Single-Step Functionalization Strategy of Graphene Microtransistor Array with Chemically Modified Aptamers for Biosensing Applications.

Graphene solution-gated field-effect transistors (gSGFETs) offer high potential for chemical and biochemical sensing applications. Among the current trends to improve this technology, the functionalization processes are gaining relevance for its crucial impact on biosensing performance. Previous efforts are focused on simplifying the attachment procedure from standard multi-step to single-step strategies, but they still suffer from overreaction, and impurity issues and are limited to a particular ligand. Herein, a novel strategy for single-step immobilization of chemically modified aptamers with fluorenylmethyl and acridine moieties, based on a straightforward synthetic route to overcome the aforementioned limitations is presented. This approach is benchmarked versus a standard multi-step strategy using thrombin as detection model. In order to assess the reliability of the functionalization strategies 48-gSGFETs arrays are employed to acquire large datasets with multiple replicas. Graphene surface characterization demonstrates robust and higher efficiency in the chemical coupling of the aptamers with the single-step strategy, while the electrical response evaluation validates the sensing capability, allowing to implement different alternatives for data analysis and reduce the sensing variability. In this work, a new tool capable of overcome the functionalization challenges of graphene surfaces is provided, paving the way toward the standardization of gSGFETs for biosensing purposes.

Covalent functionalisation controlled by molecular design for the aptameric recognition of serotonin in graphene-based field-effect transistors.

In the last decade, solution-gated graphene field effect transistors (GFETs) showed their versatility in the development of a miniaturized multiplexed platform for electrophysiological recordings and sensing. Due to their working mechanism, the surface functionalisation and immobilisation of receptors are pivotal to ensure the proper functioning of devices. Herein, we present a controlled covalent functionalisation strategy based on molecular design and electrochemical triggering, which provide a monolayer-like functionalisation of micro-GFET arrays retaining the electronic properties of graphenes. The functionalisation layer as a receptor was then employed as the linker for serotonin aptamer conjugation. The micro-GFET arrays display sensitivity toward the target analyte in the micromolar range in a physiological buffer (PBS 10 mM). The sensor allows the in-flow real-time monitoring of serotonin transient concentrations with fast and reversible responses.

Correction: Organ-on-a-chip with integrated semitransparent organic electrodes for barrier function monitoring.

Correction for 'Organ-on-a-chip with integrated semitransparent organic electrodes for barrier function monitoring' by Denise Marrero et al., Lab Chip, 2023, https://doi.org/10.1039/d2lc01097f.

Organ-on-a-chip with integrated semitransparent organic electrodes for barrier function monitoring.

Organs-on-a-chip (OoC) are cell culture platforms that replicate key functional units of tissues in vitro. Barrier integrity and permeability evaluation are of utmost importance when studying barrier-forming tissues. Impedance spectroscopy is a powerful tool and is widely used to monitor barrier permeability and integrity in real-time. However, data comparison across devices is misleading due to the generation of a non-homogenous field across the tissue barrier, making impedance data normalization very challenging. In this work, we address this issue by integrating PEDOT:PSS electrodes for barrier function monitoring with impedance spectroscopy. The semitransparent PEDOT:PSS electrodes cover the entire cell culture membrane providing a homogenous electric field across the entire membrane making the cell culture area equally accountable to the measured impedance. To the best of our knowledge, PEDOT:PSS has never been used solely to monitor the impedance of cellular barriers while enabling optical inspection in the OoC. The performance of the device is demonstrated by lining the device with intestinal cells where we monitored barrier formation under flow conditions, as well as barrier disruption and recovery under exposure to a permeability enhancer. The barrier tightness and integrity, and the intercellular cleft have been evaluated by analyzing the full impedance spectrum. Furthermore, the device is autoclavable paving the way toward more sustainable OoC options.

Bidirectional Modulation of Neuronal Cells Electrical and Mechanical Properties Through Pristine and Functionalized Graphene Substrates.

In recent years, the quest for surface modifications to promote neuronal cell interfacing and modulation has risen. This course is justified by the requirements of emerging technological and medical approaches attempting to effectively interact with central nervous system cells, as in the case of brain-machine interfaces or neuroprosthetic. In that regard, the remarkable cytocompatibility and ease of chemical functionalization characterizing surface-immobilized graphene-based nanomaterials (GBNs) make them increasingly appealing for these purposes. Here, we compared the (morpho)mechanical and functional adaptation of rat primary hippocampal neurons when interfaced with surfaces covered with pristine single-layer graphene (pSLG) and phenylacetic acid-functionalized single-layer graphene (fSLG). Our results confirmed the intrinsic ability of glass-supported single-layer graphene to boost neuronal activity highlighting, conversely, the downturn inducible by the surface insertion of phenylacetic acid moieties. fSLG-interfaced neurons showed a significant reduction in spontaneous postsynaptic currents (PSCs), coupled to reduced cell stiffness and altered focal adhesion organization compared to control samples. Overall, we have here demonstrated that graphene substrates, both pristine and functionalized, could be alternatively used to intrinsically promote or depress neuronal activity in primary hippocampal cultures.

Distortion-Free Sensing of Neural Activity Using Graphene Transistors.

Graphene solution-gated field-effect transistors (g-SGFETs) are promising sensing devices to transduce electrochemical potential signals in an electrolyte bath. However, distortion mechanisms in g-SGFET, which can affect signals of large amplitude or high frequency, have not been evaluated. Here, a detailed characterization and modeling of the harmonic distortion and non-ideal frequency response in g-SGFETs is presented. This accurate description of the input-output relation of the g-SGFETs allows to define the voltage- and frequency-dependent transfer functions, which can be used to correct distortions in the transduced signals. The effect of signal distortion and its subsequent calibration are shown for different types of electrophysiological signals, spanning from large amplitude and low frequency cortical spreading depression events to low amplitude and high frequency action potentials. The thorough description of the distortion mechanisms presented in this article demonstrates that g-SGFETs can be used as distortion-free signal transducers not only for neural sensing, but also for a broader range of applications in which g-SGFET sensors are used.

A perfusion chamber for monitoring transepithelial NaCl transport in an in vitro model of the renal tubule.

Transepithelial electrical measurements in the renal tubule have provided a better understanding of how kidney regulates electrolyte and water homeostasis through the reabsorption of molecules and ions (e.g., H2 O and NaCl). While experiments and measurement techniques using native tissue are difficult to prepare and to reproduce, cell cultures conducted largely with the Ussing chamber lack the effect of fluid shear stress which is a key physiological stimulus in the renal tubule. To overcome these limitations, we present a modular perfusion chamber for long-term culture of renal epithelial cells under flow that allows the continuous and simultaneous monitoring of both transepithelial electrical parameters and transepithelial NaCl transport. The latter is obtained from electrical conductivity measurements since Na+ and Cl- are the ions that contribute most to the electrical conductivity of a standard physiological solution. The system was validated with epithelial monolayers of raTAL and NRK-52E cells that were characterized electrophysiologically for 5 days under different flow conditions (i.e., apical perfusion, basal, or both). In addition, apical to basal chemical gradients of NaCl (140/70 and 70/140 mM) were imposed in order to demonstrate the feasibility of this methodology for quantifying and monitoring in real time the transepithelial reabsorption of NaCl, which is a primary function of the renal tubule.

A compartmentalized microfluidic chip with crisscross microgrooves and electrophysiological electrodes for modeling the blood-retinal barrier.

The interconnection of different tissue-tissue interfaces may extend organ-on-chips to a new generation of sophisticated models capable of recapitulating more complex organ-level functions. Single interfaces are largely recreated in organ-on-chips by culturing the cells on opposite sides of a porous membrane that splits a chamber in two or by connecting the cells of two adjacent compartments through microchannels. However, it is difficult to interconnect more than one interface using these approaches. To address this challenge, we present a novel microfluidic device where cells are arranged in parallel compartments and are highly interconnected through a grid of microgrooves, which facilitates paracrine signaling and heterotypic cell-cell contact between multiple tissues. In addition, the device includes electrodes on the substrate for the measurement of transepithelial electrical resistance (TEER). Unlike conventional methods for measuring the TEER where electrodes are on each side of the cell barrier, a method with only electrodes on the substrate has been validated. As a proof-of-concept, we have used the device to mimic the structure of the blood-retinal barrier by co-culturing primary human retinal endothelial cells (HREC), a human neuroblastoma cell line (SH-SY5Y), and a human retinal pigment epithelial cell line (ARPE-19). Cell barrier formations were assessed by a permeability assay, TEER measurements, and ZO-1 expression. These results validate the proposed microfluidic device with microgrooves as a promising in vitro tool for the compartmentalization and monitoring of barrier tissues.

Quantitative self-powered electrochromic biosensors.

Self-powered sensors are analytical devices able to generate their own energy, either from the sample itself or from their surroundings. The conventional approaches rely heavily on silicon-based electronics, which results in increased complexity and cost, and prevents the broader use of these smart systems. Here we show that electrochromic materials can overcome the existing limitations by simplifying device construction and avoiding the need for silicon-based electronics entirely. Electrochromic displays can be built into compact self-powered electrochemical sensors that give quantitative information readable by the naked eye, simply controlling the current path inside them through a combination of specially arranged materials. The concept is validated by a glucose biosensor coupled horizontally to a Prussian blue display designed as a distance-meter proportional to (glucose) concentration. This approach represents a breakthrough for self-powered sensors, and extends the application of electrochromic materials beyond smart windows and displays, into sensing and quantification.

New trends in quantitative assessment of the corneal barrier function.

The cornea is a very particular tissue due to its transparency and its barrier function as it has to resist against the daily insults of the external environment. In addition, maintenance of this barrier function is of crucial importance to ensure a correct corneal homeostasis. Here, the corneal epithelial permeability has been assessed in vivo by means of non-invasive tetrapolar impedance measurements, taking advantage of the huge impact of the ion fluxes in the passive electrical properties of living tissues. This has been possible by using a flexible sensor based in SU-8 photoresist. In this work, a further analysis focused on the validation of the presented sensor is performed by monitoring the healing process of corneas that were previously wounded. The obtained impedance measurements have been compared with the damaged area observed in corneal fluorescein staining images. The successful results confirm the feasibility of this novel method, as it represents a more sensitive in vivo and non-invasive test to assess low alterations of the epithelial permeability. Then, it could be used as an excellent complement to the fluorescein staining image evaluation.