Denice D. Denton

Micromachining of non-fouling coatings for bio-MEMS applications

Abstract Standard photolithography is used to pattern a poly (ethylene glycol) (PEG)-like polymer onto silicon substrates. The coating has excellent non-fouling properties and good adhesion to various substrate materials, such as silicon, oxide, nitride, gold, and platinum. This method allows precise control of the shape, size and alignment of the polymer, thus providing a reliable tool to pattern protein sheets as well as cell cultures. This method also enables the incorporation of patterned cell cultures with various predefined elements such as electrodes, channels, and sensors. To demonstrate the properties of our technique, we apply it to build cell cultures and to protect metallic electrodes from protein and cell adhesion. We show that the thin coatings provide excellent protection without compromising the conductivity of the electrodes.

High-aspect ratio submicrometer needles for intracellular applications

A processing technology is presented to produce high-aspect ratio submicrometer silicon needles suited for intracellular interfacing with living cells. Pillars are created using deep reactive ion etching, and the sharpening of the pillars is achieved by reactive ion etching. A simple polyimide-based micro-fabrication approach was developed to integrate the silicon needles with a larger silicon base designed to carry elements such as amplifier, battery or memory. The current design allows convenient handling of the device during implantation and minimal mechanical load on the implanted region. Prototype devices were tested for usability and animal compatibility.

Micro-Scale Cell Patterning on Nonfouling Plasma Polymerized Tetraglyme Coatings by Protein Microcontact Printing

Nonfouling thin films were prepared by the plasma deposition of tetraethylene glycol dimethyl ether (pp4G) on fluorinated ethylene propylene polymer (FEP) and glass substrates. Ordered cell patterns were created on these surfaces by microcontact printing of proteins. Pp4G was found to be stable in aqueous environments and resistant to an ethanol sterilization procedure, as verified by surface analysis. Pp4G also reduced nonspecific protein adsorption by more than 65-fold before and after sterilization. Despite the low adsorption of proteins to pp4G in solution, protein microcontact printing was achieved and we were able to print laminin, an adhesive extracellular matrix protein, from an elastomeric stamp onto pp4G. The printed laminin supported the attachment and spreading of cardiomyocytes and the nonprinted pp4G regions remained cell repulsive in culture conditions. Microscale patterns of cardiomyocytes were maintained on printed pp4G for more than 7 days. This cell patterning process should be viable for other cell types. The potential applications include tissue engineering and microdevices for biosensor, diagnostic, and pharmacological applications.

Selective attachment of multiple cell types on thermally responsive polymer

Programmable surface chemistry has been achieved by depositing a temperature sensitive polymer onto arrays of micro-fabricated metallic heaters. Activating a single heater causes a localized change in the device surface chemistry from non-fouling to fouling in an aqueous environment. Two types of proteins and two types of cells were used to demonstrate localized immobilization of proteins and cells on such surface. These experiments show, for the first time, selective cell attachments on thermally responsive polymer controlled by a micro heater array. It suggests a new approach to realize proteomic chips and cell chips.

Protein Patterning with Programmable Surface Chemistry Chips

Programmable surface chemistry has been achieved by depositing thermally responsive polymer (plasma polymerized N-isopropylacrylamide, ppNIPAM) onto arrays ofmicro-fabricated metallic heaters. Activating a single heater causes a localized change in the device surface chemistry from non-fouling to fouling in aqueous environment. Various proteins were used to demonstrate localized immobilization of proteins on the surface ofcoated micro-heater arrays. Additional uses ofthis technique include applications such as cell patterning, tissue engineering, self-assembly, etc.

Silicon micro-needles with flexible interconnections

A flexible polyimide-based interconnect scheme was developed to realize isolated needle-like microelectrodes. A simple fabrication approach allows the integration of micromachined silicon needles with a larger silicon base designed to carry elements such as amplifiers, battery or memory. The interconnecting scheme uses two polyimide layers to sandwich a metallic layer. The metal layer forms the electrical connection between the silicon base and the micro-electrodes, while the polyimide layers provide flexible insulation. The current design allows convenient handling of the device during implantation and minimal mechanical load on the implanted region. The device can conform to the surface of neural tissue and allows convenient interfacing with rugged and dynamic tissues. Prototype devices were tested for usability and animal-compatibility. The devices were implanted in sea slugs (Tritonia diomedea) and extracellular signals were acquired. Tritonia diomedea show full recovery from surgery and implantation, and survive up to a minimum of fourteen days with the ability to perform normal behaviors.

Controlled microtubules transport on patterned non-fouling surfaces

Described is a lithographic technique used to realize high resolution tracks of the motor protein kinesin inside flow cells. The process consists of patterns of plasma polymerized PEG-like non-fouling coating on a fouling glass substrate. When in contact with a coated surface, proteins adhere exclusively to the fouling areas, thus forming kinesin tracks suited for microtubule guidance. Parallel transport along smooth kinesin track was observed for over 250 /spl mu/m. A method has been developed to seal the substrates with molded silicon flow channels in order to allow convenient control of reagent introduction and immobilization processes. Complete kinesin and BSA immobilization protocols, comprised of initial rinse, protein introduction, immobilization and final rinse have been successfully demonstrated.

Intracellular Neuronal Recording with High Aspect Ratio MEMS Probes

Micro-machined silicon needles capable of penetrating through cell membranes were fabricated and tested for intracellular sensing applications. The fabricated needles have sharp tips (diameter 300 µm) and exhibit high mechanical strength. The needles were tested for extra- and intra-cellular neuronal recording applications. To prepare the needles for neuronal recording, they were coated with metal and their shanks insulated. Using these needles, we were able to obtain extremely localized extracellular signals and to perform first recordings with silicon based micro-probes from the inside of neurons.