The effect of ouabain on the osmotic resistance of red cells from 17 splenectomized patients with congenital or hereditary spherocytosis (HS), from 5 of their relatives suspected of having a subclinical form of the disease and from unsplenectomized and splenectomized normal controls was studied. In red cells from the HS patients and the splenectomized controls a small ouabain-induced decrease in osmotic resistance was seen, whereas in the unsplenectomized controls no such change was observed. The osmotic resistance of red cells from the 5 relatives, on the other hand, was significantly decreased by ouabain. The usefulness of ouabain in unmasking HS in its subclinical form was further tested by studying monovalent cation influx into red cells. The active influx of 86Rb+ into red cells from HS suffers was significantly increased. A similar degree of ouabain-induced inhibition of 86Rb+ transport was seen in all groups studied. The passive influx of 22Na+, which was also increased in HS red cells, was not affected by ouabain in any of the four groups.