Premature and low-birth-weight infants usually require small-volume platelet transfusions to treat thrombocytopenia. Also, infants undergoing open-heart surgery with extracorporeal circulation and with compromised cardiac function are at risk for excessive intravascular volume. The small-volume platelet substitution can be achieved by dispensing an aliquot from the unit of a standard single-donor platelet concentrate (PC). Alternatively, there is an indication for volume reduction of PCs to maximize the number of platelets transfused in the smallest possible volume. We determined the spontaneous and induced activation of platelets before and after volume reduction in 20 consecutive single-donor-apheresis PCs. After a mean storage time of 2 days, the PCs were plasma-depleted by centrifugation. Spontaneous, adenosine diphosphate (ADP)-induced, and collagen-induced activation were determined by flow cytometry. Furthermore, ADP- and collagen-induced aggregation were measured. A total of 33.8% of platelets in standard PCs were activated spontaneously. Volume reduction of PCs led to a mild but significant increase of spontaneous activation of platelets (43.2%). Additionally, volume reduction resulted in an impaired ADP-induced aggregability of platelets, whereas collagen induction was unaffected. Transfusion of volume-reduced PCs is an effective alternative to use of standard PCs in patients at frequent risk for excessive intravascular volume, because equal volumes increase the platelet count twice as effectively.