Atomic force microscopy (AFM) was used to study the structure of the staphylococcal alpha-hemolysin (alpha HL) oligomer formed in supported phospholipid bilayers. In contrast to the recent X-ray crystallographic demonstration of a heptameric stoichiometry for the oligomer formed in deoxycholate (DOC) micelles, the high-resolution unprocessed AFM images unequivocally revealed a hexamer in these phospholipid bilayers. Independent support of this hexameric stoichiometry was obtained from the measurements of the lattice constant in the AFM images and from gel electrophoresis. Therefore, alpha HL can form two different, energetically stable oligomers, which differ in at least stoichiometry but perhaps subunit structure as well. Furthermore, stable, incomplete oligomers were observed in the AFM images, which may be of relevance to the mechanism by which alpha HL damages the cell.