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Introduction:The current study evaluated the value of red blood cell (RBC) indices and the corresponding cut- off points for β-thalassemia control programs in Iran. Materials and Methods:1,150 individuals (575 couples) with low RBC indices and normal hemoglobin A2 who had been referred to the Genetic Centre of Isfahan, were tested during pre-marital screening analyses, in the 2 year period, 2006-2008. β-thalassemia mutations were evaluated. Results: β-thalassemia was identified in 67.8% of the cases with both mean corpuscular volume (MCV) less than 78fl and mean corpuscular hemoglobin (MCH) less than 26 pg. However, 4.1% of the individuals with  78≤ MCV≤ 80 tested positive for thalassemia. MCV showed a higher diagnostic reliability than MCH. However, the accompaniment of MCH<26 with MCV<78 increased the risk of thalassemia 35 times. A significant association was found between the prevalence of minor β-thalassemia and educational levels, race and familial relationships. Conclusion: According to the results of this study, it seems that MCV=80fl can be used as a proper cut- off point for the screening of minor β-thalassemia. Although in the present study MCV was found to have a higher diagnostic reliability than MCH, MCH <26 along with MCV are very helpful indices for the counselor physician to estimate the risk of minor β-thalassemia more accurately.