After incubation of Ureaplasma urealyticum serotype 8 in the presence of 3H-labeled palmitic acid, about 25 acylated proteins were detected by electrophoresis and fluorography. Of these, at least six were shown to be antigenic by immunoprecipitation of solubilized palmitate-labeled cells with a homologous polyclonal serum. These six included the serotype 8-specific surface-expressed 96-kDa antigen. After phase partition of palmitate-labeled cells with Triton X-114, all but six acylated proteins partitioned entirely into the detergent phase. The others, including the 96-kDa antigen, partitioned preferentially into the detergent phase and were apparently amphipathic. These results are consistent with the acylated proteins being mainly membrane associated.