Chronic exposure to elevated saturated free fatty acid (FFA) levels has been shown to induce Endoplasmic Reticulum (ER) stress that may contribute to promoting pancreatic -cell apoptosis. Here, we compared the effects of FFAs on apoptosis and ER stress in human islets and two pancreatic -cell lines, rat INS-1 and mouse MIN6 cells. Isolated human islets cultured in vitro underwent apoptosis and markers of ER stress pathways were elevated by chronic palmitate exposure. Palmitate also induced apoptosis in MIN6 and INS-1 cells, although the former were more resistant to both apoptosis and ER stress. MIN6 cells were found to express significantly higher levels of ER chaperone proteins than INS-1 cells, which likely accounts for the ER stress resistance. We attempted to determine the relative contribution that ER stress plays in palmitate-induced beta-cell apoptosis. Although overexpressing GRP78 in INS-1 cells partially reduced susceptibility to thapsigargin, this failed to reduce palmitate-induced ER stress or apoptosis. In INS-1 cells palmitate induced apoptosis at concentrations that did not result in significant ER stress. Finally, MIN6 cells depleted of GRP78 were more susceptible to tunicamycin-induced apoptosis, but not to palmitate-induced apoptosis compared to control cells. These results suggest that ER stress is likely not the main mechanism involved in palmitate-induced apoptosis in -cell lines. Human islets and MIN6 cells were found to express high levels of stearoyl-CoA desaturase 1 compared to INS-1 cells, which may account for the decreased susceptibility of these cells to the cytotoxic effects of palmitate.