[Abstract] Objective The heart rate and behavior reactions of Labrador retrievers in the distance test (DT) was studied in this study, which may provide effective behavioral and physiological basis for early identification of potential guide dogs. Methods The 60 Labrador retrievers (8-15 months old; 22 females, 38 males) were provided by Chinese guide dog training centre of Dalian. 30 of the 60 dogs were guide dogs and the other 30 were eliminated dogs which were evaluated by exam in the training center. The “Interest”, “Exploration” and “Play Invitation” behavioral categories in the DT of the dogs were tested and the heart rate reactions were simultaneously recorded. Results The mean heart rate reactions of the guide dogs were significantly lower than those of the eliminated dogs in the DT (P<0.05). The scores of “Interest”, “Exploration” and “Play Invitation” showed no significant difference between guide dogs and eliminated dogs (P>0.05). Conclusion In the DT, the changes of the heart rate reaction in guide dogs was significantly lower than eliminated dogs. This study indicated that guide dogs have a more stable psychological quality and better control of emotions compared with eliminated dogs in the DT. This study can provide a theoretical basis for the combination of DT and heart rate to predict the success rate of the guide dog training.