To find out adequate inactivator in substitution for the high concentration of formalin, different organic and inorganic chemicals, heat and chemicals combined with heat were tested for the inactivation of a fish-pathogenic bacterium Edwardsiella tarda strain. The survivality, sustainability of the cell surface antigenicity, bacterial protein and the total cell antigens of E. tarda were determined after treated with those inactivators. A major antigen at 37 kDa was detected by Sodium Dodecyl Sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blot analysis. Formalin (0.1%) with heat (70°Cx10 min) and 0.9% citric acid killed E. tarda resulted moderate killing activity with enough antigen-sustainability. This study suggests that formalin (0.1%) combined with heat (70°Cx10 min) and citric acid (0.9%) killed E. tarda as a new vaccine candidate.