1Universitaet Karlsruhe (TH), Zoologisches Institut, Zell- und Neurobiologie, Haid-und-Neu-Strasse 9, 76131 Karlsruhe, Germany.
2Max-Planck-Institut für Entwicklungsbiologie, Spemannstrasse 35, 72076 Tuebingen, Germany.
Graded distributions of proteins are pivotal for many signaling processes during development, such as morphogenesis, cell migration, and axon guidance. Here, we describe a technique to fabricate substrate-bound stepwise protein gradients by means of a microfluidic network etched into a silicon wafer with an array of parallel 14-micrometer–wide channels, which can be filled with a series of arbitrarily chosen protein solutions. In a subsequent microcontact printing step, the protein pattern is transferred onto a surface and is used as a substrate for cell culture. Cellular responses to a defined microscopic pattern of a protein, such as guided axonal outgrowth and directed migration, cell polarization, changes in morphology, and signaling, can be thus studied in a controlled in vitro environment.