Concerns have been raised whether natural and man-made chemicals might have the potential of interfering with the endocrine system. Especially interactions with sex hormone receptors are considered as a critical issue. Weak anti-androgenicity has been demonstrated for some environmental pollutants such as p,p'-DDE, and androgenic activity was found in feedlot and pulp mill effluents. In order to be able to screen for compounds with affinity for the androgen receptor (AR), we developed an AR binding assay using a recombinant AR as receptor source and the synthetic androgen methyltrienolone (R 1881) as ligand. Experiments were performed on 96-well microtitre plates. Following method optimization, compounds recently recommended for the validation of assays characterizing AR-mediated effects and those being used for the OECD validation of the Hershberger assay were employed amongst others to standardize the method. The assay readily detected and discriminated compounds with strong and weak affinity for the AR such as natural and synthetic androgens, anti-androgens in therapeutic use, and a variety of chemicals with weak anti-androgenic side effects, whereas in line with previous findings, AR binding properties of dibutylphthalate and its metabolites could not be demonstrated. Detergents interfered with receptor binding, but showed characteristic effects different from that of true AR binding compounds. The assay is simple and sensitive, avoids the use of animals as a receptor source, and should be of value when screening for endocrine-modulating compounds.