BACKGROUND: Iron deficiency is common after Roux-en-Y gastric bypass (RYGB) surgery, but there is no consensus on the optimal diet quality and quantity for restoring and preserving iron status. OBJECTIVES: The authors explored the impact of dietary and supplemental sources of iron and absorptive factors on iron status. SETTING: Academic, United States. METHODS: In a cross-sectional cohort of individuals who underwent RYGB, nutrient intakes from food and supplements were measured using 3-day food records. Blood biomarkers of iron status, including concentrations of ferritin, total iron binding capacity, serum transferrin receptor (sTfR), and the sTfR:ferritin ratio, were assessed by a reference laboratory; iron deficiency was defined as having at least 2 abnormal measures. Associations between iron status biomarkers and dietary predictors were determined using regression analysis. RESULTS: Of the 36 participants, 97% were female, the mean age was 45 years (95% confidence interval, 41-48 years), and body mass index was 32 (30-35) kg/m(2). Iron deficiency was found in 42% of participants. Dietary intake of heme iron, found in meats, was favorably associated with 3 iron status biomarkers (ferritin, β = .366; sTfR:ferritin ratio, β = -.459; and total iron binding capacity, β = -18.26; all P<.05), independent of obesity-induced inflammation. Intake of vitamin C from food contributed to iron status (ferritin, β = .010 and sTfR:ferritin ratio, β = -.011; P<.05). Use of supplementary non-heme iron, at doses recommended for prophylaxis (45 mg/d), was positively associated with serum ferritin (β = .964; P = .029). CONCLUSIONS: For patients who have undergone RYGB, consuming high, but realistic amounts of heme iron in meat, vitamin C from food, and adherence to recommended iron supplements can prevent iron deficiency.