Objective To observe the contribution of acute normovolemic hemodilution (ANH) in experiment of cryopreserved dog limb replantation. Methods Sixteen healthy Beagle dogs (male:female=1:1) were divided into two groups. Dogs in the experiment group (Group B) received ANH in the limb replantation, and dogs in the control group (Group A) received the same amount of lactate Ringer's solution intravenously during the surgical operation. We recorded and compared the hemodynamic indexes, HB, HCT, the resuscitation time, the first rising head time, the first standing time and the first eating and drinking time between the two groups. Results (1) During the operation, both PaO₂ and PCO₂ in the two groups were normal, as well as the breathing rate. The heart rate in the group B was lower than that in the group A. Before blood transfusion, there was no statistically significant difference in HB and HCT between the two groups, but after transfusion they were significantly higher in the group B than in the group A. (2) The resuscitation time, the first rising head time, the first standing time and the first eating and drinking time of the group B were all better than those in the group A. Conclusions In cryopreserved dog limb replantation experiments, acute normovolemic hemodilution is helpful to improve the general condition and facilitate the recovery of animals after limb replantation.