Inhibition of mitochondrial reactive oxygen species (ROS) and subsequent damage‐associated molecular patterns (DAMPs)‐induced inflammatory responses could be a novel target in clinical islet transplantation. We investigated the protective effects of NecroX‐7, a novel clinical‐grade necrosis inhibitor that specifically targets mitochondrial ROS, against primary islet graft failure. Islets from heterozygote human islet amyloid polypeptide transgenic (hIAPP+/−) mice and nonhuman primates (NHPs) were isolated or cultured with or without NecroX‐7 in serum‐deprived medium. Supplementation with NecroX‐7 during hIAPP+/− mouse islet isolation markedly increased islet viability and adenosine triphosphate content, and attenuated ROS, transcription of c‐Jun N‐terminal kinases, high mobility group box 1, interleukin‐1beta (IL‐1 β), IL‐6, and tumor necrosis factor‐alpha. Supplementation of NecroX‐7 during serum‐deprived culture also protected hIAPP+/− mouse and NHP islets against impaired viability, serum deprivation‐induced ROS, proinflammatory response, and accumulation of toxic IAPP oligomer. Supplementation with NecroX‐7 during isolation or serum‐deprived culture of hIAPP+/− mouse and NHP islets also improved posttransplant glycemia in the recipient streptozotocin‐induced diabetic hIAPP−/− mice and BALB/c‐nu/nu mice, respectively. In conclusion, pretransplant administration of NecroX‐7 during islet isolation and serum‐deprived culture suppressed mitochondrial ROS injury, generation of DAMPs‐induced proinflammatory responses, and accumulation of toxic IAPP oligomers ex vivo, and improved posttransplant glycemia in vivo. NecroX‐7, a novel clinical‐grade necrosis inhibitor, protects against primary islet graft failure in murine and nonhuman primate models through inhibition of mitochondrial reactive oxygen species and subsequent inflammatory responses induced by damage‐associated molecular patterns.