Sperm from 10 fertile donors were incubated for 8 h in a capacitating medium (BWW 3.5% HSA) and treated with 1.25 microliters of 2 mM Ca-ionophore solution. Sperm motion was analysed using the Hamilton-Thorn system before and after incubation and treatment. The acrosome reaction was detected with PSA-FITC labelling of the acrosome. Critical values of sperm movement parameters were defined for objective identification of hyperactive movement by the Hamilton-Thorn system. The incidence of hyperactive movement in the sperm suspensions was approximately 2%. No significant changes of hyperactive movement could be observed after the incubation period or the ionophore treatment, in contrast to a significant rise in the percentage of acrosome-reacted cells after ionophore treatment. The implications of these findings for monitoring hyperactive cells to test sperm function are discussed.