Objective: Studies show that immobilization stress has a variety of effects on serum levels of liver enzymes. The main aim of this study was to determine the effects of immobilization stress on serum level of creatine kinase and alkaline phosphatase in male rats. Method: 50 male Wistar rats weighing 200±30 g were randomly divided into control, acutely or chronically immobilized animals of 5 in each group. Animals were immobilized for 2 h/day or 8 h/day for a period of 3 weeks or 1 week in chronically or acutely immobilized groups, respectively. Blood samples were collected using cardiac puncture method. Following serum collection, creatine kinase or alkaline phosphatase level was measured by spectrophotometery method. Data were statistically analyzed and compared between groups using ANOVA. Results: The results indicated that serum creatine kinase level was significantly increased in rats enduring acute or chronic immobilization compared with control animals (P< 0.001), however, there was not significant difference between serum alkaline phosphatase levels in immobilized animals compared with control rats. Conclusion: Immobilization stress may leave pathological effects in liver or other organs leading to enhanced serum creatine kinase level.