Hepatitis B virus (HBV) and C virus (HCV) are major public health concerns in Pakistan. We conducted a baseline analysis of first-time replacement blood donors at the blood bank of Jinnah Postgraduate Medical Centre in the year 2000 with the view to developing low-cost sentinel surveillance for these infections. Among 7325 such donors, 264 (3.6%) were positive for anti-HCV and 344 (4.7%) for HBsAg. HCV seroprevalence was significantly positively associated with age and lower education. Those speaking Sindhi or other minor languages had a significantly higher prevalence of HBV infection. Blood bank data could provide reliable information to monitor trends in prevalence of these infections.