Defects in the androgen receptor cause a spectrum of abnormalities in genetic males ranging from phenotypic women with testicular feminization to men with minor defects in fertility and/or virilization. The diagnosis of androgen resistance can be quite cumbersome, including analysis of the family history, karyotyping, endocrine studies, measurement of androgen binding in genital skin fibroblasts, and, in some instances, sequencing of mutant cDNAs. Furthermore, androgen-binding studies may be normal in patients with qualitative receptor abnormalities or mutations in the DNA-binding domain of the receptor. To circumvent these difficulties, we have used a recombinant adenovirus to deliver an androgen-responsive reporter gene (mouse mammary tumor virus-luciferase) to fibroblasts cultured from genital skin from 12 normal controls and from eight individuals with complete testicular feminization. Following incubation with androgen (2 nM mibolerone) for 72 h, luciferase activity in normal fibroblasts increased > 10-fold (range 11-200-fold) in a manner that corresponded with the level of androgen receptor detected in ligand-binding assays. By contrast, luciferase activity increased negligibly in fibroblasts from individuals with testicular feminization (average = 1.2-fold increase). This assay permits a direct assessment of endogenous androgen receptor function in cells and should be a powerful aid in the diagnosis of androgen resistance.