© 2018 European Association for the Study of the Liver Background & Aims: Lactation lowers blood glucose and triglycerides, and increases insulin sensitivity. We hypothesized that a longer duration of lactation would be associated with lower prevalence of non-alcoholic fatty liver disease (NAFLD), which is the leading cause of chronic liver disease in the United States. Methods: Participants from the Coronary Artery Risk Development in Young Adults cohort study who delivered ≥ 1 child post-baseline (Y0: 1985–1986), and underwent CT quantification of hepatic steatosis 25 years following cohort entry (Y25: 2010–2011) were included (n = 844). The duration of lactation was summed for all post-baseline births, and NAFLD at Y25 was assessed by central review of CT images and defined by liver attenuation ≤ 40 Hounsfield Units after exclusion of other causes of hepatic steatosis. Unadjusted and multivariable logistic regression analyses were performed using an a priori set of confounding variables; age, race, education, and baseline body mass index. Results: Of 844 women who delivered after baseline (48% black, 52% white, mean age 49 years at Y25 exam), 32% reported lactation duration of 0 to 1 month, 25% reported >1 to 6 months, 43% reported more than 6 months, while 54 (6%) had NAFLD. Longer lactation duration was inversely associated with NAFLD in unadjusted logistic regression. For women who reported >6 months lactation compared to those reporting 0–1 month, the odds ratio for NAFLD was 0.48 (95% CI 0.25–0.94; p = 0.03) and the association remained after adjustment for confounders (adjusted odds ratio 0.46; 95% CI 0.22–0.97; p = 0.04). Conclusions: A longer duration of lactation, particularly greater than 6 months, is associated with lower odds of NAFLD in mid-life and may represent a modifiable risk factor for NAFLD. Lay summary: A longer duration of breastfeeding has been associated with multiple potential health benefits for the mother including reduction in heart disease, diabetes and certain cancers. In this study we found that breastfeeding for longer than 6 months was associated with a lower risk of non-alcoholic fatty liver disease in mid-life.