BACKGROUND Roux-en-Y gastric bypass (RYGB) may reduce the absorption of iron, but the extent to which this absorption is impeded is largely unknown. First, we determined the prevalence of iron deficiency following RYGB and explored the risk factors for its development. Second, we examined to what extent oral iron supplements are absorbed after RYGB. METHODS Monocentric retrospective study in 164 patients (123 females, 41 males; mean age 43 years) who underwent RYGB between January 2006 and November 2010 was done. Pre- and postoperative data on gender, age, BMI, serum levels of iron, ferritin, hemoglobin, vitamin B12, 25-hydroxy vitamin D, and use of proton pump inhibitors and H2 antagonists were collected. Generalized linear mixed models were used for the analysis of the data. In 23 patients who developed iron deficiency after surgery, an oral challenge test with 100 mg FeSO4 · 7H2O was performed. RESULTS Following RYGB, 52 (42.3 %) female patients and 9 male (22.0 %) patients developed iron deficiency (serum ferritin concentration ≤ 20 μg/L). The prevalence of iron deficiency was significantly higher in females than males (p = 0.0170). Young age (p = 0.0120), poor preoperative iron status (p = 0.0004), vitamin B12 deficiency (p = 0.0009), and increasing time after surgery (p < 0.0001) were also associated with iron deficiency. In the oral iron challenge test, only one patient out of 23 showed sufficient iron absorption. CONCLUSIONS Iron deficiency is extremely frequent after RYGB and is linked with different risk factors. Iron supplementation seems essential, but the effect of oral tablets may be limited as absorption of oral iron supplements is insufficient post-RYGB.