To study the function behind the molecular arrangement of single integrins in cell adhesion, we designed a hexagonally close-packed rigid template of cell-adhesive gold nanodots coated with cyclic RGDfK peptide by using block-copolymer micelle nanolithography. The diameter of the adhesive dots is < 8 nm, which allows the binding of one integrin per dot. These dots are positioned with high precision at 28, 58, 73, and 85 nm spacing at interfaces. A separation of > or = 73 nm between the adhesive dots results in limited cell attachment and spreading, and dramatically reduces the formation of focal adhesion and actin stress fibers. We attribute these cellular responses to restricted integrin clustering rather than insufficient number of ligand molecules in the cell-matrix interface since "micro-nanopatterned" substrates consisting of alternating fields with dense and no nanodots do support cell adhesion. We propose that the range between 58-73 nm is a universal length scale for integrin clustering and activation, since these properties are shared by a variety of cultured cells.

Au-nanoclusters between 2 and 8 nm in diameter were deposited onto solid substrates in different pattern geometries. The basis of this approach is the self-assembly of polystyrene-b-poly[2-vinylpyridine (HAuCl 4)] diblock copolymer micelles into uniform monomicellar films on solid supports such as Si-wafers or glass cover slips. HAuCl 4 as metallic precursor or a single solid Au-nanoparticle caused by reduction of the precursor are embedded in the centre of diblock copolymer micelles. Subsequent hydrogen, oxygen or argon gas plasma treatment of the dry film causes deposition of Au-nanoparticles onto the substrate by entire removal of the polymer. The Au-dot patterns resemble the micellar patterns before the plasma treatment. Separation distances between the dots is controlled by the molecular weight of the diblock copolymers. The limitation of the separation distance between individual dots or the pattern geometry is overcome by combining self-assembly of diblock copolymer micelles with pre-structures formed by photo or e-beam lithography. Capillary forces of a retracting liquid film due to solvent evaporation on the pre-structured substrate push micelles in the corners of these defined topographies. A more direct process is demonstrated by applying monomicellar films as negative e-beam resist. Micelles that are irradiated by electrons are chemically modified and can hardly be dissolved from the substrate while non-exposed micelles can be lifted-off by suitable solvents. This process is also feasible on electrical isolating substrates such as glass cover slips if the monomicellar film is coated in addition with a 5 nm thick conductive layer of carbon before e-beam treatment. The application of cylindrical block copolymer micelles also allows for the formation of 4 nm wide lines which can be 1-50 µm long and also be organized in defined aperiodic structures.

This paper introduces an approach where the match of two different length scales, i.e., pattern from self‐assembly of block copolymer micelles (< 100 nm) and electron‐beam (e‐beam) writing (> 50 nm), allow the grouping of nanometer‐sized gold clusters in very small numbers in even aperiodic pattern and separation of these groups at length scales that are not accessible by pure self‐assembly. Thus, we could demonstrate the grouping of Au nanoclusters in different geometries such as squares, rings, or spheres.

A multistep procedure for creating nanohole‐patterned gold films (see Figure) is presented. The steps include the self‐assembly of metal‐loaded polystyrene‐block‐poly(2‐vinylpyridine) micelles on GaAs substrates, hydrogen gas plasma treatment, directional reactive ion etching, and gold sputtering. The size and separation of holes resemble that of the gold cluster pattern.

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