Background: Blood transfusion is lifesaving but it is also an important mode of transmission of infections to recipients. With every unit of blood there is a 1% chance of transfusion associated problems including transfusion transmitted infections. Aim: The aim of the present study was to find out prevalence of transfusion transmitted infections (TTI) in voluntary and replacement donors in our hospital transfusion service set up. This study also aids in evaluating the safety of the collected donations. Material and Methods: This five years record based descriptive retrospective study was conducted from January 2009 to December 2013 at the blood bank, P.D.U. Government Hospital and Medical College, Rajkot, Gujarat, India. Donors and cases of TTI were identified from computer records of blood bank of hospital and monthly report sent to GSACS (Gujarat State AIDS Control Society), Ahmadabad. The data were analysed in computer software, Microsoft excel 2010. Result: A total of 48162 donors were tested, out of which 10478 (21.76%) were replacement donors and 37684 (78.24%) were voluntary donors. The seroprevalence of HIV, HBV, HCV and syphilis was 0.21%, 0.95%, 0.24% and 0.19% respectively in total donors. Replacement donors had a high incidence of TTI was 274(2.62%) as compared to voluntary donors were 492(1.31%). Conclusion: The seroprevalence was more in replacement donors as compared to voluntary donors. It concludes that motivating voluntary blood donors are the most effective way of ensuring adequate supplies of safe blood on a continuing basis.