2-Hydroxyethyl methacrylate polymer brushes with various grafting densities and chain lengths were prepared through surface-initiated atom transfer radical polymerization. X-ray photoelectron spectra, ellipsometry measurement, contact angle measurement, and atom force microscope were used to characterize the prepared polymer brush. The biofouling assays of polymer brush were investigated by adhesion of Dunaliella tertiolecta, Navcular sp., and Bovine Serum Albumin protein and by static marine immersion field test. Besides, hydroxyl and sulfonate-terminated self-assembled monolayers, anionic charged 3-sulfopropyl methacrylate potassium salt polymer brush were prepared for comparison. Results suggest that the settlement of microorganisms can be largely reduced by polymer with enough polymer chain length and grafting density. More importantly, static immersion field tests indicate that hydrophilic polymer film with enough hydration layer thickness is necessary for long-term marine antifouling application. This comprehensive investigation is of great importance to understanding their influence on the adhesion of marine microorganism. (c) 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2018, 135, 46232.