Polyethylene glycol (PEG) is a well-known non-fouling material used for protein adsorption and cellular adhesion. Recently, we successfully fabricated plasma-polymerized PEG (PP-PEG) thin films on various substrates by using a plasma enhanced chemical vapor deposition method and PEG200 as a precursor. The structure of our PP-PEG thin film was found to be very similar to the chemical structure of a PEG polymer and also bore a non-fouling characteristic for numerous proteins and cells without in vitro or in vivo toxicity. In this study, we modified this PP-PEG thin film with a simple H2/He plasma treatment to change the non-fouling surface property to a fouling one for proteins and cells. To make the patterned surfaces for proteins or cells on the protein- or cell-resistant areas, depositing a new fouling PP thin film was unnecessary since it was possible to perform a simple H2/He plasma treatment through a metal shadow mask on the PP-PEG thin film. Various surface analysis techniques such as water contact angle, atomic force microscopy, X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry were used to confirm the drastic changes in the surface chemical properties resulting from the plasma treatment. These results show that a PP-PEG film, together with its plasma treatment through a shadow mask, is a very simple and useful patterning technique for various biological applications. Copyright © 2012 John Wiley & Sons, Ltd.