Rat and mouse parotid acinar cells have been dispersed into small acinar groups and individual cells. Amylase release in response to secretagogues has been determined. Cytochemical localization of adenylate cyclase in these cells has demonstrated isoproterenol-stimulated adenylate cyclase in the same location as the reported in tissue slices, at the extracellular aspect of the lumenal plasma membrane: adenylate cyclase activity was also visualized in completely separate cells, restricted to the lumenal pole of the cell. These results support the validity of other localizations of this enzyme in intact tissue blocks, and illustrate cytochemical similarity of cells in dispersed preparations to cells in tissue slices. The specialized location of plasma membrane adenylate cyclase at the locus of secretory granule exocytosis is retained even without intercellular junctions.