Poly(propylene sulfide-bl-ethylene glycol) (PPS-PEG) is an amphiphilic block copolymer that spontaneously adsorbs onto gold from solution. This results in the formation of a stable polymeric layer that renders the surface protein-resistant when an appropriate architecture is chosen. The established molecular-assembly patterning by lift-off (MAPL) technique can convert a prestructured resist film into a pattern of biointeractive chemistry and a non-interactive background. Employing the MAPL technique, we produced a micron-scale PPS-PEG pattern on a gold substrate, and then characterized the patterned structure with time-of-flight secondary ion mass spectrometry (ToF-SIMS) and atomic force microscopy (AFM). Subsequent exposure of the PPS-PEG/gold pattern to protein adsorption (full human serum) was monitored in situ; SPR-imaging (i-SPR) shows a selective adsorption of proteins on gold, but not on PPS-PEG areas. Analysis shows a reduction of serum adsorption up to 93% on the PPS-PEG areas as compared to gold, in good agreement with previous analysis of homogenously adsorbed PPS-PEG on gold. MAPL patterning of PPS-PEG block copolymers is straightforward, versatile and reproducible, and may be incorporated into biosensor-based surface analysis methods. (C) 2008 Elsevier B.V. All rights reserved.