The subcellular distribution and characteristics of protein carboxymethylase (protein methylase II) and its methyl-acceptor proteins have been studied in human platelets and erythrocytes. Human platelets constitute a rich source of protein carboxymethylase with the majority of the active enzyme localized to the cytosol. In contrast, the highest specific activity of protein substrate(s) for the enzyme, i.e. methyl-acceptor proteins, appears to be associated with the particulate fraction containing indoleamine storage vesicles. Thrombin promotes a 60 per cent increase in the carboxymethylation of platelet protein after pulse labeling of the cells with [ 3H]methionine. Human erythrocytes also contain protein carboxymethylase which is localized almost entirely to the cytosol. Unlike platelet cytosol, erythrocyte cytosol appears to be devoid of methyl-acceptor proteins. Erythrocyte membrane ghosts, however, can be readily methylated and probably constitute the sole source of methyl-acceptor proteins in these cells.