Tetanus is an important disease which created by tetanospasmin toxin of Clostridium tetani. In this study we surveyed six important factors including LF, KF, MLD, pH, OD and total protein assay of Harvard 52 (H52), G5 (HG5) and 49205 (H49205) strains of the bacterium to determine which of them were more suitable for use in vaccine production. The mentioned strains were seperatedly reactivated in thioglycolate medium, 2ml of this suspension was used to inoculate the Muller-Miller medium of each of sixteen 500 ml Erlenmeyer flasks (7,6 and 3 flasks for H52, HG5 and H49205 respectively), where the fermentation runs were performed. Over a period of seven days of experiment, several tests for evaluation of the six mentioned factors on samples of medium cultures were carried out. Results revealed that H52 strain had significantly lower values in LF and OD compared to strains HG5 & H49205 (P