| This cross-sectional study was aimed to determine the seroprevalence of potential transfusion based transmissible infectious agents among apparently healthy blood donor community of Multan, Pakistan. For this purpose, blood specimens were collected from volunteer donors through standard method of venipuncture. The presence of antigens or antibodies against five major infectious agents was detected by the use of respective good quality standard immunochromatographic (ICT) assay kits. A total of 48020 blood samples were received at the respective tertiary care hospital of Multan during the study period. Among them 94.08% were found free from infection and therefore safe for transfusion. While 5.92% blood donations were infected with one or another infectious agent. Among infected blood donors, Hepatitis C virus (HCV) infection exhibited highest prevalence of 3.44% followed by Hepatitis B virus (HBV) (2.32%), Treponema pallidum (TP) (0.07%), malarial parasite (MP) (0.06%) and Human immunodeficiency virus (HIV) (0.01%). Therefore, we conclude a higher prevalence of HCV and HBV among blood donors of Multan, Pakistan during the year 2013. While, Syphilis, MP and HIV cases were comparatively less but prominent malarial cases were observed in particular months of vector-favourable temperature and humidity. The values of 95% confidence interval indicated 9.26%, 16.98% interval for the prevalence of HBV and HCV among the subject population. This study provided recent scenario of transfusion transmissible infections (TTIs) among general population, therefore, it could play an important role in limiting TTIs by drawing the attention of policy makers and health officials towards this serious health issue.