Introduction: Red blood cells (RBCs) extended parameters or erythrocyte subsets are now reported by the new Sysmex XE 5000 analyzer. This study was aimed at establishing a characteristic analytical feature, including the new erythrocyte and reticulocyte parameters, in case of thalassemia trait and iron deficiency (IDA). Methods: Ninety healthy individuals, 136 b-thalassemia carriers, 121 mild IDA, and 126 severe IDA patients were analyzed. Results: The values obtained for the RBC extended parameters were significantly different (Po0.0001) in the groups; the only exception was %Hypo-He in the case of mild IDA and thalassemia (P 5 0.6226). %Hypo-He was considerably greater in severe IDA (23.4%) than in mild cases (12.4%), Po0.0001. %MicroR was more increased in thalassemia (38.6 %) than in the mild IDA (16.5%, Po0.001) and in severe IDA (21.6%, Po0.001). Immature reticulocyte fraction (IRF) mean values in the groups were statistically different; the thalassemia group had an intermediate value (8.7%) between healthy (4.4%) and IDA (16.7 and 12.9%). Conclusions: Erythrocytosis and severe microcytosis, together with a high percentage of microcytes and a moderate increase in IRF, is the profile of b-thalassemia carriers, whereas anisocytosis and the hypochromic subset correlates with the severity of the anemia in iron-deficient patients.