The adsorption of proteins on poly(2-hydroxyethyl methacrylate) (PHEMA) brushes was systematically studied by quartz crystal rnicrobalance (QCM) and fluorescence microscopy as a function of graft density and protein size. The graft density a (chains/nm(2)) ranged from 0.007 (dilute or semidilute brush regime) to 0.7 (concentrated brush regime), and the protein size ranged from 2 to 13 nm in an effective diameter. The lowest-density brush (sigma = 0.007) adsorbed all the tested four proteins, while the highest-density brush (a = 0.7) adsorbed none of them. The middle-density brush (sigma = 0.06) showed an intermediate behavior, adsorbing the smallest two proteins but effectively repelling the largest two. PHEMA cast films adsorbed a probe protein with the adsorbed amount increasing approximately proportionally to the film thickness, indicating that the adsorption mainly occurred in the bulk of the film. The noted results for the brushes support the idea of size-exclusion effect, an effect characteristic of concentrated polymer brushes, in which the graft chains are highly extended and highly oriented so that large molecules, sufficiently large compared with the distance between the nearest-neighbor graft points, are physically excluded from the entire brush layer. In this regard, the behavior of the lowest-density brush should be essentially similar to that of the cast film, as was in fact observed.