Synergy of Nanotopography and Electrical Conductivity of PEDOT/PSS for Enhanced Neuronal Development.

Biomaterials able to promote neuronal development and neurite outgrowth are highly desired in neural tissue engineering for the repair of damaged or disrupted neural tissue and restoring the axonal connection. For this purpose, the use of either electroactive or micro- and nanostructured materials has been separately investigated. Here, the use of a nanomodulated conductive poly(3,4-ethylendioxithiophene) poly(styrenesulfonate) (PEDOT/PSS) substrate that exhibits instructive topographical and electrical cues at the same time was investigated for the first time. In particular, thin films featuring grooves with sizes comparable with those of neuronal neurites (NanoPEDOT) were fabricated by electrochemical polymerization of PEDOT/PSS on a nanomodulated polycarbonate template. The ability of NanoPEDOT to support neuronal development and direct neurite outgrowth was demonstrated by assessing cell viability and proliferation, expression of neuronal markers, average neurite length, and direction of neuroblastoma N2A cells induced to differentiate on this novel support. In addition to the beneficial effect of the nanogrooved topography, a 30% increase was shown in the average length of neurites when differentiating cells were subjected to an electrical stimulation of a few microamperes for 6 h. The results reported here suggest a favorable effect on the neuronal development of the synergistic combination of nanotopography and electrical stimulation, supporting the use of NanoPEDOT in neural tissue engineering to promote physical and functional reconnection of impaired neural networks.

Organic Electrochemical Transistor Aptasensor for Interleukin-6 Detection.

We demonstrate an organic electrochemical transistor (OECT) biosensor for the detection of interleukin 6 (IL6), an important biomarker associated with various pathological processes, including chronic inflammation, inflammaging, cancer, and severe COVID-19 infection. The biosensor is functionalized with oligonucleotide aptamers engineered to bind specifically IL6. We developed an easy functionalization strategy based on gold nanoparticles deposited onto a poly(3,4-ethylenedioxythiophene) doped with polystyrenesulfonate (PEDOT:PSS) gate electrode for the subsequent electrodeposition of thiolated aptamers. During this functionalization step, the reduction of sulfide bonds allows for simultaneous deposition of a blocking agent. A detection range from picomolar to nanomolar concentrations for IL6 was achieved, and the selectivity of the device was assessed against Tumor Necrosis Factor (TNF), another cytokine involved in the inflammatory processes.

Reduced Graphene Oxide Electrolyte-Gated Transistor Immunosensor with Highly Selective Multiparametric Detection of Anti-Drug Antibodies.

The advent of immunotherapies with biological drugs has revolutionized the treatment of cancers and auto-immune diseases. However, in some patients, the production of anti-drug antibodies (ADAs) hampers the drug efficacy. The concentration of ADAs is typically in the range of 1-10 pm; hence their immunodetection is challenging. ADAs toward Infliximab (IFX), a drug used to treat rheumatoid arthritis and other auto-immune diseases, are focussed. An ambipolar electrolyte-gated transistor (EGT) immunosensor is reported based on a reduced graphene oxide (rGO) channel and IFX bound to the gate electrode as the specific probe. The rGO-EGTs are easy to fabricate and exhibit low voltage operations (≤ 0.3 V), a robust response within 15 min, and ultra-high sensitivity (10 am limit of detection). A multiparametric analysis of the whole rGO-EGT transfer curves based on the type-I generalized extreme value distribution is proposed. It is demonstrated that it allows to selectively quantify ADAs also in the co-presence of its antagonist tumor necrosis factor alpha (TNF-α), the natural circulating target of IFX.

Robust Biosensor Based on Carbon Nanotubes/Protein Hybrid Electrolyte Gated Transistors.

Semiconducting single walled carbon nanotubes (SWCNTs) are promising materials for biosensing applications with electrolyte-gated transistors (EGT). However, to be employed in EGT devices, SWCNTs often require lengthy solution-processing fabrication techniques. Here, we introduce a simple solution-based method that allows fabricating EGT devices from stable dispersions of SWCNTs/bovine serum albumin (BSA) hybrids in water. The dispersion is then deposited on a substrate allowing the formation of a SWCNTs random network as the semiconducting channel. We demonstrate that this methodology allows the fabrication of EGT devices with electric performances that allow their use in biosensing applications. We demonstrate their application for the detection of cortisol in solution, upon gate electrode functionalization with anti-cortisol antibodies. This is a robust and cost-effective methodology that sets the ground for a SWCNT/BSA-based biosensing platform that allows overcoming many limitations of standard SWCNTs biosensor fabrications.

In operando nanomechanical mapping of PEDOT:PSS thin films in electrolyte solutions with bimodal AFM.

Understanding the interplay between the nanomechanical properties of organic electronic materials and their electronic properties is central to developing sensors and transducers for applications ranging from immunosensing to e-skin. Controlling organic device operations in aqueous electrolyte solutions and their mechanical compliance with the host tissue or living systems, as for instance in active implants for the recording or stimulation of neural signals, is still largely unexplored. Here, we implemented bimodal AFM to map the nanomechanical and structural properties of thin films made from poly(3,4-ethylenedioxythiophene):poly(styrene-sulfonate) (PEDOT:PSS), the most widely used conducting polymer blend, during operation as a microelectrode in an electrolyte solution. Nanomechanical maps showed the film consisting of a granular structure made from PEDOT:PSS regions embedded in the PSS matrix. The film swelled upon immersion in an aqueous solution. In operando bimodal AFM data obtained by applying a sequence of doping/de-doping bias cycles showed a significant decrease in the modulus (70%) that saturated after about 10 cycles. A similar sequence of biases at the opposite polarity did not significantly influence the mechanical behaviour of PEDOT:PSS. The decrease in the modulus was explained by the development of persistent hydration, which was enhanced by the cations trapped inside the organic electronic material.

PEDOT: PSS promotes neurogenic commitment of neural crest-derived stem cells.

Poly (3,4-ethylendioxythiophene) polystyrene sulphonate (PEDOT:PSS) is the workhorse of organic bioelectronics and is steadily gaining interest also in tissue engineering due to the opportunity to endow traditional biomaterials for scaffolds with conductive properties. Biomaterials capable of promoting neural stem cell differentiation by application of suitable electrical stimulation protocols are highly desirable in neural tissue engineering. In this study, we evaluated the adhesion, proliferation, maintenance of neural crest stemness markers and neurogenic commitment of neural crest-derived human dental pulp stem cells (hDPSCs) cultured on PEDOT:PSS nanostructured thin films deposited either by spin coating (SC-PEDOT) or by electropolymerization (ED-PEDOT). In addition, we evaluated the immunomodulatory properties of hDPSCs on PEDOT:PSS by investigating the expression and maintenance of the Fas ligand (FasL). We found that both SC-PEDOT and ED-PEDOT thin films supported hDPSCs adhesion and proliferation; however, the number of cells on the ED-PEDOT after 1 week of culture was significantly higher than that on SC-PEDOT. To be noted, both PEDOT:PSS films did not affect the stemness phenotype of hDPSCs, as indicated by the maintenance of the neural crest markers Nestin and SOX10. Interestingly, neurogenic induction was clearly promoted on ED-PEDOT, as indicated by the strong expression of MAP-2 and ß -Tubulin-III as well as evident cytoskeletal reorganisation and appreciable morphology shift towards a neuronal-like shape. In addition, strong FasL expression was detected on both undifferentiated or undergoing neurogenic commitment hDPSCs, suggesting that ED-PEDOT supports the expression and maintenance of FasL under both expansion and differentiation conditions.

Poly(3,4-ethylenedioxythiophene)-Based Neural Interfaces for Recording and Stimulation: Fundamental Aspects and In Vivo Applications.

Next-generation neural interfaces for bidirectional communication with the central nervous system aim to achieve the intimate integration with the neural tissue with minimal neuroinflammatory response, high spatio-temporal resolution, very high sensitivity, and readout stability. The design and manufacturing of devices for low power/low noise neural recording and safe and energy-efficient stimulation that are, at the same time, conformable to the brain, with matched mechanical properties and biocompatibility, is a convergence area of research where neuroscientists, materials scientists, and nanotechnologists operate synergically. The biotic-abiotic neural interface, however, remains a formidable challenge that prompts for new materials platforms and innovation in device layouts. Conductive polymers (CP) are attractive materials to be interfaced with the neural tissue and to be used as sensing/stimulating electrodes because of their mixed ionic-electronic conductivity, their low contact impedance, high charge storage capacitance, chemical versatility, and biocompatibility. This manuscript reviews the state-of-the-art of poly(3,4-ethylenedioxythiophene)-based neural interfaces for extracellular recording and stimulation, focusing on those technological approaches that are successfully demonstrated in vivo. The aim is to highlight the most reliable and ready-for-clinical-use solutions, in terms of materials technology and recording performance, other than spot major limitations and identify future trends in this field.

Sensing Inflammation Biomarkers with Electrolyte-Gated Organic Electronic Transistors.

An overview of cytokine biosensing is provided, with a focus on the opportunities provided by organic electronic platforms for monitoring these inflammation biomarkers which manifest at ultralow concentration levels in physiopathological conditions. Specifically, two of the field's state-of-the-art technologies-organic electrochemical transistors (OECTs) and electrolyte gated organic field effect transistors (EGOFETs)-and their use in sensing cytokines and other proteins associated with inflammation are a particular focus. The overview will include an introduction to current clinical and "gold standard" quantification techniques and their limitations in terms of cost, time, and required infrastructure. A critical review of recent progress with OECT- and EGOFET-based protein biosensors is presented, alongside a discussion onthe future of these technologies in the years and decades ahead. This is especially timely as the world grapples with limited healthcare diagnostics during the Coronavirus disease (COVID-19)pandemic where one of the worst-case scenarios for patients is the "cytokine storm." Clearly, low-cost point-of-care technologies provided by OECTs and EGOFETs can ease the global burden on healthcare systems and support professionals by providing unprecedented wealth of data that can help to monitor disease progression in real time.

Label free detection of miRNA-21 with electrolyte gated organic field effect transistors (EGOFETs).

We report a dual gate/common channel organic transistor architecture designed for quantifying the concentration of one of the strands of miRNA-21 in solution. The device allows one to measure the differential response between two gate electrodes, viz. one sensing and one reference, both immersed in the electrolyte above the transistor channel. Hybridization with oligonucleotide in the picomolar regime induces a sizable reduction of the current flowing through the transistor channel. The device signal is reported at various gate voltages, showing maximum sensitivity in the sublinear regime, with a limit of detection as low as 35 pM. We describe the dose curves with an analytical function derived from a thermodynamic model of the reaction equilibria relevant in our experiment and device configuration, and we show that the apparent Hill dependence on analyte concentration, whose exponent lies between 0.5 and 1, emerges from the interplay of the different equilibria. The binding free energy characteristic of the hybridization on the device surface is found to be approximately 20% lower with respect to the reaction in solution, hinting to partially inhibiting effect of the surface and presence of competing reactions. Impedance spectroscopy and surface plasmon resonance (SPR) performed on the same oligonucleotide pair were correlated to the electronic current transduced by the EGOFET, and confirmed the selectivity of the biorecognition probe covalently bound on the gold surface.

Electrolyte-gated transistors for enhanced performance bioelectronics.

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.

Long-range selective transport of anions and cations in graphene oxide membranes, causing selective crystallization on the macroscale.

Monoatomic nanosheets can form 2-dimensional channels with tunable chemical properties, for ion storage and filtering applications. Here, we demonstrate transport of K+, Na+, and Li+ cations and F- and Cl- anions on the centimeter scale in graphene oxide membranes (GOMs), triggered by an electric bias. Besides ion transport, the GOM channels foster also the aggregation of the selected ions in salt crystals, whose composition is not the same as that of the pristine salt present in solution, highlighting the difference between the chemical environment in the 2D channels and in bulk solutions.

Anti-drug antibody detection with label-free electrolyte-gated organic field-effect transistors.

The efficacy of immunotherapy can be undermined by the development of an immune response against a drug/antibody mediated by anti-drug antibodies (ADAs) in treated patients. We present the first label-free EGOFET immunosensor that integrates a biological drug, Nivolumab (Opdivo©), as a specific recognition moiety to quantitatively and selectively detect ADAs against the drug. The limit of detection is 100 fM. This demonstration is a prelude to the detection of ADAs in a clinical setting in the treatment of different pathologies, and it also enables rapid screening of biological drugs for immunogenicity.

Label-free immunodetection of α-synuclein by using a microfluidics coplanar electrolyte-gated organic field-effect transistor.

The aggregation of α-synuclein is a critical event in the pathogenesis of neurological diseases, such as Parkinson or Alzheimer. Here, we present a label-free sensor based on an Electrolyte-Gated Organic Field-Effect Transistor (EGOFET) integrated with microfluidics that allows for the detection of amounts of α-synuclein in the range from 0.25 pM to 25 nM. The lower limit of detection (LOD) measures the potential of our integrated device as a tool for prognostics and diagnostics. In our device, the gate electrode is the effective sensing element as it is functionalised with anti-(α-synuclein) antibodies using a dual strategy: i) an amino-terminated self-assembled monolayer activated by glutaraldehyde, and ii) the His-tagged recombinant protein G. In both approaches, comparable sensitivity values were achieved, featuring very low LOD values at the sub-pM level. The microfluidics engineering is central to achieve a controlled functionalisation of the gate electrode and avoid contamination or physisorption on the organic semiconductor. The demonstrated sensing architecture, being a disposable stand-alone chip, can be operated as a point-of-care test, but also it might represent a promising label-free tool to explore in-vitro protein aggregation that takes place during the progression of neurodegenerative illnesses.

A minimal model of hospital patients' dynamics in COVID-19.

Italy has been one of the countries hardest hit by the coronavirus disease (COVID-19) pandemic. While the overall policy in response to the epidemic was to a large degree centralised, the regional basis of the healthcare system represented an important factor affecting the natural dynamics of the disease induced geographic specificities. Here, we characterise the region-specific modulation of COVID dynamics with a reduced exponential model leveraging available data on sub-intensive and intensive care unit patients made available by all regional councils from the very onset of the disease. This simple model provides a rather good fit of regional patient dynamics, particularly for regions where the affected population was large, highlighting important region-specific patterns of epidemic dynamics.

Water-Based PEDOT:Nafion Dispersion for Organic Bioelectronics.

The water dispersion of the conductive polymer poly(3,4-ethylenedioxythiophene):poly(styrenesulfonic acid) (PEDOT:PSS) is one of the most used material precursors in organic electronics also thanks to its industrial production. There is a growing interest for conductive polymers that could be alternative surrogates or replace PEDOT:PSS in some applications. A recent study by our group compared electrodeposited PEDOT:Nafion vs PEDOT:PSS in the use for neural recordings. Here, we introduce an easy and reproducible synthetic protocol to prepare a water dispersion of PEDOT:Nafion. The conductivity of the pristine material is on the order of 2 S cm-1 and was improved up to ≈6 S cm-1 upon treatment with ethylene glycol. Faster ion transfer was assessed by electrochemical impedance spectroscopy (EIS), and, interestingly, an improved adhesion was observed for coatings of the new PEDOT:Nafion dispersion on glass substrates, even without the addition of the silane cross-linker needed for PEDOT:PSS. As proof of concept, we demonstrate the use of this novel water dispersion of PEDOT:Nafion in three different organic electronic device architectures, namely, an organic electrochemical transistor (OECT), a memristor, and an artificial synapse.

Nanodecoration of electrospun polymeric fibers with nanostructured silver coatings by ionized jet deposition for antibacterial tissues.

Silver-based nanomaterials are used as antibacterial agents in a number of applications, including wound dressing, where electrospun materials can effectively promote wound healing and tissue regeneration thanks to their biomimicry, flexibility and breathability. Incorporation of such nanomaterials in electrospun nonwovens is highly challenging if aiming at maximizing stability and antibacterial efficacy and minimizing silver detachment, without neglecting process straightforwardness and scalability. In this work nanostructured silver coatings were deposited by Ionized Jet Deposition (IJD) on Polylactic acid, a medical grade polyester-urethane and Polyamide 6,6 nanofibers. The resulting materials were thoroughly characterized to gain an in-depth view of coating morphology and substrate resistance to the low-temperature deposition process used. Morphology of silver coatings with well-cohesive grains having dimensions from a few tens to a few hundreds of nanometers was analyzed by SEM, TEM and AFM. TGA, DSC, FTIR and GPC showed that the polymers well withstand the deposition process with negligible effects on their properties, the only exception being the polylactic acid that resulted more susceptible to degradation. Finally, the efficacy against S. aureus and E. coli bacterial strains was demonstrated, indicating that electrospun fibers decorated with nanostructured silver by IJD represent a breakthrough solution in the field of antibacterial devices.

Harnessing Selectivity and Sensitivity in Electronic Biosensing: A Novel Lab-on-Chip Multigate Organic Transistor.

Electrolyte gated organic transistors can operate as powerful ultrasensitive biosensors, and efforts are currently devoted to devising strategies for reducing the contribution of hardly avoidable, nonspecific interactions to their response, to ultimately harness selectivity in the detection process. We report a novel lab-on-a-chip device integrating a multigate electrolyte gated organic field-effect transistor (EGOFET) with a 6.5 µL microfluidics set up capable to provide an assessment of both the response reproducibility, by enabling measurement in triplicate, and of the device selectivity through the presence of an internal reference electrode. As proof-of-concept, we demonstrate the efficient operation of our pentacene based EGOFET sensing platform through the quantification of tumor necrosis factor alpha with a detection limit as low as 3 pM. Sensing of inflammatory cytokines, which also include TNFα, is of the outmost importance for monitoring a large number of diseases. The multiplexable organic electronic lab-on-chip provides a statistically solid, reliable, and selective response on microliters sample volumes on the minutes time scale, thus matching the relevant key-performance indicators required in point-of-care diagnostics.

Amphiphilic cationic cyclodextrin nanovesicles: a versatile cue for guiding cell adhesion.

It is well known that amphiphilic cationic ß-cyclodextrins (amßCDs) form nanovesicles able to release their cargo in aqueous solution upon applying different stimuli. In addition they can be selectively positioned onto substrates by unconventional soft lithography. This makes them a powerful tool for designing environments where different cues can be externally supplied to the cells helping to achieve good control of their fate. Lithographically controlled wetting (LCW) of amßCD nanovesicles loaded with fluorescein isothiocyanate (FITC), amßCD/FITC, has been used here to fabricate geometrically functionalized surfaces, thus achieving multiscale control of the cell environment. The amßCD functionalization was strongly influenced by the surface energy of the underlying substrates that, according to their hydrophobicity, orient the amßCD in a different way, thus "offering" different portions to the cells. The structure of the pattern was characterized both over large scales exploiting the FITC fluorescence and at the nanoscale by atomic force microscopy. Cell guidance and aCD/FITC cell internalization were demonstrated in human neuroblastoma SHSY5Y cells.

Retinoic acid/calcite micro-carriers inserted in fibrin scaffolds modulate neuronal cell differentiation.

The controlled release of cell differentiating agents is crucial in many aspects of regenerative medicine. Here we propose the use of hybrid calcite single crystals as micro-carriers for the controlled and localized release of retinoic acid, which is entrapped within the crystalline lattice. The release of retinoic acid occurs only in the proximity of stem cells, upon dissolution of the calcite hybrid crystals that are dispersed in the fibrin scaffold. These hybrid crystals provide a sustained dosage of the entrapped agent. The environment provided by this composite scaffold enables differentiation towards neuronal cells that form a three-dimensional neuronal network.

Electrodeposited PEDOT:Nafion Composite for Neural Recording and Stimulation.

Microelectrode arrays are used for recording and stimulation in neurosciences both in vitro and in vivo. The electrodeposition of conductive polymers, such as poly(3,4-ethylene dioxythiophene) (PEDOT), is widely adopted to improve both the in vivo recording and the charge injection limit of metallic microelectrodes. The workhorse of conductive polymers in the neurosciences is PEDOT:PSS, where PSS represents polystyrene-sulfonate. In this paper, the counterion is the fluorinated polymer Nafion, so the composite PEDOT:Nafion is deposited onto a flexible neural microelectrode array. PEDOT:Nafion coated electrodes exhibit comparable in vivo recording capability to the reference PEDOT:PSS, providing a large signal-to-noise ratio in a murine animal model. Importantly, PEDOT:Nafion exhibits a minimized polarization during electrical stimulation, thereby resulting in an improved charge injection limit equal to 4.4 mC cm-2 , almost 80% larger than the 2.5 mC cm-2 that is observed for PEDOT:PSS.