The acidic phospholipid phosphatidylserine (PS) has been reported to have variable effects on in vitro hydroxyapatite proliferation. PS promotes in vitro mineralization in systems in which calcium-PS-phosphate complexes are allowed to form, and inhibits in vitro mineralization when incorporated into liposomes. To investigate these diverse effects, a Langmuir adsorption isotherm was used to determine the affinity of PS for hydroxyapatite crystals, based on binding of 14C-PS to synthetic hydroxyapatite crystals of specific surface 54 m2/g. Using this model, PS was found to bind to hydroxyapatite crystals with an affinity comparable to that of the amino acid phosphoserine (K = 3.33 ml/mumol). Coating the surface of hydroxyapatite seed crystals with PS reduced their rate of proliferation in a metastable calcium phosphate solution in which calcium-PS-phosphate complexes were previously shown to promote hydroxyapatite formation. The extent of inhibition of hydroxyapatite seeded growth was directly related to the proportion of the hydroxyapatite surface covered with PS. These data suggest that PS may have multiple effects on hydroxyapatite formation in situ, and that mineral-PS interactions can retard crystal proliferation.