The effect of anticoagulation on platelet size stability was studied using blood collected in seven different anticoagulants and stored at room temperature for up to eight hours. The mean platelet volume (MPV) value was most stable in blood collected in 15% ACD and ACD/Na2EDTA. In blood collected in Na2EDTA, K3EDTA, or 11.9% ACD, there was an increase in MPV in the first two hours, after which the MPVs remained stable up to eight hours. Sodium citrate and heparin proved unreliable for the measurement of platelet volume. Platelet counts were stable (less than 5% variation) in all anticoagulants except heparin, which had 16% variation for the eight hours of study. Simultaneously, RBC counts and mean corpuscular volume (MCV) measurements were stable in all seven anticoagulants, with sodium citrate producing the most variation. A negative correlation was observed between MCV and pH of the anticoagulated blood. WBC counts showed less than 3% variation in all anticoagulants except sodium citrate and heparin. Separate experiments demonstrated that electrolyte composition, pH, tonicity, and method of calcium chelation all influenced the stability of the MPV. Of the anticoagulants studied, ACD/Na2 EDTA appeared to provide the best conditions of anticoagulation for both routine clinical and research laboratory measurement of the MPV. It inhibited platelet activation but left the platelets in their normal discoid shape. Platelets could be removed from the anticoagulant and studied in functional assays for up to eight hours after blood drawing. Both platelet counts and MPVs remained stable in blood collected in ACD/Na2 EDTA anticoagulant for up to eight hours at room temperature. In 52 volunteers studied, an inverse correlation (r = -0.72, P less than 0.001) was observed between platelet count and MPV, suggesting that the circulating platelet mass may be a more important indicator of platelet homeostasis than either the platelet count or the mean platelet volume alone.