Abstract: Low-temperature acclimation is critical for live fish transportation to reduce injuries and mortality by inducing the fish into semi-hibernate state. Thus, we investigated the suitable holding temperature for largemouth bass (Micropterus salmoides) transportation, focusing on the effect of water temperature on its muscle tissue, serum biochemical indicators, antioxidant enzyme activities and respiratory function. The results show that the respiratory rate of M. salmoides gradually decreased as temperature descended. Significant differences were observed in the muscle tissue, pH, serum biochemical indicators, and antioxidant capacity between temperatures (15 ℃ and 20 ℃) and lower temperatures (9 ℃ and 12 ℃) (p<0.05) during holding of 48-hour. M. salmoides exposed semi-hibernate state at 12 ℃, which led to a significant lower quality of total ammonia nitrogen (TAN) accumulation in the water (p<0.05) and a slower TAN increase rate. In the 12 ℃ group, the serum glucose (GLU) and lactate dehydrogenase (LDH) concentrations decreased significantly, while the total antioxidant capacity increased, with no significant changes on other indicators (p>0.05), and the blood cells in the gill tissue arranged orderly, with gill filaments showing no obvious deformation. The findings indicate that 12 ℃ holding not only effectively maintains water quality, but also effectively preserves the physiological functions and antioxidant capacity of fish, thereby reducing damage caused by oxidative stress reactions. Therefore, it is recommended to control the temperature of the water body at around 12 ° C during the actual holding and circulation process.