Selective immobilization of proteins in well-defined patterns on substrates has recently attracted considerable attention as an enabling technology for applications ranging from basic research to biosensors and bioMEMS. In this work we demonstrate a low-cost and high throughput process for nanoscale, selective immobilization of proteins on patterned Si substrates, based on colloidal lithography and plasma processing in order to define the spots (< 300nm) where proteins are adsorbed. A close- packed monolayer of PS microparticles is deposited on oxidized Si substrates and is used as etching mask to define SiO 2 spots (on Si), which after plasma-induced chemical modification are appropriate for selective protein immobilization. The immobilized proteins are detected by means of confocal microscopy and evaluated by means of atomic force microscopy. Such nanoscale immobilized proteins can be successfully integrated into BioMEMS and microanalytical systems.