[Objective] To study the application parameters of thermo-stabilizer in the manufacture of swine paratyphoid vaccine, and provide reference for the preparation of thermo-stable bacterial live vaccine. [Methods] The thermo-stabilizer was mixed with same amount of bacteria culture, Salmonella enterica serovar Choleraesuis strain C500. The mixture was lyophilized using freeze-drying curves 1, 2 and 3, and the matched freeze-drying curved was then determined by sampling the survival ratio post lyophilization. Based on the optimal freeze-drying curve, the effect of various factors on the vaccine quality post lyophilization was further investigated. The studied factors included the culture age (C500 culture fermented for 12, 15, 18 and 21 h), the culture concentration (3×1010, 5×1010 and 7×1010 CFU/mL), interaction time for thermo-stabilizer and C500 culture (0, 24 and 48 h), storage time of fresh prepared thermo-stabilizer (0, 10, 20, 30 and 40 d) and the sub-package volume (2, 3 and 4 mL). [Results] The lyophilized product by freezing-drying curve 1 had the properties of surface crystallization and bottom shrinkage. The lyophilized product by freezing-drying curve 3 had a small amount of crystallization and the lower concave on the surface. The lyophilized product by freeze-drying curve 2 had the best properties, highest survival ratio and best aging resistance. The C500 strain cultured at 37 °C for 18 h was optimal for lyophilization with a highest survival ratio of 78%?81%. The optimal culture concentration was 3×1010?5×1010 CFU/mL. The optimal incubation time for the C500 culture and thermo-stabilizer was 24 h at 2?8 °C, with the survival ratio of 85.3%?90.5%. After stored at 2?8 oC for 40 days, the thermo-stabilizer has the same effect with fresh prepared one. The survival ratio after lyophilization was almost the same for the sub-package of 3 mL or 4 mL compared with 2 mL, but the survival ratio was 3?7 percentage points higher than sub-packing with 2 mL after stored at 37 °C for 7 days. [Conclusion] The optimal parameters for the thermo-stabilizer in manufacture of swine paratyphoid live vaccine were the following: freeze-drying curve 2, culture time of 18 h, culture concentration of 3×1010?5×1010 CFU/mL, thermo-stabilizer used within 40 days if storage at 2?8 °C from fresh preparation, incubation time of 24 h for the C500 culture and thermo-stabilizer, sub-package volume of 3?4 mL.