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Background: Iron deficiency anemia (IDA) is the most common anemia followed by anemia of chronic disease (ACD). Reticulocyte indices have been shown to be helpful indicators for detecting IDA. We investigated whether RBC and reticulocyte indices can be used to differentiate ACD from IDA. Methods: A total of 85 women showing microcytic hypochromic anemia (38 IDA and 47 ACD cases) were enrolled. IDA was defined as those with ferritin level of <6 μg/dL and total iron binding capacity (TIBC) of >450 μg/dL. ACD was defined as ferritin level of ≥6 μg/dL, TIBC of ≤450 μg/dL, and presence of underlying diseases. We measured complete blood count, TIBC, iron, ferritin, and RBC and reticulocyte indices. The mean values of each item were compared between the two groups and sensitivity and specificity of each item in the differential diagnosis of ACD from IDA were determined by ROC curve analysis. Results: In ACD, most of the RBC and reticulocyte indices were significantly higher than in IDA: mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concentration (MCHC), cellular hemoglobin concentration mean (CHCM), cellular hemoglobin content (CH), red cell distribution width (RDW), reticulocyte hemoglobin content (CHr), and mature RBC cellular hemoglobin content (CHm). All these indices, except MCV showed significant correlations with ferritin and/or TIBC. CHr level of ≥24.6 pg could be used to differentiate ACD from IDA with 85.1%sensitivity and 81.6% specificity. Conclusions: The reticulocyte indices, especially CHr, are useful for the differential diagnosis of microcytic hypochromic anemias, ACD and IDA.