Neutrophil aggregation has been widely evaluated from changes in light transmission. Using direct particle counting, we demonstrated that light transmission does not accurately reflect aggregation, and showed that in contrast to previous reports, newborn neutrophils do not aggregate irreversibly. We developed a flow cytometric technique to measure the kinetics of neutrophil aggregation, including latent times for onset of aggregation, initial forward and reversal rates, and maximal extents of aggregation. The kinetics of neutrophil aggregation were related to changes in initial cell concentration, stir speed (shear), and activator type and concentration. Physiologic activators stimulated reversible aggregation, accompanied by an exponential decay in aggregatory potential with increasing time. The fraction of occupied activator receptors was found to correspond to the fraction of maximal rates or extent of aggregation. Monoclonal antibodies were used to show that neutrophil aggregation is mediated by the Mac-1 integrin (CD11b/CD18). Direct measurements of aggregation have enhanced our understanding of the (patho)physiologic process of neutrophil aggregation.