Bleeding complications are the principal cause of morbidity and mortality in infants treated with extracorporeal membrane oxygenation (ECMO). The whole blood activated clotting time (ACT) test is used universally to monitor heparin therapy during this procedure. To enhance our understanding of this test and improve our management of anticoagulation, we studied the relationship between the ACT and blood heparin concentration in nine infants during ECMO. The activated clotting time correlated with the simultaneously determined heparin concentration (r = .55, p less than .001 for all patients samples; r = .92, p less than .001 for mean patients values). Within the range of values found in our patients, platelet count, fibrinogen, and fibrin degradation products did not affect the ACT-heparin concentration relationship. However, the interpretation of an individual ACT result was limited by its low precision: the mean difference of duplicate determinations was 9.2%, and the estimation of heparin concentration by a single ACT had a coefficient of variation of 32%. Two commercially available techniques using different activators gave results that differed numerically but correlated well with each other. Both provided similar precision in the estimation of heparin concentration. The ACT is a low cost, bedside test whose accuracy and precision allow the achievement of target heparin concentrations required in infants during ECMO. Multiple determinations, either in duplicate or serially, are needed to achieve satisfactory precision. These data will be useful in designing future studies to determine the optimal serum heparin concentration to provide adequate anticoagulation, but avoid bleeding complications.