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Edible biosensors can measure a wide range of physiological and biochemical parameters, including temperature, pH, gases, gastrointestinal biomarkers, enzymes, hormones, glucose, and drug levels, providing real-time data. Edible biocatalytic biosensors represent a new frontier within healthcare technology available for remote medical diagnosis. The main challenges to develop edible biosensors are: i) finding edible materials (i.e. redox mediators, conductive materials, binders and biorecognition elements such as enzymes) complying with Food and Drug Administration (FDA), European Food Safety Authority (EFSA) and European Medicines Agency (EMEA) regulations; ii) developing bioelectronics able to operate in extreme working conditions such as low pH (∼pH 1.5 gastric fluids etc.), body temperature (between 37 °C and 40 °C) and highly viscous bodily fluids that may cause surface biofouling issues. Nowadays, advanced printing techniques can revolutionize the design and manufacturing of edible biocatalytic biosensors. This review outlines recent research on biomaterials suitable for creating edible biocatalytic biosensors, focusing on their electrochemical properties such as electrical conductivity and redox potential. It also examines biomaterials as substrates for printing and discusses various printing methods, highlighting challenges and perspectives for edible biocatalytic biosensors., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

DNA can be readily amplified through replication, enabling the detection of a single-target copy. A comparable performance for proteins in immunoassays has yet to be fully assessed. Surface-plasmon-resonance (SPR) serves as a probe capable of performing assays at concentrations typically around 10⁻⁹ molar. In this study, plasmonic single-molecule assays for both proteins and DNA are demonstrated, achieving limits-of-detections (LODs) as low as 10⁻ 2 ⁰ molar (1 ± 1 molecule in 0.1 mL), even in human serum, in 1 h. This represents an improvement in typical SPR LODs by eleven orders-of-magnitude. The single-molecule SPR assay is achieved with a millimeter-wide surface functionalized with a physisorbed biolayer comprising trillions of recognition-elements (antibodies or protein-probe complexes) which undergo an acidic or alkaline pH-conditioning. Potentiometric and surface-probing imaging experiments reveal the phenomenon underlying this extraordinary performance enhancement. The data suggest an unexplored amplification process within the biomaterial, where pH-conditioning, driving the biolayer in a metastable state, induces a self-propagating aggregation of partially misfolded proteins, following single-affinity binding. This process triggers an electrostatic rearrangement, resulting in the displacement of a charge equivalent to 1.5e per 10 2 recognition elements. Such findings open new opportunities for reliable SPR-based biosensing at the physical detection limits, with promising applications in point-of-care plasmonic systems., (© 2025 The Author(s). Advanced Materials published by Wiley‐VCH GmbH.)

A ground-breaking graphene-based biosensor designed for label-free detection of immunoglobulin M (IgM) achieving a remarkable concentration of 100 zeptomolar (10 -19 m), is reported. The sensor is a two-terminal device and incorporates a millimeter-wide gold interface, bio-functionalized with ≈10 12 anti-IgM antibodies and capacitively coupled to a bare graphene electrode through a water-soaked paper strip. In this configuration, few affinity binding events trigger a collective electrostatic reorganization of the protein layer, leading to an extended surface potential (SP) shift of the biofunctionalized Au surface. The SP shift, mediated by electrolyte capacitive coupling, induces a corresponding shift in the Fermi level of graphene. This shifts the graphene phonon frequencies, which are measured by Raman spectroscopy. Decoupling the sensing interface from the transducing graphene layer provides flexibility in surface chemistry modifications, while preserving the graphene integrity. A key aspect of this biosensor is its ability to precisely determine the graphene charge neutrality point from the voltage dependence of phonon frequency shifts, enabling detections of biomarker at unprecedented low concentrations. The integration of graphene with optical probing demonstrates a proof-of-concept and establishes a ground-breaking approach to in situ biomarker detection, setting the stage for a future generation of portable opto-electronic high-performance diagnostic tools for single-marker detection., (© 2025 The Author(s). Small Methods published by Wiley‐VCH GmbH.)

The electric dipoles of proteins in a biolayer determine their dielectric properties through the polarization density P. Hence, its reproducibility is crucial for applications, particularly in bioelectronics. Biolayers encompassing capturing antibodies covalently bound at a biosensing interface are generally preferred for their assumed higher stability. However, surface physisorption is shown to offer advantages like easily scalable fabrication processes and high stability. The present study investigates the effects of electric-field (EF)-cycling of anti-Immunoglobulin M (anti-IgM) biolayers physisorbed on Au. The impact of EF-cycling on the dielectric, optical, and mechanical properties of anti-IgM biolayer is investigated. A reduction of the dispersion (standard deviation over a set of 31 samples) of the measured P values is observed, while the set median stays almost constant. Hence, physisorption combined with EF cycling, results in a biolayer with highly reproducible bioelectronic properties. Additionally, the study provides important insights into the mechanisms of dielectric rearrangement of dipole moments in capturing biolayers after EF-cycling. Notably, EF-cycling acts as an annealing process, driving the proteins in the biolayer into a statistically more probable and stable conformational state. Understanding these phenomena enhances the knowledge of the properties of physisorbed biolayers and can inform design strategies for bioelectronic devices., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Artificial neural networks (ANNs) are versatile tools capable of learning without prior knowledge. This study aims to evaluate whether ANN can calculate minute volume during spontaneous breathing after being trained using data from an animal model of metabolic acidosis. Data was collected from ten anesthetized, spontaneously breathing pigs divided randomly into two groups, one without dead space and the other with dead space at the beginning of the experiment. Each group underwent two equal sequences of pH lowering with pre-defined targets by continuous infusion of lactic acid. The inputs to ANNs were pH, ΔPaCO 2 (variation of the arterial partial pressure of CO 2 ), PaO 2 , and blood temperature which were sampled from the animal model. The output was the delta minute volume (ΔV M ), (the change of minute volume as compared to the minute volume the animal had at the beginning of the experiment). The ANN performance was analyzed using mean squared error (MSE), linear regression, and the Bland-Altman (B-A) method. The animal experiment provided the necessary data to train the ANN. The best architecture of ANN had 17 intermediate neurons; the best performance of the finally trained ANN had a linear regression with R 2 of 0.99, an MSE of 0.001 [L/min], a B-A analysis with bias ± standard deviation of 0.006 ± 0.039 [L/min]. ANNs can accurately estimate ΔV M using the same information that arrives at the respiratory centers. This performance makes them a promising component for the future development of closed-loop artificial ventilators., Competing Interests: Declarations. Compliance with ethical standards: The study received approval from the local institutional review board for the care of animal subjects. The animals were handled in accordance with the regulations of the Swedish Board for Laboratory Animals. The animal experiments were performed at the Hedenstierna Laboratory of Uppsala University, Sweden. Statement on artificial intelligence: No method of creative artificial intelligence was used to generate the text or the figures of the present report. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

The possibility to print electronics by means of office tools has remarkedly increased the possibility to design affordable and robust point-of-care/need devices. However, conductive inks suffer from low electrochemical and rheological performances limiting their applicability in biosensors. Herein, a fast CO 2 laser approach to activate printed carbon inks towards direct enzymatic bioelectrocatalysis (3 rd generation) is proposed and exploited to build biosensors for D-fructose analysis in biological fluids. The CO 2 laser treatment was compared with two lab-grade printed transducers fabricated with solvent (SB) and water (WB) based carbon inks. The use of the laser revealed significant morpho-chemical variations on the printed inks and was investigated towards enzymatic direct catalysis, using Fructose dehydrogenase (FDH) integrated into entirely lab-produced biosensors. The laser-driven activation of the inks unveils the inks' direct electron transfer (DET) ability between FDH and the electrode surface. Sub-micromolar limits of detection (SB-ink LOD = 0.47 μM; WB-ink LOD = 0.24 μM) and good linear ranges (SB-ink: 5-100 μM; WB-ink: 1-50 μM) were obtained, together with high selectivity due to use of the enzyme and the low applied overpotential (0.15 V vs. pseudo-Ag/AgCl). The laser-activated biosensors were successfully used for D-fructose determination in complex synthetic and real biological fluids (recoveries: 93-112%; RSD ≤8.0%, n = 3); in addition, the biosensor ability for continuous measurement (1.5h) was also demonstrated simulating physiological D-fructose fluctuations in cerebrospinal fluid., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Herein we introduce a novel water-based graphite ink modified with multiwalled carbon nanotubes, designed for the development of the first wearable self-powered biosensor enabling alcohol abuse detection through sweat analysis. The stencil-printed graphite (SPG) electrodes, printed onto a flexible substrate, were modified by casting multiwalled carbon nanotubes (MWCNTs), electrodepositing polymethylene blue (pMB) at the anode to serve as a catalyst for nicotinamide adenine dinucleotide (NADH) oxidation, and hemin at the cathode as a selective catalyst for H 2 O 2 reduction. Notably, alcohol dehydrogenase (ADH) was additionally physisorbed onto the anodic electrode, and alcohol oxidase (AOx) onto the cathodic electrode. The self-powered biosensor was assembled using the ADH/pMB-MWCNTs/SPG||AOx/Hemin-MWCNTs/SPG configuration, enabling the detection of ethanol as an analytical target, both at the anodic and cathodic electrodes. Its performance was assessed by measuring polarization curves with gradually increasing ethanol concentrations ranging from 0 to 50 mM. The biosensor demonstrated a linear detection range from 0.01 to 0.3 mM, with a detection limit (LOD) of 3 ± 1 µM and a sensitivity of 64 ± 2 μW mM -1 , with a correlation coefficient of 0.98 (RSD 8.1%, n = 10 electrode pairs). It exhibited robust operational stability (over 2800 s with continuous ethanol turnover) and excellent storage stability (approximately 93% of initial signal retained after 90 days). Finally, the biosensor array was integrated into a wristband and successfully evaluated for continuous alcohol abuse monitoring. This proposed system displays promising attributes for use as a flexible and wearable biosensor employing biocompatible water-based inks, offering potential applications in forensic contexts., (© 2024. The Author(s).)

The development of ultrasensitive electronic sensors for in vitro diagnostics is essential for the reliable monitoring of asymptomatic individuals before illness proliferation or progression. These platforms are increasingly valued for their potential to enable timely diagnosis and swift prognosis of infectious or progressive diseases. Typically, the responses from these analytical tools are recorded as digital signals, with electronic data offering simpler processing compared to spectral and optical data. However, preprocessing electronic data from potentiometric biosensor arrays is still in its infancy compared to more established optical technologies. This study utilized the Single-Molecule with a Large Transistor (SiMoT) array, which has achieved a Technology Readiness Level of 5, to explore the impact of data preprocessing on electronic biosensor outcomes. A dataset consisting of plasma and cyst fluid samples from 37 patients with pancreatic precursor cyst lesions was analyzed. The findings revealed that standard signal preprocessing can produce misleading conclusions due to artifacts introduced by mathematical transformations. The study offers strategies to mitigate these effects, ensuring that data interpretation remains accurate and reflective of the underlying biochemical information in the samples., (© 2024 The Author(s). ChemPlusChem published by Wiley-VCH GmbH.)

Biosensors have demonstrated versatility across numerous applications; however, their systematic optimization remains a primary obstacle, limiting their widespread adoption as dependable point-of-care tests. Experimental design, a powerful chemometric tool, offers a solution by effectively guiding the development and optimization of ultrasensitive biosensors. This perspective review provides an overview of recent applications of experimental design in the deployment of optical and electrical ultrasensitive biosensors. Various experimental designs, including full factorial, central composite, and mixture designs, are examined as systematic methodologies for optimizing biosensor fabrication, accounting for both individual variable effects and their interactions. Illustrative examples showcasing the optimization of optical and electronic biosensors through design of experiments are presented and critically analyzed. Finally, the future prospects of experimental design in the biosensor community are outlined, highlighting its potential to expedite development and bolster the performance of biosensing devices for point-of-care diagnostics, thereby facilitating their sustainable and reliable integration., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Our society largely relies on inorganic semiconductor devices which are, so far, fabricated using expensive and complex processes requiring ultra-high vacuum equipment. Here we report on the possibility of growing a p-n junction taking advantage of electrochemical processes based on the use of aqueous solutions. The growth of the junction has been carried out using the Electrochemical Atomic Layer Deposition (E-ALD) technique, which allowed to sequentially deposit two different semiconductors, CdS and Cu 2 S, on an Ag(111) substrate, in a single procedure. The growth process was monitored in situ by Surface X-Ray Diffraction (SXRD) and resulted in the fabrication of a thin double-layer structure with a high degree of crystallographic order and a well-defined interface. The high-performance electrical characteristics of the device were analysed ex-situ and show the characteristic feature of a diode., (© 2024 Wiley-VCH GmbH.)

Pancreatic cancer, ranking as the third factor in cancer-related deaths, necessitates enhanced diagnostic measures through early detection. In response, SiMoT-Single-molecule with a large Transistor multiplexing array, achieving a Technology Readiness Level of 5, is proposed for a timely identification of pancreatic cancer precursor cysts and is benchmarked against the commercially available chemiluminescent immunoassay SIMOA (Single molecule array) SP-X System. A cohort of 39 samples, comprising 33 cyst fluids and 6 blood plasma specimens, undergoes detailed examination with both technologies. The SiMoT array targets oncoproteins MUC1 and CD55, and oncogene KRAS, while the SIMOA SP-X planar technology exclusively focuses on MUC1 and CD55. Employing Principal Component Analysis (PCA) for multivariate data processing, the SiMoT array demonstrates effective discrimination of malignant/pre-invasive high-grade or potentially malignant low-grade pancreatic cysts from benign non-mucinous cysts. Conversely, PCA analysis applied to SIMOA assay reveals less effective differentiation ability among the three cyst classes. Notably, SiMoT unique capability of concurrently analyzing protein and genetic markers with the threshold of one single molecule in 0.1 mL positions it as a comprehensive and reliable diagnostic tool. The electronic response generated by the SiMoT array facilitates direct digital data communication, suggesting potential applications in the development of field-deployable liquid biopsy., (© 2024 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

Herein, a strategy to stamp laser-produced reduced graphene oxide (rGO) onto flexible polymers using only office-grade tools, namely, roll-to-roll thermal stamping, is proposed, proving for the first time its effectiveness for direct bioelectrocatalysis. This straightforward, scalable, and low-cost approach allows us to overcome the limits of the integration of laser-induced rGO-films in bioanalytical devices. Laser-produced rGO has been thermally stamped (TS) onto different polymeric substrates (PET, PVC, and EVA) using a simple roll-laminator; the obtained TS-rGO films have been compared with the native rGO (untransferred) via morphochemical and electrochemical characterization. Particularly, the direct electron transfer (DET) reaction between fructose dehydrogenase (FDH) and TS-rGO transducers has been investigated, with respect to the influence of the amount of enzyme on the catalytic process. Remarkable differences have been observed among TS-rGO transducers; PET proved to be the elective substrate to support the transfer of the laser-induced rGO, allowing the preservation of the morphochemical features of the native material and returning a reduced capacitive current. Noteworthily, TS-rGOs ensure superior electrocatalysis using a very low amount of FDH units (15 mU). Eventually, TS-rGO-based third-generation complete enzymatic biosensors were fabricated via low-cost benchtop technologies. TS-rGO PET exhibited bioanalytical performances superior to the native rGO, allowing a sensitive (0.0289 μA cm -2 μM -1 ) and reproducible (RSD = 3%, n = 3) d-fructose determination at the nanomolar level (LOD = 0.2 μM). TS-rGO exploitability as a point-of-need device was proved via the monitoring of d-fructose during banana ( Musa acuminata ) postharvest ripening, returning accurate (recoveries 110-90%; relative error -13/+1%) and reproducible (RSD ≤ 7%; n = 3) data.

While a substantial amount of research activity has been conducted in fields related to organic photonics and electronics, including the development of devices such as organic field-effect transistors, organic photovoltaics, and organic light-emitting diodes for applications encompassing organic thermoelectrics, organic batteries, excitonic organic materials for photochemical and optoelectronic applications, and organic thermoelectrics, this perspective review will primarily concentrate on the emerging and rapidly expanding domain of organic bioelectronics and neuromorphics. Here we present the most recent research findings on organic transistors capable of sensing biological biomarkers down at the single-molecule level ( i.e. , oncoproteins, genomes, etc. ) for the early diagnosis of pathological states and to mimic biological synapses, paving the way to neuromorphic applications that surpass the limitations of the traditional von Neumann computing architecture. Both organic bioelectronics and neuromorphics exhibit several challenges but will revolutionize human life, considering the development of artificial synapses to counteract neurodegenerative disorders and the development of ultrasensitive biosensors for the early diagnosis of cancer to prevent its development. Moreover, organic bioelectronics for sensing applications have also triggered the development of several wearable, flexible and stretchable biodevices for continuous biomarker monitoring.

Screening asymptomatic organisms (humans, animals, plants) with a high-diagnostic accuracy using point-of-care-testing (POCT) technologies, though still visionary holds great potential. Convenient surveillance requires easy-to-use, cost-effective, ultra-portable but highly reliable, in-vitro-diagnostic devices that are ready for use wherever they are needed. Currently, there are not yet such devices available on the market, but there are a couple more promising technologies developed at readiness-level 5: the Clustered-Regularly-Interspaced-Short-Palindromic-Repeats (CRISPR) lateral-flow-strip tests and the Single-Molecule-with-a-large-Transistor (SiMoT) bioelectronic palmar devices. They both hold key features delineated by the World-Health-Organization for POCT systems and an occurrence of false-positive and false-negative errors <1-5% resulting in diagnostic-selectivity and sensitivity >95-99%, while limit-of-detections are of few markers. CRISPR-strip is a molecular assay that, can detect down to few copies of DNA/RNA markers in blood while SiMoT immunometric and molecular test can detect down to a single oligonucleotide, protein marker, or pathogens in 0.1mL of blood, saliva, and olive-sap. These technologies can prospectively enable the systematic and reliable surveillance of asymptomatic ones prior to worsening/proliferation of illnesses allowing for timely diagnosis and swift prognosis. This could establish a proactive healthcare ecosystem that results in effective treatments for all living organisms generating diffuse and well-being at efficient costs., (© 2023 The Authors. Advanced Materials published by Wiley‐VCH GmbH.)

Herein, we describe a novel method for producing cadmium-selenide nanoparticles (CdSe NPs) with controlled size using apoferritin as a bionanoreactor triggered by local pH change at the electrode/solution interface. Apoferritin is known for its reversible self-assembly at alkaline pH. The pH change is induced electrochemically by reducing O 2 through the application of sufficiently negative voltages and bioelectrochemically through O 2 reduction catalyzed by laccase, co-immobilized with apoferritin on the electrode surface. Specifically, a Ti electrode is modified with (3-aminopropyl)triethoxysilane, followed by glutaraldehyde cross-linking (1.5% v/v in H 2 O) of apoferritin (as the bionanoreactor) and laccase (as the local pH change triggering system). This proposed platform offers a universal approach for controlling the synthesis of semiconductor NPs within a bionanoreactor solely driven by (bio)electrochemical inputs. The CdSe NPs obtained through different synthetic approaches, namely electrochemical and bioelectrochemical, were characterized spectroscopically (UV-Vis, Raman, XRD) and morphologically (TEM). Finally, we conducted online monitoring of CdSe NPs formation within the apoferritin core by integrating the electrochemical system with LWs. The quantity of CdSe NPs produced through bioelectrochemical means was determined to be 2.08 ± 0.12 mg after 90 minutes of voltage application in the presence of O 2 . TEM measurements revealed that the bioelectrochemically synthesized CdSe NPs have a diameter of 4 ± 1 nm, accounting for 85% of the size distribution, a result corroborated by XRD data. Further research is needed to explore the synthesis of nanoparticles using different biological nanoreactors, as the process can be challenging due to the elevated buffer capacitance of biological media., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Organic electrochemical transistors (OECTs) are widely employed in several bioelectronic applications such as biosensors, logic circuits, and neuromorphic engineering, providing a seamless link between the realm of biology and electronics. More specifically, OECTs are endowed with remarkable signal amplification, the ability to operate in an aqueous environment, and the effective transduction of ionic to electrical signals. One main limiting factor preventing OECTs' wide use is the need for microfabrication processes, typically requiring specialized equipment. From this perspective, a robust and cost-effective production protocol to achieve high-performing OECT would be desirable. Herein, a straightforward stencil-printed OECT fabrication procedure is proposed, where the electrical performance can be controlled by adjusting the electronic channel fabrication conditions. An experimental design approach is undertaken to optimize OECT figures of merit by varying key parameters such as the annealing temperature and time, as well as the transistor active channel length. The resulting OECT devices, fabricated through a high-yield, cost-effective, and fast stencil printing technique, feature large transconductance values at low operating voltages. The experimental design allowed for minimizing the threshold voltage ( V T = 260 mV) while keeping a high on/off ratio (7 × 10 3 ). A signal-to-noise ratio as high as 40 dB was obtained, which is among the highest for OECTs, operating in an aqueous electrolyte operated in a DC mode. An atomic force microscopy (AFM) characterization has been undertaken to analyze the channel morphology in the OECTs, correlating the annealing conditions with the charge transport properties., Competing Interests: The authors declare no conflict of interest., (This journal is © The Royal Society of Chemistry.)

Kelvin probe force microscopy (KPFM) allows the detection of single binding events between immunoglobulins (IgM, IgG) and their cognate antibodies (anti-IgM, anti-IgG). Here an insight into the reliability and robustness of the methodology is provided. Our method is based on imaging the surface potential shift occurring on a dense layer of ∼5 × 10 7 antibodies physisorbed on a 50 μm × 90 μm area when assayed with increasing concentrations of antigens in phosphate buffer saline (PBS) standard solutions, in air and at a fixed scanning location. A comprehensive investigation of the influence of the main experimental parameters that may interfere with the outcomes of KPFM immune-assay is provided, showing the robustness and reliability of our approach. The data are supported also by a thorough polarization modulation infrared reflection-absorption spectroscopy (PM-IRRAS) analysis of the physisorbed biolayer, in the spectral region of the amide I, amide II and amide A bands. Our findings demonstrate that a 10 min incubation in 500 μL PBS encompassing ≈ 30 antigens (100 zM) triggers an extended surface potential shift that involves the whole investigated area. Such a shift quickly saturates at increasing ligand concentration, showing that the developed sensing platform works as an OFF/ON detector, capable of assessing the presence of a few specific biomarkers in a given assay volume. The reliability of the developed methodology KPFM is an important asset in single molecule detections at a wide electrode interface., Competing Interests: The authors declare no conflict of interest., (This journal is © The Royal Society of Chemistry.)

A cohort of 47 patients is screened for pancreatic cancer precursors with a portable 96-well bioelectronic sensing-array for single-molecule assay in cysts fluid and blood plasma, deployable at point-of-care (POC). Pancreatic cancer precursors are mucinous cysts diagnosed with a sensitivity of at most 80% by state-of-the-art cytopathological molecular analyses (e.g., KRAS mut DNA). Adding the simultaneous assay of proteins related to malignant transformation (e.g., MUC1 and CD55) is deemed essential to enhance diagnostic accuracy. The bioelectronic array proposed here, based on single-molecule-with-a-large-transistor (SiMoT) technology, can assay both nucleic acids and proteins at the single-molecule limit-of-identification (LOI) (1% of false-positives and false-negatives). It comprises an enzyme-linked immunosorbent assay (ELISA)-like 8 × 12-array organic-electronics disposable cartridge with an electrolyte-gated organic transistor sensor array, and a reusable reader, integrating a custom Si-IC chip, operating via software installed on a USB-connected smart device. The cartridge is complemented by a 3D-printed sensing gate cover plate. KRAS mut , MUC1, and CD55 biomarkers either in plasma or cysts-fluid from 5 to 6 patients at a time, are multiplexed at single-molecule LOI in 1.5 h. The pancreatic cancer precursors are classified via a machine-learning analysis resulting in at least 96% diagnostic-sensitivity and 100% diagnostic-specificity. This preliminary study opens the way to POC liquid-biopsy-based early diagnosis of pancreatic-cancer precursors in plasma., (© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

Herein, we report a scalable benchtop electrode fabrication method to produce highly sensitive and flexible third-generation fructose dehydrogenase amperometric biosensors based on water-dispersed 0D-nanomaterials. The electrochemical platform was fabricated via Stencil-Printing (StPE) and insulated via xurography. Carbon black (CB) and mesoporous carbon (MS) were employed as 0D-nanomaterials promoting an efficient direct electron transfer (DET) between fructose dehydrogenase (FDH) and the transducer. Both nanomaterials were prepared in water-phase via a sonochemical approach. The nano-StPE exhibited enhanced electrocatalytic currents compared to conventional commercial electrodes. The enzymatic sensors were exploited for the determination of D-fructose in model solutions and various food and biological samples. StPE-CB and StPE-MS integrated biosensors showed appreciable sensitivity (∼150 μA cm -2 mM -1 ) with μmolar limit of detection (0.35 and 0.16 μM, respectively) and extended linear range (2-500 and 1-250 μM, respectively); the selectivity of the biosensors, ensured by the low working overpotential (+0.15 V), has been also demonstrated. Good accuracy (recoveries between 95 and 116%) and reproducibility (RSD ≤8.6%) were achieved for food and urine samples. The proposed approach because of manufacturing versatility and the electro-catalytic features of the water-nanostructured 0D-NMs opens new paths for affordable and customizable FDH-based bioelectronics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Antibody physisorption at a solid interface is a very interesting phenomenon that has important effects on applications such as the development of novel biomaterials and the rational design and fabrication of high-performance biosensors. The strategy selected to immobilize biorecognition elements can determine the performance level of a device and one of the simplest approaches is physical adsorption, which is cost-effective, fast, and compatible with printing techniques as well as with green-chemistry processes. Despite its huge advantages, physisorption is very seldom adopted, as there is an ingrained belief that it does not lead to high performance because of its lack of uniformity and long-term stability, which, however, have never been systematically investigated, particularly for bilayers of capture antibodies. Herein, the homogeneity and stability of an antibody layer against SARS-CoV-2-Spike1 (S1) protein physisorbed onto a gold surface have been investigated by means of multi-parametric surface plasmon resonance (MP-SPR). A surface coverage density of capture antibodies as high as (1.50 ± 0.06) × 10 12 molecules per cm -2 is measured, corresponding to a thickness of 12 ± 1 nm. This value is compatible with a single monolayer of homogeneously deposited antibodies. The effect of the ionic strength ( i s ) of the antibody solution in controlling physisorption of the protein was thoroughly investigated, demonstrating an enhancement in surface coverage at lower ionic strength. An atomic force microscopy (AFM) investigation shows a globular structure attributed to i s -related aggregations of antibodies. The long-term stability over two weeks of the physisorbed proteins was also assessed. High-performance sensing was proven by evaluating figures of merit, such as the limit of detection (2 nM) and the selectivity ratio between a negative control and the sensing experiment (0.04), which is the best reported performance for an SPR S1 protein assay. These figures of merit outmatch those measured with more sophisticated biofunctionalization procedures involving chemical bonding of the capture antibodies to the gold surface. The present study opens up interesting new pathways toward the achievement of a cost-effective and scalable biofunctionalization protocol, which could guarantee the prolonged stability of the biolayer and easy handling of the biosensing system., Competing Interests: The authors declare no conflict of interest., (This journal is © The Royal Society of Chemistry.)

In this study, a novel sensing strategy based on double sensing/actuating pathway is demonstrated, being capable to trigger the DNA-based AND gate for the sensitive and selective detection of hepatitis B virus DNA (HBV-DNA). Such an approach encompasses an enzymatic machinery logically operated using the variation of physiologically relevant biomarkers for liver dysfunctions. Alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) are used as inputs of an AND gate generating an output signal, namely lactate. In particular, lactate is oxidized back to pyruvate at the anodic electrode by lactate oxidase connected in mediated electron transfer through ferrocene moieties (creating an amplifying recycling mechanism). The anodic electrode is further connected with a Myrothecium verrucaria bilirubin oxidase (MvBOx) based biocathode modified with SiO 2 nanoparticles (SiO 2 NPs) functionalized with phenyl boronic acid and trigonelline, triggering the release of quenching DNA (qDNA) upon local pH change at the electrode surface (notably, modified SiONPs gets negatively recharged upon local pH gradient releasing negatively charged DNA). Next, the released qDNA labeled with BHQ2 and detecting DNA (dDNA, labeled with FAM) are detecting HBV-DNA. The proposed biosensor can discriminate between the absence and presence of HBV-DNA setting the threshold at 0.05 fM in model buffer solutions and 1 fM in human serum. This enzymatic/DNA logic network can be of particular interest for future biomedical applications (e.g., early detection of liver cancer disease etc.). In the future development this technology could be easily integrated with a smartphone camera, allowing more user-friendly applications., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The development of ultrasensitive analytical detection methods for organophosphorus pesticides such as dimethoate (DMT) plays a key role in healthy food production. DMT is an inhibitor of acetylcholinesterase (AChE), which can lead to the accumulation of acetylcholine and result in symptoms related to the autonomous and central nervous systems. Herein, we report the first spectroscopic and electrochemical study on template removal after an imprinting process from a polypyrrole-based molecularly imprinted polymer (PPy-MIP) film for the detection of DMT. Several template removal procedures were tested and evaluated using X-ray photoelectron spectroscopy. The most effective procedure was achieved in 100 mM NaOH. The proposed DMT PPy-MIP sensor exhibits a limit of detection of (8 ± 2) × 10 -12 M.

Pathogens ultra-sensitive detection is vital for early diagnosis and provision of restraining actions and/or treatments. Among plant pathogens, Xylella fastidiosa is among the most threatening as it can infect hundreds of plant species worldwide with consequences on agriculture and the environment. An electrolyte-gated transistor is here demonstrated to detect X. fastidiosa at a limit-of-quantification (LOQ) of 2 ± 1 bacteria in 0.1 mL (20 colony-forming-unit per mL). The assay is carried out with a millimeter-wide gate functionalized with Xylella-capturing antibodies directly in saps recovered from naturally infected plants. The proposed platform is benchmarked against the quantitave polymerase chain reaction (qPCR) gold standard, whose LOQ turns out to be at least one order of magnitude higher. Furthermore, the assay selectivity is proven against the Paraburkholderia phytofirmans bacterium (negative-control experiment). The proposed label-free, fast (30 min), and precise (false-negatives, false-positives below 1%) electronic assay, lays the ground for an ultra-high performing immunometric point-of-care platform potentially enabling large-scale screening of asymptomatic plants., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Molecular tests are highly reliable and sensitive but lack portability and are not simple to use; conversely, easy-to-use antigenic tests still lack high performance. BioScreen combines single-molecule sensitivity and outstanding reliability with ultraportability and simplicity of use. This digital platform is capable of artificial intelligence-based binary classification at the limit of identification of a single marker/virus in 0.1 ml. The diagnostic sensitivity, specificity, and accuracy reach 99.2% as validated through 240 assays, including a pilot clinical trial. The versatile immunometric system can detect the SARS-CoV-2 virus, spike S1, and immunoglobulin G antigen proteins in saliva, blood serum, and swab. BioScreen has a small footprint comprising a disposable cartridge and a handheld electronic reader connected to a smart device. The sample handling is minimal, and the assay time to result is 21 min. Reliable and sensitive self-testing with an ultraportable and easy-to-use diagnostic system operated directly by a patient holds the potential to revolutionize point-of-care testing and early diagnosis.

Early diagnosis in a premalignant (or pre-invasive) state represents the only chance for cure in neoplastic diseases such as pancreatic-biliary cancer, which are otherwise detected at later stages and can only be treated using palliative approaches, with no hope for a cure. Screening methods for the purpose of secondary prevention are not yet available for these cancers. Current diagnostic methods mostly rely on imaging techniques and conventional cytopathology, but they do not display adequate sensitivity to allow valid early diagnosis. Next-generation sequencing can be used to detect DNA markers down to the physical limit; however, this assay requires labeling and is time-consuming. The additional determination of a protein marker that is a predictor of aggressive behavior is a promising innovative approach, which holds the potential to improve diagnostic accuracy. Moreover, the possibility to detect biomarkers in blood serum offers the advantage of a noninvasive diagnosis. In this study, both the DNA and protein markers of pancreatic mucinous cysts were analyzed in human blood serum down to the single-molecule limit using the SiMoT (single-molecule assay with a large transistor) platform. The SiMoT device proposed herein, which exploits an inkjet-printed organic semiconductor on plastic foil, comprises an innovative 3D-printed sensing gate module, consisting of a truncated cone that protrudes from a plastic substrate and is compatible with standard ELISA wells. This 3D gate concept adds tremendous control over the biosensing system stability, along with minimal consumption of the capturing molecules and body fluid samples. The 3D sensing gate modules were extensively characterized from both a material and electrical perspective, successfully proving their suitability as detection interfaces for biosensing applications. KRAS and MUC1 target molecules were successfully analyzed in diluted human blood serum with the 3D sensing gate functionalized with b-KRAS and anti-MUC1, achieving a limit of detection of 10 zM and 40 zM, respectively. These limits of detection correspond to (1 ± 1) KRAS and (2 ± 1) MUC1 molecules in the 100 μL serum sample volume. This study provides a promising application of the 3D SiMoT platform, potentially facilitating the timely, noninvasive, and reliable identification of pancreatic cancer precursor cysts., (© 2022. The Author(s).)

Single-molecule detection at a nanometric interface in a femtomolar solution, can take weeks as the encounter rate between the diffusing molecule to be detected and the transducing nanodevice is negligibly small. On the other hand, several experiments prove that macroscopic label-free sensors based on field-effect-transistors, engaging micrometric or millimetric detecting interfaces are capable to assay a single-molecule in a large volume within few minutes. The present work demonstrates why at least a single molecule out of a few diffusing in a 100 µL volume has a high probability to hit a large capturing and detecting electronic interface. To this end, sensing data, measured with an electrolyte-gated FET whose gate is functionalized with 10 12 capturing anti-immunoglobulin G, are here provided along with a Brownian diffusion-based modeling. The EG-FET assays solutions down to some tens of zM in concentrations with volumes ranging from 25 µL to 1 mL in which the functionalized gates are incubated for times ranging from 30 s to 20 min. The high level of accordance between the experimental data and a model based on the Einstein's diffusion-theory proves how the single-molecule detection process at large-capturing interfaces is controlled by Brownian diffusion and yet is highly probable and fast., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Herein, we report a combined strategy encompassing electrochemical and X-ray photoelectron spectroscopy (XPS) experiments to investigate self-assembled monolayer (SAM) conformational reorganization onto an electrode surface due to the application of an electrical field. In particular, 3-mercaptopriopionic acid SAM (3MPA SAM) modified gold electrodes are activated with a 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N -hydroxysulfosuccinimide (NHSS) (EDC-NHSS) mixture by shortening the activation time, from 2 h to 15/20 min, labelled as Protocol-A, -B and -C, respectively. This step, later followed by a deactivation process with ethanolamine (EA), plays a key role in the reaction yields (formation of N -(2-hydroxyethyl)-3-mercaptopropanamide, NMPA) but also in the conformational rearrangement observed during the application of the electrical field. This study aims at explaining the high performance (i.e., single-molecule detection at a large electrode interface) of bioelectronic devices, where the 3MPA-based SAM structure is pivotal in achieving extremely high sensing performance levels due to its interchain interaction. Cyclic voltammetry (CV) experiments performed in K 4 Fe(CN) 6 :K 3 Fe(CN) 6 for 3MPA SAMs that are activated/deactivated show similar trends of anodic peak current (I A ) over time, mainly related to the presence of interchain hydrogen bonds, driving the conformational rearrangements (tightening of SAMs structure) while applying an electrical field. In addition, XPS analysis allows correlation of the deactivation yield with electrochemical data (conformational rearrangements), identifying the best protocol in terms of high reaction yield, mainly related to the shorter reaction time, and not triggering any side reactions. Finally, Protocol-C's SAM surface coverage, determined by CV in H 2 SO 4 and differential pulse voltammetry (DPV) in NaOH, was 1.29 * 10 13 molecules cm -2 , being similar to the bioreceptor surface coverage in single-molecule detection at a large electrode interface.

Bioelectronic transducing surfaces that are nanometric in size have been the main route to detect single molecules. Though enabling the study of rarer events, such methodologies are not suited to assay at concentrations below the nanomolar level. Bioelectronic field-effect-transistors with a wide (μm 2 -mm 2 ) transducing interface are also assumed to be not suited, because the molecule to be detected is orders of magnitude smaller than the transducing surface. Indeed, it is like seeing changes on the surface of a one-kilometer-wide pond when a droplet of water falls on it. However, it is a fact that a number of large-area transistors have been shown to detect at a limit of detection lower than femtomolar; they are also fast and hence innately suitable for point-of-care applications. This review critically discusses key elements, such as sensing materials, FET-structures, and target molecules that can be selectively assayed. The amplification effects enabling extremely sensitive large-area bioelectronic sensing are also addressed.

The early detection of the human immunodeficiency virus (HIV) is of paramount importance to achieve efficient therapeutic treatment and limit the disease spreading. In this perspective, the assessment of biosensing assay for the HIV-1 p24 capsid protein plays a pivotal role in the timely and selective detection of HIV infections. In this study, multi-parameter-SPR has been used to develop a reliable and label-free detection method for HIV-1 p24 protein. Remarkably, both physical and chemical immobilization of mouse monoclonal antibodies against HIV-1 p24 on the SPR gold detecting surface have been characterized for the first time. The two immobilization techniques returned a capturing antibody surface coverage as high as (7.5 ± 0.3) × 10 11 molecule/cm 2 and (2.4 ± 0.6) × 10 11 molecule/cm 2 , respectively. However, the covalent binding of the capturing antibodies through a mixed self-assembled monolayer (SAM) of alkanethiols led to a doubling of the p24 binding signal. Moreover, from the modeling of the dose-response curve, an equilibrium dissociation constant K D of 5.30 × 10 -9 M was computed for the assay performed on the SAM modified surface compared to a much larger K D of 7.46 × 10 -5 M extracted for the physisorbed antibodies. The chemically modified system was also characterized in terms of sensitivity and selectivity, reaching a limit of detection of (4.1 ± 0.5) nM and an unprecedented selectivity ratio of 0.02.

Electrolyte-gated transistors (EGTs), capable of transducing biological and biochemical inputs into amplified electronic signals and stably operating in aqueous environments, have emerged as fundamental building blocks in bioelectronics. In this Primer, the different EGT architectures are described with the fundamental mechanisms underpinning their functional operation, providing insight into key experiments including necessary data analysis and validation. Several organic and inorganic materials used in the EGT structures and the different fabrication approaches for an optimal experimental design are presented and compared. The functional bio-layers and/or biosystems integrated into or interfaced to EGTs, including self-organization and self-assembly strategies, are reviewed. Relevant and promising applications are discussed, including two-dimensional and three-dimensional cell monitoring, ultra-sensitive biosensors, electrophysiology, synaptic and neuromorphic bio-interfaces, prosthetics and robotics. Advantages, limitations and possible optimizations are also surveyed. Finally, current issues and future directions for further developments and applications are discussed., Competing Interests: Competing interests The authors declare no competing interests.

Single-molecule sensing is becoming a major driver in biomarker assays as it is foreseen to enable precision medicine to enter into everyday clinical practice. However, among the single-molecule detection methods proposed so far, only a few are fully exploitable for the ultrasensitive label-free assay of biofluids. Firstly introduced single-molecule sensing platforms encompass low-background-noise fluorescent microscopy as well as plasmonic and electrical nanotransducers; these are generally able to sense at the nanomolar concentration level or higher. Label-based single-molecule technologies relying on optical transduction and microbeads that can scavenge and detect a few biomarkers in the bulk of real biofluids, reaching ultralow detection limits, have been recently commercialized. These assays, thanks to the extremely high sensitivity and convenient handling, are new trends in the field as they are paving the way to a revolution in early diagnostics. Very recently, another new trend is the label-free, organic bioelectronic electrolyte-gated large transistors that can potentially be produced by means of large-area low-cost technologies and have been proven capable to detect a protein at the physical limit in real bovine serum. This article offers a bird's-eye view on some of the more significant single-molecule bioanalytical technologies and highlights their sensing principles and figures-of-merit such as limit of detection, need for a labelling step, and possibility to operate, also as an array, directly in real biofluids. We also discuss the new trend towards single-molecule proof-of-principle extremely sensitive technologies that can detect a protein at the zeptomolar concentration level involving label-free devices that potentially offer low-cost production and easy scalability.

Thiolated self-assembled monolayers (SAMs) are typically used to anchor on a gold surface biomolecules serving as recognition elements for biosensor applications. Here, the design and synthesis of N -(2-hydroxyethyl)-3-mercaptopropanamide (NMPA) in biotinylated mixed SAMs is proposed as an alternative strategy with respect to on-site multistep functionalization of SAMs prepared from solutions of commercially available thiols. In this study, the mixed SAM deposited from a 10:1 solution of 3-mercaptopropionic acid (3MPA) and 11-mercaptoundecanoic acid (11MUA) is compared to that resulting from a 10:1 solution of NMPA:11MUA. To this end, surface plasmon resonance (SPR) and attenuated total reflectance infrared (ATR-IR) experiments have been carried out on both mixed SAMs after biotinylation. The study demonstrated how the fine tuning of the SAM features impacts directly on both the biofunctionalization steps, i.e., the biotin anchoring, and the biorecognition properties evaluated upon exposure to streptavidin analyte. Higher affinity for the target analyte with reduced nonspecific binding and lower detection limit has been demonstrated when NMPA is chosen as the more abundant starting thiol. Molecular dynamics simulations complemented the experimental findings providing a molecular rationale behind the performance of the biotinylated mixed SAMs. The present study confirms the importance of the functionalization design for the development of a highly performing biosensor., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

The increasing interest in technologies capable of tracking a biomarker down to the physical limit points toward new opportunities in early diagnostics of progressive diseases. Indeed, single-molecule detection technologies are foreseen to enable clinicians to associate the tiniest increase in a biomarker with the progression of a disease, particularly at its early stage. Bioelectronic organic transistors represent an extremely powerful tool to achieve label-free and single-molecule detection of clinically relevant biomarkers. These electronic devices are millimetric in size and in the future could be mass-produced at low cost. The core of the single molecule with a large transistor (SiMoT) platform, based on an electrolyte-gated field-effect transistor, is a gold gate electrode biofunctionalized with a self-assembled monolayer, a densely packed layer of recognition elements. So far, only the SiMoT detection of proteins, using the corresponding antibodies as recognition elements, has been reported. In this study, the SiMoT sensing response toward genomic biomarkers is proposed. Herein, the gate is functionalized with a genomic biomarker for multiple sclerosis (miR-182). This is relevant, not only because a limit of detection of a single molecule is achieved but also because it proves that the SiMoT label-free, single-molecule detection principle is the only one of its kind that can detect, by means of the same platform, both protein and genomic markers.

A point-of-care (POC) device to enable de-centralized diagnostics can effectively reduce the time to treatment, especially in case of infectious diseases. However, many of the POC solutions presented so far do not comply with the ASSURED (affordable, sensitive, specific, user-friendly, rapid and robust, equipment free, and deliverable to users) guidelines that are needed to ensure their on-field deployment. Herein, we present the proof of concept of a self-powered platform that operates using the analysed fluid, mimicking a blood sample, for early stage detection of HIV-1 infection. The platform contains a smart interfacing circuit to operate an ultra-sensitive electrolyte-gated field-effect transistor (EGOFET) as a sensor and facilitates an easy and affordable readout mechanism. The sensor transduces the bio-recognition event taking place at the gate electrode functionalized with the antibody against the HIV-1 p24 capsid protein, while it is powered via paper-based biofuel cell (BFC) that extracts the energy from the analysed sample itself. The self-powered platform is demonstrated to achieve detection of HIV-1 p24 antigens in fM range, suitable for early diagnosis. From these developments, a cost-effective digital POC device able to detect the transition from "healthy" to "infected" state at single-molecule precision, with no dependency on external power sources while using minimal components and simpler approach, is foreseen., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Zinc oxide (ZnO) nanostructures are widely applied materials, and are also capable of antimicrobial action. They can be obtained by several methods, which include physical and chemical approaches. Considering the recent rise of green and low-cost synthetic routes for nanomaterial development, electrochemical techniques represent a valid alternative to biogenic synthesis. Following a hybrid electrochemical-thermal method modified by our group, here we report on the aqueous electrosynthesis of ZnO nanomaterials based on the use of alternative stabilizers. We tested both benzyl-hexadecyl-dimetylammonium chloride (BAC) and poly-diallyl-(dimethylammonium) chloride (PDDA). Transmission electron microscopy images showed the formation of rod-like and flower-like structures with a variable aspect-ratio. The combination of UV-Vis, FTIR and XPS spectroscopies allowed for the univocal assessment of the material composition as a function of different thermal treatments. In fact, the latter guaranteed the complete conversion of the as-prepared colloidal materials into stoichiometric ZnO species without excessive morphological modification. The antimicrobial efficacy of both materials was tested against Bacillus subtilis as a Gram-positive model microorganism.

Early diagnosis of the infection caused by human immunodeficiency virus type-1 (HIV-1) is vital to achieve efficient therapeutic treatment and limit the disease spreading when the viremia is at its highest level. To this end, a point-of-care HIV-1 detection carried out with label-free, low-cost, and ultra-sensitive screening technologies would be of great relevance. Herein, a label-free single molecule detection of HIV-1 p24 capsid protein with a large (wide-field) single-molecule transistor (SiMoT) sensor is proposed. The system is based on an electrolyte-gated field-effect transistor whose gate is bio-functionalized with the antibody against the HIV-1 p24 capsid protein. The device exhibits a limit of detection of a single protein and a limit of quantification in the 10 molecule range. This study paves the way for a low-cost technology that can quantify, with single-molecule precision, the transition of a biological organism from being "healthy" to being "diseased" by tracking a target biomarker. This can open to the possibility of performing the earliest possible diagnosis.

There is an increasing interest for low cost, ultrasensitive, time saving yet reliable, point-of-care bioelectronic sensors. Electrolyte gated organic field effect transistors (EGOFETs) are proven compelling transducers for various sensing applications, offering direct electronic, label-free transduction of bio-recognition events along with miniaturization, fast data handling and processing. Given that field effect transistors act as intrinsically signal amplifiers, even a small change of a chemical or biological quantity may significantly alter the output electronic signal. In EGOFETs selectivity can be guaranteed by the immobilization of bioreceptors able to bind specifically a target analyte. The layer of receptors can be linked to one of the electronic active interfaces of the transistor, and the interactions with a target molecule affect the electronic properties of the device. The present chapter discusses main aspects of EGOFETs transducers along with detailed examples of how to tailor the device interfaces with desired functionality. The development of an "electronic tongue" based on an EGOFET device coupled to odorant binding proteins (OBPs) for enantiomers differentiation is presented., (© 2020 Elsevier Inc. All rights reserved.)

The rapid spreading of resistance among common bacterial pathogens towards the misused antibiotics/disinfectant agents has drawn much attention worldwide to bacterial infections. In light of this, the present work aimed at the realization of core-shell nanoparticles possessing remarkable antimicrobial properties thanks to the synergistic action of the metal core and the disinfectant shell. Copper nanoparticles stabilized by benzalkonium chloride were prepared, characterized, and implemented in poly-vinyl-methyl ketone to obtain nanoantimicrobial composite coatings. Bioactivity tests are reported, proving the excellent disinfectant properties of the proposed nanomaterials, as compared to one of the well-known and strongest silver-based nanoantimicrobials. Applications are also briefly described.

Robust electrolyte-gated organic field-effect-transistors (OFETs) are particularly needed for the development of biosensing devices. However, when a FET biosensor operates in aqueous environments or even in real biological fluids, some critical issues may arise due to the possible lack of environmental long-term and/or operational stability. An important source of instability is associated with the degradation of the organic electronic channel materials such as for instance, poly-3-hexylthiophene (P3HT), a benchmark commercially available p-type organic semiconductor. In this work, the investigation of critical parameters, such as the control over spurious electrochemical phenomena as well as the operating conditions that can affect water-gated OFETs lifetime, is reported, together with a proposed modeling of the P3HT stability curve over 1 week in water. The investigation of possible morphological/chemical modifications occurring at the polymer surface after operating in water for 2 weeks was carried out. Moreover, it is proven how the addition of a gel layer can extend the P3HT based water-gated OFET shelf life up to 2 months., (Copyright © 2019 Picca, Manoli, Macchia, Tricase, Di Franco, Scamarcio, Cioffi and Torsi.)

In the last decade, saliva has been suggested as non-invasive diagnostic fluid, suitable for clinical use alternatively to blood serum and plasma. However, the clinical applicability of saliva has been hampered so far by the inadequate sensitivity of current methods to detect the lower salivary concentrations of many biomarkers monitored in blood products. Herein, a label-free biosensor based on electrolyte-gated organic thin-film transistor (EGOTFT) has been developed for the detection at the physical limit of C-reactive protein (CRP) in human saliva. CRP is a key relevant biomarker for inflammatory processes and is routinely monitored for many clinical purposes. Herein, an electrolyte-gated thin-film transistor (EGOTFT) has been proposed as a transducer of the biorecognition event taking place at the gate electrode, functionalized with a self-assembled monolayer (SAM) of highly densely packed capturing anti-CRP proteins. Thanks to the SAM, the biosensing platform herein proposed is endowed with ultra-high sensitivity, along with an extremely high selectivity, assessed by measuring the dose curves of CRP interacting with a bovine serum albumin-functionalized gate. Moreover, the biosensing platform is compatible with low-cost fabrication techniques and applicable to the ultra-sensitive detection of a plethora of clinically relevant biomarkers. Therefore, the EGOTFT device herein proposed, being able to operate in physiologically relevant fluids such as saliva, will set the ground to a major revolution in biosensing applications for early clinical detection.

Label-free single-molecule detection has been achieved so far by funnelling a large number of ligands into a sequence of single-binding events with few recognition elements host on nanometric transducers. Such approaches are inherently unable to sense a cue in a bulk milieu. Conceptualizing cells' ability to sense at the physical limit by means of highly-packed recognition elements, a millimetric sized field-effect-transistor is used to detect a single molecule. To this end, the gate is bio-functionalized with a self-assembled-monolayer of 10 12 capturing anti-Immunoglobulin-G and is endowed with a hydrogen-bonding network enabling cooperative interactions. The selective and label-free single molecule IgG detection is strikingly demonstrated in diluted saliva while 15 IgGs are assayed in whole serum. The suggested sensing mechanism, triggered by the affinity binding event, involves a work-function change that is assumed to propagate in the gating-field through the electrostatic hydrogen-bonding network. The proposed immunoassay platform is general and can revolutionize the current approach to protein detection.

Herein a label-free immunosensor based on electrolyte-gated organic field-effect transistor (EGOFET) was developed for the detection of procalcitonin (PCT), a sepsis marker. Antibodies specific to PCT were immobilized on the poly-3-hexylthiophene (P3HT) organic semiconductor surface through direct physical adsorption followed by a post-treatment with bovine serum albumin (BSA) which served as the blocking agent to prevent non-specific adsorption. Antibodies together with BSA (forming the whole biorecognition layer) served to selectively capture the procalcitonin target analyte. The entire immunosensor fabrication process was fast, requiring overall 45min to be completed before analyte sensing. The EGOFET immunosensor showed excellent electrical properties, comparable to those of bare P3HT based EGOFET confirming reliable biosensing with bio-functional EGOFET immunosensor. The detection limit of the immunosensor was as low as 2.2pM and within a range of clinical relevance. The relative standard deviation of the individual calibration data points, measured on immunosensors fabricated on different chips (reproducibility error) was below 7%. The developed immunosensor showed high selectivity to the PCT analyte which was evident through control experiments. This report of PCT detection is first of its kind among the electronic sensors based on EGOFETs. The developed sensor is versatile and compatible with low-cost fabrication techniques., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Bioconjugated gold surfaces constitute interesting platforms for biosensing applications. The immobilization of antibodies such as anti-immunoglobulin G and M (anti-IgG and anti-IgM) on gold electrodes via self-assembled monolayers (SAMs) is here studied as a model system for further immunoassays development. The biolayer is characterized by means of X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), a dedicated thin-film transistor (TFT)-based platform and electrochemical surface plasmon resonance (EC-SPR). XPS analysis confirms the presence of all the chemical species involved in the fabrication process as well as the covalent attachment of the antibodies with high reproducibility. Visualization of the biolayer topography by AFM shows nanostructures with a thickness consistent with the actual size of the protein, which is also verified by SPR measurements. EC-SPR allows taking advantage of complementary electrochemical and optical signals during the functionalization steps. Moreover, the functionalization of gold leads to a change in the work function, which is demonstrated in an electrolyte gated thin-film transistor configuration. Such configuration enables also to evaluate the electrostatic changes occurring on the gate that are connected with the threshold voltage shifts. The data support that functional biomodified gold surfaces can be reproducibly prepared, which is a prerequisite for further biosensor development., (© 2017 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Electrical double layer (EDL) thin film transistors (TFTs) are an interesting class of transistors that use an electrolyte as the gating medium. Recently it has been demonstrated that pure organic solvents can also be used as gating media for TFTs without the addition of exogenous electrolytes. Here we present a systematic study of the performances of TFTs based on two different semiconductors (P3HT and ZnO) gated through nine different solvents either pure or loaded with NaCl. The nature of the solvent impacts the transfer characteristics of the TFT through a change in the gating capacitance while the threshold voltage remains unaffected. Depending on the polarity of solvents, addition of NaCl gives rise to different responses. TFTs gated through highly polar solvents are unaffected by the salt concentration while for low polarity solvents the output current increases with salt up to a plateau. Furthermore, when the semiconductor surface is covered with a high capacitance thin dielectric layer, the TFT output current becomes dependent on the NaCl concentration also for high polarity solvents. This phenomenology was rationalized considering the different contributions of Helmholtz and Guy-Chapman EDLs to the capacitance and the dielectric saturation that decreases the solvent dielectric constant within the Helmholtz EDL.

Multiwalled carbon nanotube (MWCNT)-based chemiresistors were electrochemically decorated with Au and Pd nanoparticles (NPs), resulting in an improvement in the detection of gaseous pollutants as compared to sensors based on pristine MWCNTs. Electrophoresis was used to decorate MWCNTs with preformed Au or Pd NPs, thus preserving their nanometer-sized dimensions and allowing the metal content to be tuned by simply varying the deposition time. The sensing response of unmodified and metal-decorated MWCNTs was evaluated towards different gaseous pollutants (e.g., NO 2 , H 2 S, NH 3 and C 4 H 10 ) at a wide range of concentrations in the operating temperature range of 45-200 °C. The gas sensing results were related to the presence, type and loading of metal NPs used in the MWCNT functionalization. Compared to pristine MWCNTs, metal-decorated MWCNTs revealed a higher gas sensitivity, a faster response, a better stability, reversibility and repeatability, and a low detection limit, where all of these sensing properties were controlled by the type and loading of the deposited metal catalytic NPs. Specifically, in the NO 2 gas sensing experiments, MWCNTs decorated with the lowest Au content revealed the highest sensitivity at 150 °C, while MWCNTs with the highest Pd loading showed the highest sensitivity when operated at 100 °C. Finally, considering the reported gas sensing results, sensing mechanisms have been proposed, correlating the chemical composition and gas sensing responses.

Pristine and electrochemically Pd-modified ZnO nanorods (ZnO NRs) were proposed as active sensing layers in chemiresistive gas sensors for hydrocarbon (HC) gas detection (e.g., CH 4 , C 3 H 8, C 4 H 10 ). The presence of Pd nanoparticles (NPs) on the surface of ZnO NRs, obtained after the thermal treatment at 550 °C, was revealed by morphological and surface chemical analyses, using scanning electron microscopy and X-ray photoelectron spectroscopy, respectively. The effect of the Pd catalyst on the performance of the ZnO-based gas sensor was evaluated by comparing the sensing results with those of pristine ZnO NRs, at an operating temperature of 300 °C and for various HC gas concentrations in the range of 30-1000 ppm. The Pd-modified ZnO NRs showed a higher selectivity and sensitivity compared to pristine ZnO NRs. The mean sensitivity of Pd-modified ZnO NRs towards the analyzed HCs gases increased with the length of the hydrocarbon chain of the target gas molecule. Finally, the evaluation of the selectivity revealed that the presence or the absence of metal nanoparticles on ZnO NRs improves the selectivity in the detection of specific HCs gaseous molecules.