Novel high-resolution micropatterning for neuron culture using polylysine adsorption on a cell repellant,...

The ability to organize individual neurons and their processes in culture provides important benefits to both basic neuroscience research applications and the development of biomedical microdevices. While numerous methods have been used to produce such micropatterning of neurons and cells in general, there has yet been no method to simultaneously provide high-resolution patterns with high compliance of cells to desired patterns and good manufacturability. To develop such a process, this work used a plasma polymerized, nonfouling poly ethylene oxide (PEO)-like film to provide a cell repellant substrate on which cell adhesive micropatterns can be selectively laid down. While the use of plasma polymerized, organic films have been used for cell micropatterning, this process exploits the often-overlooked tendency for the surface of this PEO-like material to adsorb polylysine from aqueous solution while remaining nonfouling with respect to other species, such as bovine serum albumin (BSA) and immunoglobulin G (IgG). When the adsorption of polylysine was enhanced by brief plasma oxidation, which slightly alters the surface chemistry of the material, simple photolithographic liftoff could be used to micropattern stable, cell adhesive areas on an otherwise cell repellant background. We showed that the application of photolithography itself on the PEO-like material did not alter its chemical properties, nor did it result in the erosion of the micropatterned polylysine on its surface. Hippocampal neurons from embryonic mice flourished on these micropatterned substrates and exhibited viability comparable to neurons cultured on polylysine coated glass. Furthermore, the compliance of cell bodies and outgrowing neurites to the micropatterns was nearly perfect. In addition to providing cell adhesive regions, the micropatterned polylysine coating also served as a template mediating the immobilization of other bioactive species such as IgG and laminin. Using this "piggybacking" of laminin on polylysine, we were also able to culture and micropattern retinal ganglion cells (RGC).