Sensitive detection of protein adsorption to supported lipid bilayers by frequency-dependent capacitance measurements and microelectrophoresis.

In the first part, we report experiments which enable the sensitive detection of protein adsorption to lipid bilayers deposited onto chromium electrodes on glass substrates by frequency-dependent capacitance measurements. The sensitivity of the present type of sensor (better than 0.3 nm average protein layer thickness) is at least equivalent to that of ellipsometry. A high specific resistance of the supported bilayer of (1-5).10(5) omega.cm2 is achieved by deposition of a tightly packed (crystalline) cadmium arachidate monolayer in contact with the substrate, whereas the outer monolayer can be more loosely packed (fluid phase or state of fluid-solid coexistence) which is essential for the incorporation of receptors. In the present work, charged lipids are incorporated as nonspecific receptors for polylysine and cytochrome c. The capacitance measurements provide a very sensitive test of the tightness and the long-time stability of the supported bilayers and, in combination with ellipsometric thickness measurements, enable estimations of dielectric properties of protein layers (such as the permittivity). In the second part, we report first electrophoresis experiments in asymmetric bilayers on substrates which enable simultaneous measurements of lateral diffusion coefficients and frictional coefficients between monolayers. The potential application of the electrophoretic effect for the differentiation between different receptors and the amplification of signals in biosensors is discussed.

On the chemical modification of pacemaker electrodes and patterned surface functionalization of planar substrates.

We report on experiments towards the chemical modification of metal electrodes in order to enhance biocompatibility or improve cell adhesion properties. In the first example pacemaker electrodes were modified with a thin polysiloxane network which allowed for further derivatization with a poly(ethylene glycol) layer. The primary goal was to suppress inflammatory response of tissue after implantation of electrodes. FTIR, ESCA and a.c.-impedance spectroscopy show the integrity of the ultrathin membrane. No significant reduction of the electrode capacitance was observed, providing further proof for the deposition of a homogeneously thin membrane. The second example deals with the patterned chemical modification of planar surfaces. The goal was to eventually effect selective adhesion of electrosensitive cells above microelectrodes for stimulation and/or recording. First results demonstrate the compatibility of monolayer deposition techniques with common photolithography. It is thus possible to create surfaces with patterned chemical functionality. A gas-phase silylation process was developed in order to control more precisely surface hydration and reaction parameters than is possible with common solution-based silylation procedures.

[Subretinal microphotodiode array as replacement for degenerated photoreceptors?]. / Subretinales Mikrophotodioden-Array als Ersatz für degenerierte Photorezeptoren?

A survey is given on the status of developments, concerning a subretinal electronic microphotodiode array that aims at replacing degenerated photoreceptors. Various prototypes have been developed, tested, and implanted in various experimental animals up to 18 months. The fact that electrical responses were recorded from the visual cortex of pigs after electrical stimulation by subretinal electrodes and the fact that responses are also recorded in-vitro in degenerated rat retinae, shows the feasibility of this approach. However, there are a number of open questions concerning the biocompatibility, the long-time stability, and the type of transmitted image to be solved before application in patients can be considered.

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.

Two-dimensional microelectrophoresis in supported lipid bilayers.

We report the application of supported bilayers for two-dimensional microelectrophoresis. This method allows the lateral separation and accumulation of charged amphiphilic molecular probes in bilayers by application of an electric field parallel to the bilayer surface. Diffusion coefficient and mobility of the fluorescent probes are determined by observation of the fluorescence recovery after photobleaching (pattern bleaching). The diffusion coefficients and the mobilities of oppositely charged fluorescent probes in one bilayer can be determined independently from a single measurement. By analysis of the motion of charged and uncharged probes in one membrane one can distinguish between the motion caused by the electric field acting on the charge of individual probes and that caused by frictional forces due to electroosmosis.

On-chip electrophoretic accumulation of DNA oligomers and streptavidin.

A micro-chamber for electrophoretic accumulation of charged biomolecules has been designed and evaluated. The system is based on a chip with an array of planar focusing electrodes. Particular attention was devoted to a design which enables penetration of a large sample volume by the electric field of the focusing electrodes. General design principles for a cylindrically symmetrical arrangement of the focusing electrodes were derived. Accumulation of DNA oligomers and streptavidin in aqueous solution was demonstrated. The concentration of biomolecules in the centre of the chip was enhanced by up to a factor of 200. The major fraction of the total charge delivered during electrophoretic accumulation results from Faradaic processes. The maximum charge density deliverable without visible gas formation was determined. By careful control of the voltage and current density applied to the electrodes, evolution of gas bubbles could be avoided for the time required to accumulate analyte molecules in the centre of the micro-chamber. On-chip electrophoretic accumulation of biomolecules can be applied to sample pre-conditioning in lab-on-a-chip devices for analysis of DNA and protein samples.

Microdevices for separation, accumulation, and analysis of biological micro- and nanoparticles.

Microfabrication and performance of a novel microsystem for separation, accumulation and analysis of biological micro- and nanoparticles is reported. Versatile chip functions based on dielectrophoresis and microfluidics were integrated to isolate particles from complex sample solutions such as serum. A bead-based assay for virus detection is proposed. Separation of micro- and sub-mum beads employing dielectrophoretic deflector and bandpass structures is demonstrated. Individual antibody coated beads with hepatitis A virus bound to their surface were trapped by negative dielectrophoresis in a field cage and analysed by fluorescence microscopy.