systems have not been published. In order to compare the sensitivity of the instruments, the responses of the brain microvascular endothelial cell line hCMVEC to the inflammatory cytokines TNFα and IL1β were measured on all three instruments simultaneously. All three instruments showed transient decreases, followed by prolonged increases in impedance. Although xCELLigence could detect these changes, it was unable to determine which component of the barrier was affected. In contrast, ECIS and cellZscope were both able to attribute responses to particular barrier components, and ECIS had a higher sensitivity than cellZscope. Finally, as cellZscope uses Transwells, it allows access to the basolateral compartment, an important advantage of this technology. Furthermore, although xCELLigence readings are equivalent to ECIS, the reduced frequency range greatly limits interpretation. This work demonstrates that instruments must be carefully selected in order to ensure that they are appropriate for the experimental questions being asked. Full article

The increasing demand for fast and on-site information has generated great interest in developing simple and portable analytical devices that provide reliable responses. Electroanalytical devices fit perfectly with these purposes because of their ease of use, low cost and facility of miniaturization. Moreover, the growing interest in the construction of do-it-yourself electronic devices has spread the use of common and mass-produced materials for the development of analytical devices. In this context, it is presented here the use of gold-plated pins, from standard connections, and stainless-steel pins, from needlework, as electrodes in (bio)electroanalytical platforms. Three different analytical platforms combining those pins with paper, transparency sheets or micropipette tips were constructed and applied in food and environmental analyses: glucose determination in beverages and surfactant analysis in water. Full article

The increasing use of fertilisers rises the risk of eutrophication, a sudden algal bloom that seriously damage ecosystems due to critical oxygen depletion. Continuous monitoring of oxygen in environmental waters could improve the detection of eutrophication and prevent anoxic conditions. However, online and in situ dissolved oxygen sensors are yet to be implemented due to poor portability and power requirements. Here, we propose a ceramic soil microbial fuel cell as a self-powered sensor for algal growth detection via monitoring of dissolved oxygen in water. The sensor signal follows the characteristic photosynthetic cycle, with a maximum day current of 0.18 ± 0.2 mA and a minimum night current of 0.06 ± 0.34 mA, which correlates with dissolved oxygen (R² = 0.85 (day); R²= 0.5 (night)) and algal concentration (R² = 0.63). A saturated design of experiments on seven factors suggests that temperature, dissolved oxygen, nitrates, and pH are the most influential operational factors in the voltage output. Moreover, operating the system at maximum power point (R_ext = 2 kΩ) improves the sensor sensitivity. To the best of our knowledge, this is the first proposed MFC-based biosensor for in-field, early detection of eutrophic events. Full article