Summary.Sperm cell plasma membrane and the outer acrosomal membrane fuse profusely during the acrosome reaction. The process is triggered by extracellular signals that elicit several intracellular events leading ultimately to membrane fusion. We have developed a streptolysin O permeabilizing protocol that selectively affects the spermatozoon plasma membrane without causing a significant loss of the acrosomal content. Most of the acrosomal acid phosphatase remained sperm‐associated even after a 20 min incubation at 37°C. However, the presence of 100 μM Ca2+in the incubation buffer stimulates the release of the enzyme. The reaction was followed biochemically, measuring the acid phosphatase activity released to the medium and morphologically by the binding of fluorescein isothiocynate‐conjugated peanut agglutinin and by electron microscopy. The results show that the streptolysin O permeabilized spermatozoon is a promising model for studying the complex set of events mediating and regulating the acrosome reaction.Sperma