Study of endocytosis in human erythrocytes and their ghosts provides an opportunity for the elucidation of the steps involved in membrane invagination and fusion. The specific preparation being studied should be described with great care since circumstances that produce endocytosis may differ among the several available preparations. For example, in resealed red ghosts the addition of 0.5 to 1 mM Ca stimulates Mg-ATP-induced endocytosis, whereas in intact erythrocytes Ca addition stimulates primaquine-induced endocytosis and inhibits vinblastine-induced endocytosis. Furthermore, Ca inhibits endocytosis in white ghosts. EDTA can produce endocytosis in white ghosts but not in resealed red ghosts. In studies with white ghosts, incubation with EDTA, trypsin, or Mg-ATP produced endocytosis, whereas the prior addition of an antispectrin antibody prevented endocytosis. When the white ghost vacuoles were harvested they were found to be depleted of spectrin. These observations lead to the hypothesis that endocytic vacuoles are formed in areas of the membrane that have been substantially freed of spectrin.