Conjugated polymers provide a unique toolbox for establishing electrical communication with biological systems. In this talk, I will show how modulating the chemistry of these materials can lead to interfaces that maximize interactions with biological systems while maintaining electronic transport properties. These materials are then used in organic electrochemical transistors (OECTs) to detect biological species in physiological media. I will introduce two types of OECT based sensors; one that detects metabolites with performance exceeding the state-of-the-art, and the other that detects coronavirus spike proteins at the physical limit. Having challenged these sensors with patient samples and cellular media, I will discuss areas where proof-of-concept platforms may fail and how to prolong the operation in biological environments. By tackling each of these problems, we improve device performance to a level that marks a considerable step toward label-free diagnostics.

The interface between biological cells and non-biological materials has profound influences on cellular activities, chronic tissue responses, and ultimately the success of medical implants and bioelectronic devices. The optimal coupling between cells and materials is mainly based on surface interaction, electrical communication and sensing1.
In the last years, many efforts have been devoted to engineer materials to recapitulate both the environment (i.e., dimensionality, curvature, dynamicity)2 and the functionalities (i.e., long and short term synaptic plasticity)3 of the neuronal tissue to ensure a better integration of the bioelectronic platform and cells. In this scenario, resembling the operation and the composition of the neuronal membrane might be beneficial to reconstitute synaptic proteins’ arrangement (i.e. synaptic receptors) and electronic functionalities to further optimize the communication between neuronal cells and in vitro bioelectronic platforms2.
Here, we explore how organic neuromorphic devices and supported lipid bilayers (SLBs) can recapitulate short and long term plasticity in biohybrid synapses.
Through the neurotransmitters’ oxidation (i.e. catecholamines) first, we were capable of modulating the synaptic potentiation and ultimately the coupling with biological cells to form a functional synapse.
and artificial membranes we were capable of tuning the response of a neuromorphic platform to modulate its conductance over time. Then, we functionalized the organic neuromorphic transistor with a supported lipid bilayer and investigated how the PEDOT:PSS – artificial membrane interface can affect the ion flow and thus the device’s conductance over time and the short term plasticity of the biohybrid synapse.
In turn, this could represent a first step toward in vitro adaptive neurohybrid interfaces to engineering neuronal networks with biomimetic structural and functional connections at synaptic level.
References
1. Lubrano, C. et al. Towards biomimetic electronics that emulate cells. MRS Commun. 10, 398–412 (2020).
2. Mariano, A. et al. Advances in cell-conductive polymer biointerfaces and role of the plasma membrane (2021).
3. Keene, S. T. A biohybrid synapse with neurotransmitter-mediated plasticity. Nat. Mater. 19, 16 (2020).

Thiophene-based trimers is a new class of organic electronic molecular units, which enable the organization and polymerization of conjugated structures within living biological systems and inside operating electronic devices and systems. The trimers can be equipped with side groups that targets and promotes coupling to specific surfaces and (bio-)chemical cues making self-organization and -assembly of organic bioelectronics possible in a novel manner. The chemical and physical fundamentals of the trimers, the route of polymerization, and their performance while operating in neuromorphic and in vivo-manufactured bioelectronics will be reported. Specifically, neuromorphic systems based on organic electrochemical transistors including the trimers will be described along with bioelectronic systems formed inside living plants, cells and animal models. Our findings promise for radically new ways of forming bioelectronic systems in living systems, which mimicks the structures and functions of the signaling of biology.

Organic electrochemical transistors have garnered strong interest due to ability to efficiently transduce biological signals into electronic ones thanks to the mixed ion and electron conduction of the semiconducting film in the channel. However, the lack of understanding of these mechanisms during the operation of OECTs hinders the use of OECT-based bioelectronic technologies in circuits. Here we investigate how polymer hydration in aqueous biological media affects mixed conduction properties of n-type (electron transporting) semiconductor transistors. We use electrochemical quartz crystal microbalance with dissipation monitoring (eQCM-D) coupled with UV-Vis and impedance spectroscopy to observe the hydration of n-type polymers with varying lengths of ethylene glycol side-chains attached to NDI-T2 backbone. By varying the hydrophilic content as well as conformation of the n-type polymer, we modulate the hydration which in turn governs the in situ (dynamic) morphology of the film and hence the electronic and ionic mobilities. Our study suggests that hydration is not de facto beneficial for the n-type OECT performance, as has recently been observed for the p-type counterparts.¹

Among the existing two-dimensional materials, MXenes, i.e. transition metal carbides, nitrides and/or carbonitrides, stand out for their excellent electrochemical properties. Due to their high charge storage capacity, metal-like conductivity, biocompatibility as well as hydrophilicity, Ti3C2Tx MXene-based inks hold great potential for scalable production of skin conformable electronics via direct printing methods. Herein, we develop an aqueous MXene ink and inkjet-print MXene films on freestanding, flexible, and conducting polymer-based substrates. These skin-adherent MXene electrodes detect electrocardiography signals with high signal-to-noise ratio while exhibiting preserved electrical performance after 1000 cycles of bending with a 50 d long shelf life in ambient conditions. We show that printed MXene films can be further functionalized to perform as multifunctional biosensing units. When integrated with a sodium (Na+) ion selective membrane, MXene electrodes detect Na+ in artificial sweat with a sensitivity of 40 mV per decade. When the films are functionalized with antibodies, they generate an electrical signal in response to a pro-inflammatory cytokine protein (interferon gamma) with a sensitivity of 3.9 mV per decade. Our findings demonstrate how inkjet-printed MXene films simplify the fabrication of next-generation wearable electronic platforms that comprise multimodal sensors.

The unmatchable capability of living cells to fabricate complex structure starting from simple building blocks offers new paradigms to seamlessly integrate new electronic structures into the living matter, creating new hybrid devices. We exploited this potential in a simple tissue-like organism, the freshwater polyp Hydra vulgaris, and here we provide an overview of recent and on-going results obtained with this model together with our methodological approaches ranging from behavioural to optical, spectroscopical, and electrical characterization. By simple exposure to conjugated oligomers we show the capability of Hydra to fabricate new electronically conducting and electrochemically active microstructures in specific cell types [1]. In another case, fluorescent and conductive microfibers embedded into the tentacle tissues were produced starting from oligothiophene compounds, showing the feasibility to use these organisms as biofactories to fabricate and to test novel bioelectronic interface [2]. Tissues with integrated biocompatible electronics, manufactured in vivo in localized regions, may inspire new devices to manipulate biological functions by adding or augmenting conductivity in physiological or pathological contexts with spatiotemporal control, paving the way to new bioengineering concepts.

References

[1] G. Tommasini, G. Dufil, F. Fardella, X. Strakosas, E. Fergola, T. Abrahamsson, D. Bliman, R. Olsson, M. Berggren, A. Tino, E. Stavrinidou, C. Tortiglione, Seamless integration of bioelectronic interface in an animal model via in vivo polymerization of conjugated oligomers, Bioactive Materials 10 (2022) 107-116.

[2] M. Moros, F. Di Maria, P. Dardano, G. Tommasini, H. Castillo-Michel, A. Kovtun, M. Zangoli, M. Blasio, L. De Stefano, A. Tino, G. Barbarella, C. Tortiglione, In Vivo Bioengineering of Fluorescent Conductive Protein-Dye Microfibers, iScience 23(4) (2020) 101022.

Leveraging the biocatalytic machinery of living organisms for fabricating functional bioelectronic interfaces in-vivo not only enables seamless integration of microdevices into the tissue but also a greener solution for advanced microfabrication. Previously we have demonstrated that plants can polymerize conjugated oligomers in-vivo forming conductors within their structure. We showed that the polymerization is enzymatically catalyzed by endogenous peroxidases and we developed a series of conjugated oligomers that can be enzymatically polymerized in physiological conditions. The conjugated polymers integrate within the plant cell wall structure adding electronic functionality into the plant that is then explored for energy storage. Recently we demonstrated intact plants with electronic roots that continue to grow and develop enabling plant-biohybrid systems that maintain fully their biological processes. Furthermore, we have extended this concept into an animal model system. We demonstrated that Hydra, an invertebrate animal, can polymerize intracellularly conjugated oligomers in cells that expresses peroxidase activity. The conjugated polymer forms electronically conducting and electrochemically active domains in the µm range integrated within the hydra tissue. Our work paves the way for self-organized electronics in plant and animal tissue for modulating biological functions and in-vivo bio-fabrication of hybrid functional materials and devices.

Skin is the body’s largest organ, and is responsible for the transduction of a vast amount of information. This conformable, stretchable, self-healable and biodegradable material simultaneously collects signals from external stimuli that translate into information such as pressure, pain, and temperature. The development of electronic materials, inspired by the complexity of this organ is a tremendous, unrealized materials challenge. However, the advent of organic-based electronic materials may offer a potential solution to this longstanding problem. Over the past decade, we have developed materials design concepts to add skin-like functions to organic electronic materials without compromising their electronic properties. These new materials and new devices enabled arrange of new applications in medical devices, robotics and wearable electronics. In this talk, I will discuss basic material design concepts for realizing stretchable, self-healable and biodegradable conductive or semiconductive materials. I will show our methods for scalable fabrication of stretchable electronic circuit blocks. Finally, I will show a few examples of applications we are pursuing uniquely enabled by skin-like organic electronics when interfacing with biological systems, such as low-voltage electrical stimulation, high-resolution large area electrophysiology, “morphing electronics” that grows with biological system and genetically targeted chemical assembly - GTCA.

Bioelectronic devices have found use at the interface with neural tissue to investigate and treat nervous system disorders. We have developed and characterized a very thin flexible polyimide-based bioelectronic implant that is inserted along the thoracic spinal cord in rats directly in contact with the dorsal surface of the spinal cord. There was no negative impact on hind-limb functionality nor any change in the volume or shape of the spinal cord. The bioelectronic implant was maintained in rats for a period of 3 months. We have obtained the first subdural recordings of spinal cord activity in freely moving animals. Recordings contained multiple distinct voltage waveforms spatially localized to individual electrodes. A clinically relevant spinal contusion injury can be achieved with the implant in place. This device has great potential to monitor electrical signaling in the spinal cord after an injury, and in the future, this implant will facilitate the identification of biomarkers in spinal cord injury and recovery. We are exploring further development of this implant to deliver localized treatments to the spinal cord towards regeneration of damaged tissues to recover lost function. Localized therapies are being developed in the form of pharmaceutical treatment through the local delivery of neurotrophic factors, and electroceutical treatment through precise electrical field stimulation.

Miniaturized neural implants cover the wet interface between electronic and biological circuits and systems. They need to establish stable and reliable functional interfaces to the target structure in chronic application in neuroscientific experiments but especially in clinical applications in humans. Proper selection of substrate, insulation and electrode materials is of utmost importance to bring the interface in close contact with the neural target structures, minimize foreign body reaction after implantation and maintain functionality over the complete implantation period. Our work has focused on polymer substrates with integrated thin-film metallization as core of our flexible neural interfaces approach and silicone rubber with metal sheets. Micromachining and laser structuring are the main technologies for electrode array manufacturing. Different design and development aspects from the first idea to first-in-human studies are presented and challenges in translational research are discussed. Reliability data from long-term ageing studies and chronic experiments show the applicability of thin-film implants for stimulation and recording and ceramic packages for electronics protection. Examples of sensory feedback after amputation trauma, vagal nerve stimulation to treat hypertension and chronic recordings from the brain surface display opportunities and challenges of these miniaturized implants. System assembly and interfacing microsystems to robust cables and connectors still is a major challenge in translational research and transition of research results into medical products.

Neuroscience in non-human primates, as well as neurotherapy in patients, can benefit tremendously from recent developments in engineering flexible interfaces to record neuronal activity. In particular thin-film electrodes offer the potential for electrophysiological recordings over very long periods of time, accessing any part and potentially covering large parts of the brain. I will present some of the work that we have performed, in collaboration with engineers, to develop and use polyimide-based electrodes for recordings from the cortical surface and the cortical neuropil. I will then try to identify those potential future developments that are most urgently needed and those that will enable the largest further steps.

Successful translation of electronic devices, operated as sensors and/or transducers, to clinical settings demands precise knowledge of their working principles as well as of the peculiar needs arising from the targeted application scenarios. This demands a tight mutual exchange between the scientific fields of device engineering and neurosciences, which metaphorically mirrors the most ambitious goal of Neuroelectronics: namely the bidirectional communication of electronic devices with the living matter, and, in particular, with the central nervous system.

The technological and scientific platform of organic electronics provides materials, tools and procedures to effectively tackle this goal, giving rise to the field of Organic Neuroelectronics.

Typically, operation of Organic Neuroelectronic devices demands driving voltages across the biotic/abiotic interface, which may result in undesired electrochemical reactions that are harmful both for the patient and for the device.

This talk reports on a novel device architecture which bypasses these drawbacks: the common-drain/grounded-source electrolyte-gated organic transistor. When implanted, this architecture allows one to maximize brain-signal amplification while applying null net bias, and consequently eliciting no parasitic currents, across the tissue. We demonstrate the viability of the proposed configuration by recording in vivo the somatosensory evoked activity from the barrel cortex of rats, achieving a superior signal-to-noise ratio which enables the detection of evoked activity at the single-trial level.

In drug-resistant partial epilepsies, resective surgery is the treatment of choice to suppress seizures, provided that the epileptogenic zone (EZ) is clearly identified and that it can be safely removed. In this context, the capacity to rely on objective biomarkers of the EZ is fundamental to define the optimal surgical approach in the specific context of each patient. One of the most important biomarkers are electrophysiological signals which represent local field potential variations generated by a network of neurons. Here our focus is a specific oscillation band knows as fast ripple (FR) with the frequency band between 250 Hz to 600 Hz. FRs are transit signals which mixed with ongoing activities and last only few tens of millisecond. There are fundings that show FRs are specific to epileptogenic zone. However, routine clinical use of FRs is a long path since we do not know which information is carried by FRs and how FRs correlates with epileptogenic seizure  [1] .

The aim of this project is to optimize the recording electrodes properties to improve the detection of epileptic FRs. In first stage we benefited from the epileptic computational mode and tuned the electrode’s characteristics including size, impedance, geometry, and material.  According to the simulated results, microelectrodes are fabricated. The impedance of electrodes were tuned by controlling the electrodeposition of PEDOT:PSS conductive polymer.  Electrode’s recording sites are characterized in terms of morphology, impedance, electricidal conductivity, uv-visible and furrier transform infrared transmission. The double layer capacitance of the electrodes was measured experimentally and modeled with an equivalent circuit  [2-3].

In the next step, microelectrodes are implanted in kainate mouse model of temporal lobe epilepsy for 28 days.  The recorded signals are segmented and processed to extract the fast ripples. We found that the amplitude of the FRs signals to the background activity increases by lowering the impedance. In addition, the cut-off frequency at which electrodes the electrode transits from predominantly resistive to capacitive, highly influence the FRs resolvability.

In vitro models of biological systems are essential for our understanding of biological systems. In many cases where animal models have failed to translate to useful data for human diseases, physiologically relevant in vitro models can bridge the gap. Many difficulties exist in interfacing complex, 3D cell biology models with technology adapted for monitoring function. Polymeric electroactive materials and devices can bridge the gap between hard inflexible materials used for physical transducers and soft, compliant biological tissues. An additional advantage of these electronic materials is their flexibility for processing and fabrication in a wide range of formats. In this presentation, I will discuss our recent progress in pushing conducting polymer devices, to 3D, to simultaneously host and monitor complex multi-cellular models of tissues and organs. Building on our previous work that showcased a bioelectronic model of the human intestine, we are now incorporating elements of the microbiome and the immune system as well as the enteric nervous system. Coupling this model with our model of the neuro-vascular unit (including blood brain barrier) currently in progress, will bring us to our goal of a physiologically representative in vitro model of the gut-brain-microbiome axis.

Organic electrochemical transistors (OECTs) are electronic devices having organic mixed
ionic/electronic conductors as core components. The efficient ion-to-electron conversion of
organic materials allows for low operating voltages, high amplification, and adaptability to various
form factors. These characteristics, together with operation in physiological conditions make
OECTs promising devices for bioelectronics, including the recording of action potentials from
electrogenic cells.[1].

When interfacing with cells in vitro, the active material should satisfy several requirements
to allow both device performance and a suitable platform for cell culture. On the device side, the
conjugated polymer should provide a path for ionic and electronic conductivity as well as stability
in contact with aqueous electrolytes. Additionally, for in vitro cell culture, the polymer should be
resistant to sterilization, provide long-term stability in conditions needed for cell survival (i.e., 37
ºC in cell culture media), and offer a suitable substrate for cell attachment.

Conjugated polymers with ethylene glycol side chains provide films that are stable in
contact with water and that have the mixed ionic/electronic conductivity needed for efficient
OECT operation. However, ethylene glycol side chains, with their antifouling properties, offer a
poor substrate for cell adhesion.[2]

Here, we focus our attention to the cell-device interface. We develop polymer blends
combining efficient OECT operation and the functionalities needed for conjugation with specific
cues for cell attachment. Isoindigo donor-acceptor copolymers with hybrid alkyl/ethylene glycol
side chains provide a path for electronic and ionic conductivity.[3] PEDOT having carboxyl side
groups [3,4] is used to couple functional groups for click-chemistry with cues for cell attachment.

The blends enabled long-term device stability, for up to 6 months in cell culture media.
Covalent attachment of an extracellular matrix protein (i.e., fibronectin) provided a suitable
platform for attachment and recording of human embryonic stem cells-derived cardiomyocytes.
Overall, the blends offer a versatile strategy to attach cues that can be used to promote and/or
modulate cell-device interaction. As active materials for OECTs, the blends provide a suitable
platform to monitor the activity of electrogenic cells in vitro.