Vanessa Maybeck

Graphene Transistor Arrays for Recording Action Potentials from Electrogenic Cells

The development of the future generation of neuroprosthetic devices will require the advancement of novel solid-state sensors and actuators with a further improvement in the signal detection capability, a superior stability in biological environments, and a more suitable compatibility with living tissue. To date, interfacing of living cells and tissue with solid-state electronic devices has mainly been based on conventional silicon technology, in particular using Si metal-oxide-semiconductor fi eld-effect transistor (MOSFET) structures. [ 1 ] However, some of the drawbacks associated with this technology, such as its limited stability in aqueous environments [ 2 ] and a relatively high electrical noise, [ 3 ] have triggered the study of alternative materials and technologies. [ 4–11 ] In this respect, solution-gated fi eldeffect transistors (SGFETs) based on Si-nanowires, [ 4 ] AlGaN/ GaN heterostructures, [ 5 ] H-terminated diamond, [ 6 , 7 ] carbon nanotubes, [ 8 ] and, more recently, graphene [ 9–11 ] have been investigated as sensing devices. Among these materials, graphene is a particularly attractive candidate for bioelectronic applications, due to its remarkable physical and chemical properties. The extremely high charge carrier mobility in graphene [ 12 ] leads to a fi eld-effect transistor (FET) performance that is superior to most known semiconductors. [ 13 ] In addition, graphene is known to possess good chemical stability [ 14 ] and biocompatibility, [ 15 ] which is crucial not only for integration with biological systems, but also for the operation of fi eld-effect devices without a protective dielectric layer. Furthermore, the facile integration of graphene electronics with fl exible substrates paves the way for the development of fl exible devices, an important requirement for the design of biomedical implants with reduced tissue damage and scarring. [ 16 ] The use of graphene-based solutiongated fi eld-effect transistors (G-SGFETs) for the detection of cell signals has already been demonstrated on a fundamental level by using a single transistor on exfoliated graphene. [ 8 ] Despite this promising preliminary result, a successful graphene-based technology for the applications envisioned above requires the demonstration of arrays of graphene transistors, which can

Nanostructured gold microelectrodes for extracellular recording from electrogenic cells

We present a new biocompatible nanostructured microelectrode array for extracellular signal recording from electrogenic cells. Microfabrication techniques were combined with a template-assisted approach using nanoporous aluminum oxide to develop gold nanopillar electrodes. The nanopillars were approximately 300-400 nm high and had a diameter of 60 nm. Thus, they yielded a higher surface area of the electrodes resulting in a decreased impedance compared to planar electrodes. The interaction between the large-scale gold nanopillar arrays and cardiac muscle cells (HL-1) was investigated via focused ion beam milling. In the resulting cross-sections we observed a tight coupling between the HL-1 cells and the gold nanostructures. However, the cell membranes did not bend into the cleft between adjacent nanopillars due to the high pillar density. We performed extracellular potential recordings from HL-1 cells with the nanostructured microelectrode arrays. The maximal amplitudes recorded with the nanopillar electrodes were up to 100% higher than those recorded with planar gold electrodes. Increasing the aspect ratio of the gold nanopillars and changing the geometrical layout can further enhance the signal quality in the future.

Boron-Doped Nanocrystalline Diamond Microelectrode Arrays Monitor Cardiac Action Potentials

The expansion of diamond-based electronics in the area of biological interfacing has not been as thoroughly explored as applications in electrochemical sensing. However, the biocompatibility of diamond, large safe electrochemical window, stability, and tunable electronic properties provide opportunities to develop new devices for interfacing with electrogenic cells. Here, the fabrication of microelectrode arrays (MEAs) with boron-doped nanocrystalline diamond (BNCD) electrodes and their interfacing with cardiomyocyte-like HL-1 cells to detect cardiac action potentials are presented. A nonreductive means of structuring doped and undoped diamond on the same substrate is shown. The resulting BNCD electrodes show high stability under mechanical stress generated by the cells. It is shown that by fabricating the entire surface of the MEA with NCD, in patterns of conductive doped, and isolating undoped regions, signal detection may be improved up to four-fold over BNCD electrodes passivated with traditional isolators.

Versatile Flexible Graphene Multielectrode Arrays

Graphene is a promising material possessing features relevant to bioelectronics applications. Graphene microelectrodes (GMEAs), which are fabricated in a dense array on a flexible polyimide substrate, were investigated in this work for their performance via electrical impedance spectroscopy. Biocompatibility and suitability of the GMEAs for extracellular recordings were tested by measuring electrical activities from acute heart tissue and cardiac muscle cells. The recordings show encouraging signal-to-noise ratios of 65 ± 15 for heart tissue recordings and 20 ± 10 for HL-1 cells. Considering the low noise and excellent robustness of the devices, the sensor arrays are suitable for diverse and biologically relevant applications.

On-chip optical stimulation and electrical recording from cells

Abstract. We present an optoelectrical device capable of in vitro optical stimulation and electrophysiological recording. The device consists of an array of micropixellated InGaN light-emitting diodes coupled to a custom-made ultrathin planar microelectrode array. Cells can be cultured directly on the chip for short- and long-term electrophysiological experiments. To show the functionality of the device, we transfected a cardiomyocyte-like cell line (HL-1) with a light-sensitive protein channelrhodopsin. We monitored action potentials of individual, spontaneously beating, HL-1 cells growing on the chip by extracellular electrical recordings. On-chip optical stimulation was demonstrated by triggering network activity in a confluent HL-1 cell culture and visualized by calcium imaging. We see the potential of our system for electrophysiological experiments with optogenetically modified cells. Optical stimulation can be performed directly on the chip without additional optical components or external light sources.

Reconstitution of Fusion Proteins in Supported Lipid Bilayers for the Study of Cell Surface Receptor–Ligand Interactions in Cell–Cell Contact

Bioactive molecules such as adhesion ligands, growth factors, or enzymes play an important role in modulating cell behavior such as cell adhesion, spreading, and differentiation. Deciphering the mechanism of ligand-mediated cell adhesion and associated signaling is of great interest not only for fundamental biophysical investigations but also for applications in medicine and biotechnology. In the presented work, we developed a new biomimetic platform that enables culturing primary neurons and testing cell surface-receptor ligand interactions in cell-cell contacts as, e.g., in neuronal synapses. This platform consists of a supported lipid bilayer modified with incorporated neuronal adhesion proteins conjugated with the Fc-domain of IgG (ephrin A5 Fc-chimera). We extensively characterized properties of these protein containing bilayers using fluorescence recovery after photobleaching (FRAP), quartz crystal microbalance with dissipation (QCM-D), and immunostaining. We conclude that the Fc-domain is the part responsible for the incorporation of the protein into the bilayer. The biomimetic platform prepared by this new approach was able to promote neuronal cell adhesion and maintain growth as well as facilitate neuronal maturation as shown by electrophysiological measurements. We believe that our approach can be extended to insert other proteins to create a general culture platform for neurons and other cell types.

Graphene transistors for interfacing with cells: towards a deeper understanding of liquid gating and sensitivity

This work is focused on the fabrication and analysis of graphene-based, solution-gated field effect transistor arrays (GFETs) on a large scale for bioelectronic measurements. The GFETs fabricated on different substrates, with a variety of gate geometries (width/length) of the graphene channel, reveal a linear relation between the transconductance and the width/length ratio. The area normalised electrolyte-gated transconductance is in the range of 1–2 mS·V−1·□ and does not strongly depend on the substrate. Influence of the ionic strength on the transistor performance is also investigated. Double contacts are found to decrease the effective resistance and the transfer length, but do not improve the transconductance. An electrochemical annealing/cleaning effect is investigated and proposed to originate from the out-of-plane gate leakage current. The devices are used as a proof-of-concept for bioelectronic sensors, recording external potentials from both: ex vivo heart tissue and in vitro cardiomyocyte-like HL-1 cells. The recordings show distinguishable action potentials with a signal to noise ratio over 14 from ex vivo tissue and over 6 from the cardiac-like cell line in vitro. Furthermore, in vitro neuronal signals are recorded by the graphene transistors with distinguishable bursting for the first time.

Graphene Multielectrode Arrays as a Versatile Tool for Extracellular Measurements

Graphene multielectrode arrays (GMEAs) presented in this work are used for cardio and neuronal extracellular recordings. The advantages of the graphene as a part of the multielectrode arrays are numerous: from a general flexibility and biocompatibility to the unique electronic properties of graphene. The devices used for extensive in vitro studies of a cardiac-like cell line and cortical neuronal networks show excellent ability to extracellularly detect action potentials with signal to noise ratios in the range of 45 ± 22 for HL-1 cells and 48 ± 26 for spontaneous bursting/spiking neuronal activity. Complex neuronal bursting activity patterns as well as a variety of characteristic shapes of HL-1 action potentials are recorded with the GMEAs. This paper illustrates that the potential applications of the GMEAs in biological and medical research are still numerous and diverse.

Characterization of the mechanical properties of HL-1 cardiomyocytes with high throughput magnetic tweezers

We characterized the mechanical properties of cardiomyocyte-like HL-1 cells using our recently developed multi-pole magnetic tweezers. With the optimized design, both high force and high throughput are achieved at the same time. Force up to 100 pN can be applied on a 1 μm diameter superparamagnetic bead in a workspace with 60 μm radius, which is encircled symmetrically by 3 sharp magnetic tips. By adjusting the coil currents, both the strength and direction of force can be controlled. The result shows that both viscosity and shear elastic modulus of HL-1 cells exhibit an approximately log-normal distribution. The cells became stiffer as they matured, consistent with a transition from proliferating cells to contractile muscle tissue. Moreover, the mechanical properties of HL-1 cells show high heterogeneity, which agrees well with their physiological structure.

Interfacing neurons on carbon nanotubes covered with diamond

A recently discovered material, carbon nanotubes covered with diamond (DCNTs) was tested for its suitability in bioelectronics applications. Diamond shows advantages for bioelectronics applications (wide electro chemical window and bioinertness). This study investigates the effect of electrode surface shape (flat or three dimensional) on cell growth and behavior. For comparison, flat nanocrystalline diamond substrates were used. Primary embryonic neurons were grown on top of the structures and neither incorporated the structures nor did they grow in between the single structures. The interface was closely examined using focused ion beam (FIB) and scanning electron microscopy. Of special interest was the interface between cell and substrate. 5% to 25% of the cell membrane adhered to the substrate, which fits the theoretical estimated value. While investigating the conformity of the neurons, it could be observed that the cell membrane attaches to different heights of the tips of the 3D structure. However, the aspect ratio of the structures had no effect on the cell viability. These results let us assume that not more than 25% of cell attachment is needed for the survival of a functional neuronal cell.