Herein we report on a preparation and performance of stable, hydrophilic and biocompatible polymeric material suitable for functionalization of disposable substrates used in biosensors. This new material features COOH surface groups cross-linked with ethylene glycol molecules and was prepared in situ on disposable, plastic substrate by high-throughput and environmentally friendly technique called plasma-enhanced chemical vapor deposition (PECVD). The film is grafted to the plasma activated plastic by sequential deposition of tetraethylorthosilicate, forming a bonding layer, and mixed vapors of acrylic acid and diethyleneglycol dimethylether (AA/PEG) that provide the desired functional groups forming a sensing, contact layer. A superior performance of the AA/PEG coating as suitable material for substrates in biomedical devices was demonstrated in a model fluorescence linked immunosorbent assay. The results were compared with other commonly used surface materials prepared by wet chemistry methods. The unique characteristic of the AA/PEG film is that the immunoassay can be executed without the need for a blocking step, typically using albumins, without negative consequences on the bioassay results. In fact, the superior quality of the materials modified with AA/PEG film was highlighted by improving the sensitivity of an immunoassay by two orders of magnitude when compared with substrates prepared by standard surface chemistry methods. © 2011 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2012.