C. D. W. Wilkinson

The use of materials patterned on a nano- and micro-metric scale in cellular engineering

Biological cells form distinct groupings in tissue depending on their function. This is as essential in the regrowth of damaged tissue as it is in the development of the mature animal from its egg. For a number of years we have been exploring the effect of surfaces patterned with a topographic relief pattern and with patterns of bound active bio-molecules. The response of cells to micron-sized features, the response of cells to nano-metric features and the effects of soft materials are discussed.

Substratum nanotopography and the adhesion of biological cells. Are symmetry or regularity of nanotopography important

Animal cells live in environments where many of the features that surround them are on the nanoscale, for example detail on collagen molecules. Do cells react to objects of this size and if so, what features of the molecules are they responding to? Here we show, by fabricating nanometric features in silica and by casting reverse features in polycaprolactone and culturing vertebrate cells in culture upon them, that cells react in their adhesion to the features. With cliffs, adhesion is enhanced at the cliff edge, while pits or pillars in ordered arrays diminish adhesion. The results implicate ordered topography and possibly symmetry effects in the adhesion of cells. Parallel results were obtained in the adhesion of carboxylate-surfaced 2-microm-diameter particles to these surfaces. These results are in agreement with recent predictions from non-biological nanometric systems.

Switching fields and magnetostatic interactions of thin film magnetic nanoelements

Switching fields of magnetic elements with nanometric dimensions have been investigated by Lorentz microscopy using a transmission electron microscope. Acicular elements of Co and Ni80Fe20 were fabricated by electron beam lithography and lift-off techniques. They were 1.6–3.5 μm long, 200 nm wide, and 20–50 nm thick, with flat rectangular ends or triangular pointed ends, and were patterned in linear arrays with center-to-center spacing ranging from 7 μm to 250 nm. Switching fields and reversal behavior of the elements were found to depend strongly on the shape of the ends and, in a closely packed array, on element separation, thereby providing a way of controlling their magnetic properties.

Adhesion formation of primary human osteoblasts and the functional response of mesenchymal stem cells to 330 nm deep microgrooves

The surface microtexture of an orthopaedic device can regulate cellular adhesion, a process fundamental in the initiation of osteoinduction and osteogenesis. Advances in fabrication techniques have evolved to include the field of surface modification; in particular, nanotechnology has allowed for the development of experimental nanoscale substrates for investigation into cell nanofeature interactions. Here primary human osteoblasts (HOBs) were cultured on ordered nanoscale groove/ridge arrays fabricated by photolithography. Grooves were 330 nm deep and either 10, 25 or 100 μm in width. Adhesion subtypes in HOBs were quantified by immunofluorescent microscopy and cell–substrate interactions were investigated via immunocytochemistry with scanning electron microscopy. To further investigate the effects of these substrates on cellular function, 1.7 K gene microarray analysis was used to establish gene regulation profiles of mesenchymal stem cells cultured on these nanotopographies. Nanotopographies significantly affected the formation of focal complexes (FXs), focal adhesions (FAs) and supermature adhesions (SMAs). Planar control substrates induced widespread adhesion formation; 100 μm wide groove/ridge arrays did not significantly affect adhesion formation yet induced upregulation of genes involved in skeletal development and increased osteospecific function; 25 μm wide groove/ridge arrays were associated with a reduction in SMA and an increase in FX formation; and 10 μm wide groove/ridge arrays significantly reduced osteoblast adhesion and induced an interplay of up- and downregulation of gene expression. This study indicates that groove/ridge topographies are important modulators of both cellular adhesion and osteospecific function and, critically, that groove/ridge width is important in determining cellular response.

Integrated optical waveguiding structures made by silver ion-exchange in glass. 1: The propagation characteristics of stripe ion-exchanged waveguides; a theoretical and experimental investigation

An account of the formation and characterization of stripe waveguides formed by silver/sodium ion exchange is given. It is shown that the variation of effective index with guide width can be predicted with good accuracy using a computer model to both solve the exchange equation to obtain the refractive index profile and to calculate the modes of the resulting structure. It is found that while the use of an anodized aluminum mask results in low-loss waveguides, a metallic mask causes deposition of silver at the edges of the guide. The experimental methods used for aluminum anodization and for characterization of the waveguides are described in detail.

Switching of nanoscale magnetic elements

We have investigated the magnetic properties of ultra-small-patterned elements of Co and NiFe thin films. The elements were rectangular with an aspect ratio in the range 3.75–20. The smallest were 200×40 nm2 with 50 nm gaps between them, corresponding to an areal density of 27 Gbit/in2 if used as discrete-patterned media for magnetic recording. The elements were fabricated by electron-beam lithography and lift-off patterning and high-resolution magnetic images were obtained by Lorentz microscopy in a transmission electron microscope. In situ magnetization reversal experiments showed that the strong dependence of the switching field on element width extended to the smallest elements of both materials. The switching field for 40-nm-wide Co elements was 1200 Oe and for 40-nm-wide NiFe elements was 800 Oe. Element length and aspect ratio had little effect.

Lorentz microscopy of small magnetic structures (invited)

Domains and domain walls in micron and submicron sized magnetic elements can be studied at high resolution using Lorentz microscopy in the transmission electron microscope. In situ magnetizing experiments are possible in which magnetization reversal processes can be viewed directly in the presence of varying magnetic fields. These techniques have been used to investigate small magnetic structures fabricated by electron beam lithography on electron transparent membrane substrates. Patterned elements as small as 200 ×40 nm have been imaged magnetically. Detailed studies have been carried out into the properties of high aspect ratio (acicular) elements of Co and a soft NiFe alloy. It has been found that the coercivity increases as the elements become narrower, down to ultrasmall elements with a width of 40 nm. Element length has no effect so long as the aspect ratio is sufficiently high. Magnetization reversal in acicular elements is known to begin from the ends of the elements, therefore the shape of the ends—flat, elliptical, or pointed—has a significant effect on the coercivity. The magnetic environment of an element is also highly important in determining its properties. A one-dimensional array of closely spaced elements has the same average switching field as an isolated element but the spread in values is greatly increased when the gap between elements is made smaller than the width of an element. Adding rows of elements to make a two-dimensional array also has an effect, even if the rows are spaced further apart than the length of the elements.

Integrated optical ring resonators made by silver ion-exchange in glass

The first (to our knowledge) integrated optical ring resonators to be fabricated using silver ion-exchanged waveguides are reported. Both circular and racetrack shaped resonators have been made, both types being capable of high finesse (>15) and efficiency (>90%). The circular resonators are much more difficult to make, however, requiring a double-diffusion process and precise control of the ion-exchange. For this reason, the racetrack resonators have been the more successful and have behaved exactly as expected from the previous work on losses and directional couplers.

Grooved substrata facilitate in vitro healing of completely divided flexor tendons

Multiple grooved substrata with groove depth 5 μm were found to facilitate the healing of completely divided rat flexor tendons in vitro. Sections of tendons cultured on plain substrata showed only partial healing with incompletely sealed epitenon layers and immature thin collagen fibres. Tendons cultured on patterned substrata healed with complete restoration of the epitenon layer and reconstitution of the internal structure of collagen fibres. Epitenon fibroblasts isolated from the surface of rat flexor tendons were shown to be more sensitive to topographical features than fibroblasts of the same size BHK fibroblasts. They remained more elongated and better aligned to the groove direction than BHK cells. Multiple grooved substrata facilitated epitenon cell movement. Cells were found to move with higher speed on patterned substrata than on plain substrata. In summary, we conclude that the use of multiple grooved substrata promotes tendon healing in vitro and may find application in clinical practice in tendon repair.

Embossing of nanoscale features and environments

Abstract A process has been developed whereby nanoscale features can be easily and uniformly transferred into a thermoplastic with the use of embossing. Masters for the embossing procedure are made using electron beam lithography, dry etching and electroplating and with these dies features as small as 60nm have been produced in the polymer cellulose acetate. In addition, it is shown that the plastic cellulose acetate can be patterned using photolithography with sub 250nm UV light and that a conjunction of these two pattern transfer methods allows fabrication of multiple layered plastic devices. An application for this two layer process is presented where sub micron features can be easily and controllably placed at the bottom of trenches, a normally very difficult procedure.