Martin Stelzle

Novel thin film titanium nitride micro-electrodes with excellent charge transfer capability for cell stimulation and sensing applications

Microelectrodes with high specific charge transfer capability are an important prerequisite for high resolution stimulation, and recording of neuronal and muscular tissue. Novel thin film titanium nitride (TiN) microelectrodes were fabricated onto a microelectrode array by reactive sputtering in a nitrogen/argon atmosphere. The electrodes showed excellent charge transfer capacity of 40 mC/cm/sup 2/ and low ac-impedance along with high pulse stability. Scanning electron microscopy revealed a regular columnar morphology providing for a large internal surface area. With respect to their electrical and mechanical properties the TiN electrodes proved superior to electrodes produced by anodic oxidation of iridium. In this type of iridium oxide irregular cracks formed probably due to the high density ratio between metal and oxide. Furthermore, in spite of high charge capacity at very low frequencies (Q/sub cap/) the iridium oxide electrodes exhibited poor safe charge injection limit under pulsing conditions (Q/sub inj/).

“Artificial micro organs”—a microfluidic device for dielectrophoretic assembly of liver sinusoids

In order to study possible toxic side effects of potential drug compounds in vitro a reliable test system is needed. Predicting liver toxicity presents a major challenge of particular importance as liver cells grown in a cell culture suffer from a rapid loss of their liver specific functions. Therefore we are developing a new microfluidic test system for liver toxicity. This test system is based on an organ-like liver 3D co-culture of hepatocytes and endothelial cells. We devised a microfluidic chip featuring cell culture chambers with integrated electrodes for the assembly of liver sinusoids by dielectrophoresis. Fluid channels enable an organ-like perfusion with culture media and test compounds. Different chamber designs were studied and optimized with regard to dielectrophoretic force distribution, hydrodynamic flow profile, and cell trapping rate using numeric simulations. Based on simulation results a microchip was injection-moulded from COP. This chip allowed the assembly of viable hepatocytes and endothelial cells in a sinusoid-like fashion.

Multiwalled Carbon-Nanotube-Functionalized Microelectrode Arrays Fabricated by Microcontact Printing: Platform for Studying Chemical and Electrical Neuronal Signaling

A facile method is proposed for the deposition of multiwalled carbon nanotube (MWCNT) layers onto microelectrode arrays by means of a microcontact printing technique, leading to the fabrication of MEAs characterized by well defined electrical and morphological properties. Using polydimethyl siloxane stamps, produced from different mold designs, a flexibility of printing is achieved that provides access to microscale, nanostructured electrodes. The thickness of MWCNT layers can be exactly predetermined by evaluating the concentration of the MWCNT solution employed in the process. The electrode morphology is further characterized using laser scanning and scanning electron microscopy. Next, by means of impedance spectroscopy analysis, the MWCNT-electrode contact resistance and MWCNT film resistance is measured, while electrochemical impedance spectroscopy is used to estimate the obtained electrode-electrolyte interface. Structural and electrochemical properties make these electrodes suitable for electrical stimulation and recording of neurons and electrochemical detection of dopamine. MWCNT-functionalized electrodes show the ability to detect micromolar amounts of dopamine with a sensitivity of 19 nA μm(-1) . In combination with their biosensing properties, preliminary electrophysiological measurements show that MWCNT microelectrodes have recording properties superior to those of commercial TiN microelectrodes when detecting neuronal electrical activity under long-term cell-culture conditions. MWCNT-functionalized microelectrode arrays fabricated by microcontact printing represent a versatile and multipurpose platform for cell-culture monitoring.

MicroPrep: Chip-based dielectrophoretic purification of mitochondria

We have developed a microfluidic system – microPrep – for subcellular fractionation of cell homogenates based on dielectrophoretic sorting. Separation of mitochondria isolated from a human lymphoblastoid cell line was monitored by fluorescence microscopy and further characterized by western blot analysis. Robust high throughput and continuous long‐term operation for up to 60 h of the microPrep chip system with complex biological samples became feasible as a result of a comprehensive set of technical measures: (i) coating of the inner surfaces of the chip with BSA, (ii) application of mechanical actuators to induce periodic flow patterns, (iii) efficient cooling of the device to ensure integrity of organelle, (iv) a wide channel to provide for high fluidic throughput, and (v) integration of a serial arrangement of 10 dielectrophoretic deflector units to enable separation of samples with a high particle load without clogging. Hence, microPrep yields tens of micrograms of enriched and purified mitochondria within hours. Western blots of mitochondria fractions showed that contaminating endoplasmatic reticulum was reduced by a factor 6 when compared with samples prepared by state of the art centrifugation.

Electrical Properties of Micro-Photodiode Arrays for Use as Artificial Retina Implant

Micro-photodiode arrays have been developed which are intended to eventually replace the function of degenerated photoreceptor cells in the retina. Electric current generated by tiny micro-photodiodes is delivered to the neuronal network in the retina via micro-electrodes. Since the coupling between electrode and tissue is capacitive of nature, only transient signals may be used for stimulation. Therefore, high capacitance of the interface between electrodes and tissue is an important prerequisite for efficient charge transfer. In addition, the electrical properties of the micro-photodiodes as are reflected in current/voltage traces have a profound influence on the charge delivery. For the first time the electrical properties of the entire system consisting of micro-photodiode array, electrode/electrolyte interface and tissue were studied. Our results indicate charged balanced operation of the device and the presence of a non-zero electrode polarization. The latter may be avoided, if an active current sink is employed in order to draw current to discharge the electrode capacitance.

Towards plug and play filling of microfluidic devices by utilizing networks of capillary stop valves

Robust bubble-free priming of complex microfluidic chips represents a critical, yet often unmet prerequisite to enable their practical and widespread application. Towards this end, the usage of a network of capillary stop valves as a generic design feature is proposed. Design principles, numerical simulations, and their application in the development of a microfluidic cell culture device are presented. This chip comprises eight parallel chambers for the assembly and cultivation of human hepatocytes and endothelial cells. The inlet channel divides into cell chambers, after which the flows are reunited to a single chip outlet. Dimensions and geometry of channels and cell chambers are designed to yield capillary burst pressures sequentially increasing towards the chip outlet. Thus, progress of liquid flow through the device is predefined by design and enclosure of air bubbles inside the microfluidic structures is efficiently avoided. Capillary stop valves were designed using numerical simulations. Devices were fabricated in cyclic olefin polymer. Pressure during filling was determined experimentally and is in good agreement with data obtained from simulation.

A microsensor system to probe physiological environments and tissue response

Foreign body responses and bio-fouling caused by the physiological environment impair sensor performance due to alteration of the sensor/tissue interface. For in vivo applications longterm stability is a critical prerequisite and often affected due to host response towards the implant. In order to assess tissue response towards implants, we propose continuous measurements at the implant/tissue interface employing a microsensor device placed in contact with the chorioallantoic membrane (CAM) of the avian embryo. We introduce a biostable microsensor implant (MSI) to measure oxygen, pH and electrical impedance in situ. These parameters were chosen for their sensitivity with respect to the composition and properties of biological tissue. Micro fabrication technology in combination with electrochemical electrode functionalization was used to combine all sensors in a small planar array. The chorioallantoic membrane assay (CAM-assay) of avian ex ovo cultures served as a quasi-in vivo environment. Here we established an immune-active and -deficient in vivo model, enabling comparison between weak and strong immune responses in the same organism. A miniaturized potentiostat unit (“MiniPot”) was developed for controlling the MSI in humid culture environments. Here we performed continuous measurements of all sensor parameters at the implant/tissue interface with the microsensor device placed in contact with the CAM of the avian embryo.

Microfluidic chip system for the selection and enrichment of cell binding aptamers

Aptamers are promising cell targeting ligands for several applications such as for the diagnosis, therapy, and drug delivery. Especially, in the field of regenerative medicine, stem cell specific aptamers have an enormous potential. Using the combinatorial chemistry process SELEX (Systematic Evolution of Ligands by Exponential enrichment), aptamers are selected from a huge oligonucleotide library consisting of approximately 10(15) different oligonucleotides. Here, we developed a microfluidic chip system that can be used for the selection of cell specific aptamers. The major drawbacks of common cell-SELEX methods are the inefficient elimination of the unspecifically bound oligonucleotides from the cell surface and the unspecific binding/uptake of oligonucleotides by dead cells. To overcome these obstacles, a microfluidic device, which enables the simultaneous performance of dielectrophoresis and electrophoresis in the same device, was designed. Using this system, viable cells can be selectively assembled by dielectrophoresis between the electrodes and then incubated with the oligonucleotides. To reduce the rate of unspecifically bound sequences, electrophoretic fields can be applied in order to draw loosely bound oligonucleotides away from the cells. Furthermore, by increasing the flow rate in the chip during the iterative rounds of SELEX, the selection pressure can be improved and aptamers with higher affinities and specificities can be obtained. This new microfluidic device has a tremendous capability to improve the cell-SELEX procedure and to select highly specific aptamers.

Introduction to polymer-based solid-contact ion-selective electrodes-basic concepts, practical considerations, and current research topics.

AbstractThis review aims at providing an introductory overview for researchers new to the field of ion-selective electrodes. Both state of the art technology and novel developments towards solid-contact reference (sc-RE) and solid-contact ion selective electrodes (sc-ISE) are discussed. This technology has potentially widespread and important applications provided certain performance criteria can be met. We present basic concepts, operation principles, and theoretical considerations with regard to their function. Analytical performance and suitability of sc-RE and sc-ISE for a given application depend on critical parameters, which are discussed in this review. Comprehensive evaluation of sensor performance along this set of parameters is considered indispensable to allow for a well-founded comparison of different technologies. Methods and materials employed in the construction of sc-RE and sc-ISE, in particular the solid contact and the polymer membrane composite, are presented and discussed in detail. Operation principles beyond potentiometry are mentioned, which would further extend the field of ISE application. Finally, we conclude by directing the reader to important areas for further scientific research and development work considered particularly critical and promising for advancing this field in sensor R&D. Graphical Abstractᅟ

Characterization of orientation of perfluorostearic acid Langmuir–Blodgett multilayers by infrared spectroscopic methods

Abstract The molecular orientation of perfluorostearic acid [CF3(CF2)16COOH] deposited by Langmuir–Blodgett technique has been investigated by grazing incidence reflection absorption and transmission infrared spectroscopy. In the former, the electric field vector is normal to the film surface whereas in the latter, it is parallel to the surface. The intensities of the infrared bands that tend to exhibit perpendicular polarization, e.g. symmetric stretching of COO−, are much higher in reflection absorption than in transmission for deposited perfluorostearic acid. Therefore, these Fourier transform infrared results strongly support that the perfluorostearic acid molecules are preferentially oriented with their chain axes perpendicular to the substrate surface.