Human type 1 diabetes is an autoimmune disease resulting from T-cell-mediated destruction of pancreatic islet beta cells. Maternal environment has been suggested to be important in the development of diabetes. In this study, to investigate the effect of maternal nutrition, in particular the essential fatty acid (EFA) ratio (n-6/n-3) and composition, on the development of type 1 diabetes in the offspring, we prepared different kinds of chows with different n-6/n-3 ratios, and provided them to pregnant and lactating mothers and post-weaning female offspring of non-obese diabetic (NOD) mice, a type 1 diabetes model. The EFA ratio in breast milk and serum of NOD dams became nearly the same as that of the maternal diet. Overt diabetes was not suppressed in the offspring from dams provided with a diet with an n-6/n-3 ratio of 14.5 during gestation and lactation, but it was strongly suppressed in those from dams provided with a diet with an n-6/n-3 ratio of 3.0 during the same period. At 2 and 4 weeks after birth, insulin autoantibodies (IAA) were detected in the offspring from dams provided with a diet with an n-6/n-3 ratio of 14.5, and not detected in those from dams provided with a diet with an n-6/n-3 ratio of 3.0. These findings suggest that n-6/n-3 ratio of the maternal diet during gestation and lactation rather than that of the diet intake by offspring after weaning strongly affects the development of overt diabetes in NOD mice.