It is necessary to establish a highly efficient method of marking, which providing technical guarantee for a more accurate assessment of stock-enhancement effect of the cuttlefish (Sepiella japonica). The cuttlefish were chosen as the experimental material in this study. The larval cuttlefish were immersed in alizarin complexone (ALC) to dye the cuttlebone. The experiment was divided into 5 groups with concentration gradient and 2 time periods, and doral mantle length, body weight and survival rate of cuttlefish were measured and observed. The trial group and control group (50 ind/group) were reared in 2.5 m diameter circular concrete ponds, respectively. The cuttlefish were sampled randomly in order to examine the dye-retention of the tagged cuttlebone after being tagged 5, 12, 19, 26 d. Moreover, the tagged cuttlefish were examined at high temperature treatment for 5 min. The results showed that the optimised dye concentration and immerged time were 90 mg/L and 24 h, respectively. The doral mantle length of cuttlefish had a significant positive correlation with body weight (P<0.01), and there was no significant difference in growth between trial group and control group (P>0.05). The tagged oval pink ring was still clear in the cuttlebone even after being tagged 26 d. The range and color of tagging parts were not changed before and after high temperature treatment. Additionally, the proposed method has effect on both internal signs and external signs, and the survival rate and dye-retention rate of the tagged cuttlefish are 100%. It is concluded that this method can be successfully applied for marking cuttlefish, and offer a number of benefits, including simple, efficient, high sample throughput, and a wide range of application prospect.