To determine the osmotic limits of human platelets, cells in plasma were exposed to either hypertonic or hypotonic conditions. Platelet rich plasma (PRP) was placed in dialysis tubes which were then immersed into non-isotonic solutions at room temperature. When the cells were exposed to concentrations of 150 mOsm or 790 mOsm and returned to isotonic condition, a substantial proportion of platelets lost their discoid shape and assumed spherical or dendritic form. The decrease in morphologic score was gradual towards either side of isotonicity. The loss in the number of platelets was independent of the osmolalities and averaged 12%. The drop in pH was small. Platelet aggregation induced by calcium ionophore (A23187) was not changed when the cells were exposed to 150 or 750 mOsm and returned to isotonicity, but aggregation induced by ADP or collagen was greatly reduced. The response of platelets to the hypotonic stress reversal assay was also impaired. These results suggest that the ability of human platelets to tolerate non-isotonic conditions is limited and that care should be exercised in the addition and removal of cryoprotective agents during the process of platelet freezing.