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To date, limited information is available describing the incidence and impact of de novo donor-specific anti–human leukocyte antigen (HLA) antibodies (dnDSA) in the primary renal transplant patient. This report details the dnDSA incidence and actual 3-year post-dnDSA graft outcomes.The study includes 189 consecutive nonsensitized, non-HLA-identical patients who received a primary kidney transplant between March 1999 and March 2006. Protocol testing for DSA via LABScreen single antigen beads (One Lambda) was done before transplantation and at 1, 3, 6, 9, and 12 months after transplantation then annually and when clinically indicated.Of 189 patients, 47 (25%) developed dnDSA within 10 years. The 5-year posttransplantation cumulative incidence was 20%, with the largest proportion of patients developing dnDSA in the first posttransplantation year (11%). Young patients (18–35 years old at transplantation), deceased-donor transplant recipients, pretransplantation HLA (non-DSA)–positive patients, and patients with a DQ mismatch were the most likely to develop dnDSA. From DSA appearance, 9% of patients lost their graft at 1 year. Actual 3-year death-censored post-dnDSA graft loss was 24%.We conclude that 11% of the patients without detectable DSA at transplantation will have detectable DSA at 1 year, and over the next 4 years, the incidence of dnDSA will increase to 20%. After dnDSA development, 24% of the patients will fail within 3 years. Given these findings, future trials are warranted to determine if treatment of dnDSA-positive patients can prevent allograft failure.