The platelet binding characteristics of platelet-activating factor (PAF), a basophil-derived lipid that causes aggregation and secretion in rabbit platelets, were studied. In an assay in which binding was quantitated by loss of PAF activity, suspensions of washed rabbit platelets rapidly removed PAF from solution. Rabbit erythrocytes, lymphocytes, and neutrophils also bound PAF, but their binding capacity was less than that of the platelets. PAF binding to rabbit platelets was saturable and dependent on temperature and concentration of PAF and platelets but not on the presence of extracellular Ca2+. Initial rates of PAF binding and platelet secretion were identical, although temperature dependence studies suggested that binding was not the rate-controlling step in PAF-induced platelet secretion. Analysis of binding kinetics suggested that a PAF concentrations saturable for platelet secretion, only some of the available platelet binding sites were occupied.