Human islet amyloid polypeptide (hIAPP) is co-secreted with insulin from pancreatic islet β cells. This peptide spontaneously aggregates in the form of fibrils, and amyloid deposits are associated with dead or degenerating β cells, a hallmark of noninsulin-dependent diabetes mellitus. We demonstrated that COS-1 cells transfected with vectors expressing hIAPP exhibited intracellular amyloid deposits that were associated with cell death (O'Brien, Butler, Kreutter, Kane, Eberhardt, Am J Pathol 1995, 147:609–616). To establish the mechanism of cell death, we transfected COS-1 cells with vectors expressing amyloidogenic hIAPP or nonamyloidogenic rat IAPP and mutant hIAPP constructs and assayed them for markers characteristic of apoptosis and necrosis by fluorescence-activated cell sorting analysis. Amyloidogenic hIAPP-transfected COS cells contained up to threefold more apoptotic cells present at 96 hours after transfection compared with the nonamyloidogenic vector controls. The hIAPP-induced apoptosis was negligible at 24 and 48 hours after transfection and was maximal at 96 hours which parallels the time course of amyloidogenesis. Immunohistochemical staining and confocal microscopy showed that hIAPP is localized with distinct clustering in the endoplasmic reticulum and Golgi apparatus with no discernable extracellular staining. These experiments provide direct evidence that intracellular hIAPP amyloid causes cell death by triggering apoptotic pathways.