Graphene – Coated Microelectrode Arrays

Objective. The goal of this study was to assess the electrochemical properties of boron-doped diamond (BDD) electrodes in relation to conventional titanium nitride (TiN) electrodes through in vitro and in vivo measurements. Approach. Electrochemical impedance spectroscopy, cyclic voltammetry and voltage transient (VT) measurements were performed in vitro after immersion in a 5% albumin solution and in vivo after subcutaneous implantation in rats for 6 weeks. Main results. In contrast to the TiN electrodes, the capacitance of the BDD electrodes was not significantly reduced in albumin solution. Furthermore, BDD electrodes displayed a decrease in the VTs and an increase in the pulsing capacitances immediately upon implantation, which remained stable throughout the whole implantation period, whereas the opposite was the case for the TiN electrodes. Significance. These results reveal that BDD electrodes possess a superior biofouling resistance, which provides significantly stable electrochemical properties both in protein solution as well as in vivo compared to TiN electrodes.

Objective. To determine whether changes in electrochemical properties of porous titanium nitride (TiN) electrodes as a function of time after implantation are different from those of smooth TiN electrodes. Approach. Eight smooth and 8 porous TiN coated electrodes were implanted in 8 rats. Before implantation, voltage transients, cyclic voltammograms and impedance spectra were recorded in phosphate buffered saline (PBS). After implantation, these measurements were done weekly to investigate how smooth and porous electrodes were affected by implantation. Main results. The electrode capacitance of the porous TiN electrodes decreased more than the capacitance of the smooth electrodes due to acute implantation under fast measurement conditions (such as stimulation pulses). This indicates that protein adhesion presents a greater diffusion limitation for counter-ions for the porous than for the smooth electrodes. The changes in electrochemical properties during the implanted period were similar for smooth and porous TiN electrodes, indicating that cell adhesion poses a similar diffusion limitation for smooth and porous electrodes. Significance. This knowledge can be used to optimize the porous structure of the TiN film, so that the effect of protein adhesion on the electrochemical properties is diminished. Alternatively, an additional coating could be applied on the porous TiN that would prevent or minimize protein adhesion.

Microelectrode Array (MEA) devices are well established platforms for biosensing applications, however, limitations in electrode materials and cell coupling to electrodes still exist. Signal-to-noise ratio (SNR), a fabrication limitation, can be improved by reducing the impedance of the microelectrodes without increasing their physical size using conductive nanomaterials. Platinum is a well-studied conductive material that has been determined to be compatible with MEA devices, contains high electronic conductivity, is non-toxic to cells and has excellent catalytic activity. Nano-porous platinum, as compared to pure platinum, has larger surface area enabled by nano-scale definition, and as a result can lower the impedance of microelectrodes. Therefore, in this study nano-porous platinum was electroplated onto thin film gold electrodes to increase SNR in a single-well MEA device. Numerous commercial and research grade MEAs are available and have been subject to for various applications such as disease modeling and pharmaceutical screening. Although high densities of cells can increase the probability of cell-electrode coupling, only signals near electrodes will be detected. Our approach introduces a method of precise patterning of cells on electrodes to increase the probability of cell-electrode coupling and reducing inaccuracies and expenses associated with cell plating. The single-well MEA glass die was designed and fabricated to contain 64 microelectrodes (30 µm diameter with a pitch of 200 µm) and 4 internal on-chip reference electrodes. The MEA glass die fabrication was performed on a 4-inch, 500 µm thick glass wafer. First, metal traces composed of titanium and gold were deposited using lift-off lithography. Subsequently, 5 µm of SU-8 was spin coated and defined as an insulation layer. A separate printed circuit board (PCB) was designed and fabricated to contain 72 contact pads and routing traces. The biocompatible epoxy 353ND was used to package the glass die and PCB in specific orientations determined by alignment markers. Ball bonding with 20 µm diameter gold wire was used to wirebond the glass die to the PCB. A packaging technique was developed that would allow the characterization of key properties of the MEA. This technique involved using copper wire, 100 µm in diameter, and attaching it to the contact pads on the backside of PCBs using a conductive epoxy. Finally, a polystyrene culture well and lid were attached to the MEA using the epoxy, 353ND. Nano-Porous platinum electroplating solution was prepared with a combination of chloroplantinic acid, HCl and lead acetate, all diluted in DI water. Fully packaged devices containing approximately 3 mL of electrolyte were placed within the Axion BioSystems Muse for nano-porous platinum electrodeposition, utilizing the custom Application Specific Integrated Circuit (ASIC) in the system at room temperature. The nano-porous platinum electroplated devices were subsequently characterized using electrical impedance spectroscopy and cyclic voltammetry against the control devices, which contained gold electrodes. The impedance of each device was measured across a scanned frequency range of 10 Hz-100 kHz between the microelectrode, the internal reference electrode and cellular conductive media. The impedance of the nano-porous platinum devices is an order of magnitude lower than the devices with gold electrodes. Cyclic voltammograms were obtained at a scan rate of 20 mV/s in the range from -0.28 to 1.2 V vs. Ag/AgCl. RMS noise comparison between nano-porous platinum electroplated devices and control devices (gold electrodes) was determined using the Muse system and Axion’s Integrated Studio (AxIS) software. Biosensing applications for the devices were explored using electrogenic cells in combination with a precise plating technique. As depicted in Figure 1, cells were positioned in a pattern on and around electrodes to increase the probability of cell-electrode coupling. The electrical response was obtained and analyzed using the Muse system and AxIS software. SNR of action potentials recorded from cells growing on devices enhanced with nano-porous platinum electrodeposition and cells on control devices with gold electrodes were compared. Activity within neuronal networks / beat frequencies of cell patches on electrodes were monitored over the course of 2-3 weeks. The methods explored in this study not only decreases impedance of microelectrodes to increase SNR but also increases cell-electrode coupling by warranting the placement of cells next or on the electrodes by precision patterning. Future studies will aim to optimize electrode sensitivity using novel electrode materials, the porosity of different materials, and characterization using emerging analytical methods. Eventually these devices will provide in vitro systems to monitor both acute and chronic effects of drugs and toxins as well as to perform functional studies under physiological or induced pathophysiological conditions that mimic in vivo damages. Figure 1: Electrogenic cells patterned near nano-porous platinum electrode Figure 1

physica status solidi (a)Volume 66, Issue 1 p. K31-K35 Short Note Single longitudinal mode operation due to excessive gain suppression? A comparative study of laser structures with and without optical confinement W. Rühle, W. Rühle Instituto de Fisica „Gleb Wataghin”︁, Universidade Estadual de Campinas On leave from the Physikalisches Institut der Universität Stuttgart.Search for more papers by this authorP. Brosson, P. Brosson Instituto de Fisica „Gleb Wataghin”︁, Universidade Estadual de Campinas Search for more papers by this author W. Rühle, W. Rühle Instituto de Fisica „Gleb Wataghin”︁, Universidade Estadual de Campinas On leave from the Physikalisches Institut der Universität Stuttgart.Search for more papers by this authorP. Brosson, P. Brosson Instituto de Fisica „Gleb Wataghin”︁, Universidade Estadual de Campinas Search for more papers by this author First published: 16 July 1981 https://doi.org/10.1002/pssa.2210660156 Caixa Postal 1170, 13100 Campinas, Sǎo Paulo, Brazil. AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Volume66, Issue116 July 1981Pages K31-K35 RelatedInformation

Measuring pH level on foetal scalp is a common practice to prevent the risk of acidosis in monitoring foetal health during labour. However, the current technique of collecting capillary sample (blood gas analyser method) is not continuous and has high rate of failures. In this work, a prototype of miniaturized pH sensor based on porous Titanium Nitride (TiN) electrodes to get continuous measurement of pH has been developed, targeting foetal health monitoring. By exploiting wet etching techniques on flat TiN electrodes, porosity of the working surface area has been increased, exhibiting better properties and performances of our pH sensor. Precision of the developed TiN pH sensor is 0.03 in pH, which is close to the requirement of the application. The sensitivity is determined to be 67.06 mV/pH within pH range of interest from 6 to 8. In addition, the Potentiostatic Electro Impedance spectroscopy (PEIS) and Cyclic Voltammetry (CV) were recorded, showing a 6.3-time smaller impedance at 100 Hz frequency and a better conductivity compared to the flat TiN electrode. This work is under the next step of fabrication to transfer the system from glass onto a flexible Polyimide substrate and to be integrated into a specific miniaturised packaged device for in-vivo testing.

Event Abstract Back to Event Long-term culture and spontaneous recordings of primary neurons on conducting polymer microelectrode arrays Jolien Pas1, Dimitrios A. Koutsouras1 and George Malliaras1 1 Ecole Nationale Supérieure des Mines de Saint-Etienne, Bioelectronics, France Microelectrode arrays (MEAs) have widely been used as diagnostic tool to improve our understanding of the beautiful yet extremely complex nervous system. MEAs are used to record and stimulate brain tissue or neuronal cells, thereby providing a platform for both in vivo and in vitro experiments. To circumvent the complexity of the nervous system, in vitro measurements enable us to simplify the biological environment and help us to attain specific essential neuronal knowledge. By using one of the most promising conducting polymers poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT: PSS), unique MEAs are developed[1] with an ideal dual character for neuronal interfacing due to their potential to conduct both electronic and ionic carriers. Nevertheless, it remains a challenge to culture and maintain a healthy primary neural cell culture over a long period of time to record their spontaneous electrical cell activity. Here we show long-term survival of primary cortex cells on PEDOT:PSS coated MEAs and show high quality recording of electrophysiological signals. We find that the combination of dropcasting poly-D-lysine (PDL, 50 µg/mL) and laminin (20 µg/mL) provide a vital adhesive support for E18 cortical rat neurons. This enables culture of these cells for more than 28 days which is followed by simultaneous multichannel recordings of spontaneous signals at 1 kHz with an average amplitude of 100 µV. Moreover, with the addition of bicuculline (50 µM) as a GABAA-antagonist during recording, bursts of action potentials are obtained similar to previously demonstrated results of Arnold et al[2]. It means that we have created the necessary cell-friendly environment for culture of such fragile primary neurons on state-of-the art MEAs. This work thereby paves the way towards a more diagnostic and eventually potentially therapeutic PEDOT:PSS platform to enable the study of neural electrophysiology and neuropathology.

Over the past century, robust methods were developed that enable the isolation, culture, and dynamic observation of mammalian neuronal networks in vitro. But even if neuronal culture cannot yet fully recapitulate the normal brain, the knowledge that has been acquired from these surrogate in vitro models is invaluable. Indeed, neuronal culture has continued to propel basic neuroscience research, proving that in vitro systems have legitimacy when it comes to studying either the healthy or diseased human brain. Furthermore, scientific advancement typically parallels technical refinements in the field. A pertinent example is that a collective drive in the field of neuroscience to better understand the development, organization, and emergent properties of neuronal networks is being facilitated by progressive advances in micro-electrode array (MEA) technology. In this chapter, we briefly review the emergence of neuronal cell culture as a technique, the current trends in human stem cell-based modeling, and the technologies used to monitor neuronal communication. We conclude by highlighting future prospects that are evolving specifically out of the combination of human neuronal models and MEA technology.

Whether it is about harnessing living cells as sensing elements or basic cell research, microelectrode arrays (MEAs) can be used for recording electrical response of various cell types in vitro, especially cardiac and neuronal cells. Even if commercial MEAs with standard layouts are easily available, several research groups end up fabricating or at least designing their own MEAs to meet better the needs of their specific research questions. In theory, the resistance of the tracks that connect the electrodes to the contact pads is several orders of magnitude smaller than the total electrode impedance, and thus there should be plenty of room to play with the track dimensions and conductivity. Still, relatively large track widths, thicknesses and highly conductive (= expensive) track materials are usually favored - just in case. In this study, we tested several track widths, thicknesses and materials. The results confirmed that playing too safe with the tracks has been unnecessary and, on the other hand, one cannot gain any benefit by exaggerating the track dimensions or conductivity. On the contrary, by reducing the track dimensions and choosing less conductive materials one could e.g., get better visibility and cost savings as well as improve the adhesion of microfluidic add-ons.

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Recent technological advances have introduced diverse engineered nanoparticles (ENPs) into our air, water, medicine, cosmetics, clothing, and food. However, the health and environmental effects of these increasingly common ENPs are still not well understood. In particular, potential neurological effects are one of the most poorly understood areas of nanoparticle toxicology (nanotoxicology), in that low-to-moderate neurotoxicity can be subtle and difficult to measure. Culturing primary neuron explants on planar microelectrode arrays (MEAs) has emerged as one of the most promising in vitro techniques with which to study neuro-nanotoxicology, as MEAs enable the fluorescent tracking of nanoparticles together with neuronal electrical activity recording at the submillisecond time scale, enabling the resolution of individual action potentials. Here we examine the dose-dependent neurotoxicity of dextran-coated iron oxide nanoparticles (dIONPs), a common type of functionalized ENP used in biomedical applications, on cultured primary neurons harvested from postnatal day 0–1 mouse brains. A range of dIONP concentrations (5–40 µg/ml) were added to neuron cultures, and cells were plated either onto well plates for live cell, fluorescent reactive oxidative species (ROS) and viability observations, or onto planar microelectrode arrays (MEAs) for electrophysiological measurements. Below 10 µg/ml, there were no dose-dependent cellular ROS increases or effects in MEA bursting behavior at sub-lethal dosages. However, above 20 µg/ml, cell death was obvious and widespread. Our findings demonstrate a significant dIONP toxicity in cultured neurons at concentrations previously reported to be safe for stem cells and other non-neuronal cell types.

The lifetime of three types of AMTEC electrodes is predicted from the rate of grain growth in the electrode. Grain size is related to electrode performance, allowing performance to be correlated with grain growth rate. The rate of growth depends on physical characteristics of each material, including the rates of surface self-diffusion and molecule mobility along grain boundaries. Grain growth rates for molybdenum, rhodium-tungsten and titanium nitride electrodes have been determined experimentally and fit to models in order to predict operating lifetimes of AMTEC electrodes. For lifetimes of 10 years or more, RhxW electrodes may be used at any operating temperature supportable by the electrolyte. TiN electrodes may be used in AMTEC cells only at operating temperatures under 1150 K, and Mo may be used only below 1100 K.

The collection of good quality extracellular neuronal spikes from neuronal cultures coupled to Microelectrode Arrays (MEAs) is a binding requirement to gather reliable data. Due to physical constraints, low power requirement, or the need of customizability, commercial recording platforms are not fully adequate for the development of experimental setups integrating MEA technology with other equipment needed to perform experiments under climate controlled conditions, like environmental chambers or cell culture incubators. To address this issue, we developed a custom MEA interfacing system featuring low noise, low power, and the capability to be readily integrated inside an incubator-like environment. Two stages, a preamplifier and a filter amplifier, were designed, implemented on printed circuit boards, and tested. The system is characterized by a low input-referred noise (<1 μV RMS), a high channel separation (>70 dB), and signal-to-noise ratio values of neuronal recordings comparable to those obtained with the benchmark commercial MEA system. In addition, the system was successfully integrated with an environmental MEA chamber, without harming cell cultures during experiments and without being damaged by the high humidity level. The devised system is of practical value in the development of in vitro platforms to study temporally extended neuronal network dynamics by means of MEAs.

Cortical neuronal activity depends on a balance between excitatory and inhibitory influences. Culturing of neurons on multi-electrode arrays (MEAs) has provided insight into the development and maintenance of neuronal networks. Herein, we seeded MEAs with murine embryonic cortical/hippocampal neurons at different densities (<150 or >1000 cells mm−2) and monitored resultant spontaneous signaling. Sparsely seeded cultures displayed a large number of bipolar, rapid, high-amplitude individual signals with no apparent temporal regularity. By contrast, densely seeded cultures instead displayed clusters of signals at regular intervals. These patterns were observed even within thinner and thicker areas of the same culture. GABAergic neurons (25% of total neurons in our cultures) mediated the differential signal patterns observed above, since addition of the inhibitory antagonist bicuculline to dense cultures and hippocampal slice cultures induced the signal pattern characteristic of sparse cultures. Sparsely seeded cultures likely lacked sufficient inhibitory neurons to modulate excitatory activity. Differential seeding of MEAs can provide a unique model for analyses of pertubation in the interaction between excitatory and inhibitory function during aging and neuropathological conditions where dysregulation of GABAergic neurons is a significant component.

This paper demonstrates a novel and facile technique for the production of microelectrode arrays based on either TiO2 or metal nanoparticles, which combines photolithography and photocatalytic deposition. A procedure that involves photolithographic selective decomposition of superhydrophobic n-octadecyltriethoxysilane (ODS) has been developed to create superhydrophobic/superhydrophilic TiO2 patterns (i.e., microelectrode arrays based on TiO2 nanoparticles). The generated TiO2 patterns can function as molecular microtemplates, in which elevated metal nanoparticle-based microelectrode arrays are produced by photocatalytic deposition to form microelectrode arrays based on metal nanoparticles. Microscopy and atomic force microscopy have demonstrated that the metal nanoparticles grow site-selectively inside the TiO2 microtemplates. This developed approach can be used to create microelectrode arrays composed by either TiO2 nanoparticles or various metal nanoparticles, such as gold, silver, and platinum, with different patterns. The electrochemical behavior of the as-prepared microelectrode arrays has also been characterized by cyclic voltammetry.

Surface Functionalized Titanium Nitride Electrode for CMOS Compatible Bioelectronic Devices.