Helicobacter pylori has a particular affinity to cholesterol. It is not known, however, whether other steroidal substances are bound as well. In order to characterize the specificity and nature of the H. pylori-steroid interaction, the affinity of H. pylori to cholesterol and several steroidal hormones was investigated. Seven strains of H. pylori (five reference strains, two wild strains) and one strain each of Staphylococcus epidermidis and Escherichia coli were cultured on a cholesterol-free medium. Cholesterol-free bacteria were incubated with cyclodextrin-mediated cholesterol and several cyclodextrin-mediated steroidal hormones (beta-estradiol, testosterone, progesterone, hydrocortisone, dexamethasone). The steroid contents of the bacteria were determined by gas liquid chromatography. High amounts of cholesterol were detected in all H. pylori strains, whilst steroidal hormones were not found. Neither S. epidermidis nor E. coli showed an appreciable amount of cholesterol in the chromatographic examinations. Bacterial pretreatment with proteinase K diminished cholesterol adsorption of H. pylori. These data indicate a specific affinity of H. pylori to cholesterol. This unique property might serve as a pathogenicity component enabling survival and colonization of H. pylori in the gastric environment.