Iron deficiency anemia (IDA) is the most frequent type of anemia. The use of biochemical markers is a challenging way of diagnosis in case of inflammation and functional iron deficiency. The role of bone marrow aspirate iron staining is criticized because it is an invasive method and has no accurate standardization. This study was planned to find the value of bone marrow iron staining in diagnosis of iron deficiency. Four hundred and seven cases who had bone marrow aspirate iron staining and simultaneous serum iron tests have been reviewed retrospectively and 47 such cases were evaluated prospectively. Bone marrow iron was negative in 125 (%30.7) cases in retrospective cohort. There was significant differences in serum iron, ferritin, transferrin saturation between bone marrow iron negative and positive cohorts (p<0.001). Low serum transferrin saturation (<15%) and ferritin (<15 ng/ml) levels are consistent with iron deficiency. On this standard, in both retrospective and prospective cohorts, rates for specificiy (85.5% and 90.4%, respectively) and negative predictivity (97.5% and 100%, respectively) of bone marrow iron staining were relatively high compared with the sensitivity and positive predictivity of the same test. The results of this study show that the evaluation of bone marrow iron staining is a more reliable test for the elimination of IDA rather than its diagnosis.