Introduction: Chronic iron deficiency is a well-recognized complication of regular blood donation [1]. Cut-off value of hemoglobin of 12.5 g/dl is used for blood donation [2]. Since hemoglobin levels may be normal in the presence of reduced iron stores, individuals potentially at risk for developing iron deficiency anemia may be detected by serum ferritin estimation [1] Materials and methods: The prospective, cross-sectional study was carried out on blood donors who had donated twice or thrice previously in a year and were donating voluntarily for the third or fourth time in blood bank of BLDEU Shri B.M. Patil Medical College, Hospital and Research Centre, Bijapur from October 2013 to March 2015. Routine hematological parameters like Hb, hematocrit, MCV, MCH, MCHC, RDW of the sample were measured. Serum ferritin estimation was done using ELISA technique. Result: Inter-group comparison of serum ferritin was done. Among male donors the mean serum ferritin values showed a decreasing trend with increasing number of donations per year which was significant but the same was not observed with female donors. Hematological parameters such as hemoglobin, hematocrit, MCV, MCH, MCHC and RDW did not show significant fall with increasing number of donations. Conclusion: The present study underscores the utility of Serum Ferritin level estimation as an adjuvant to hemoglobin assessment in multiple time donors. While there is clear cut consensus among blood banks to consider hemoglobin as a method for blood donor anemia screening, serum ferritin estimation can be planned before bleeding a prospective non anemic multiple time donor to correctly assess the iron status.