Objective To explore the effects of gender and stain on formalin test in mice. Methods Analyzing the licking/biting time of male and female Kunming,BALB/c and C57BL/6 mice in formalin test. Results （1）In the earlyphase of formalin test, there was no difference among all animal groups. In the late--phase, 20± 25 min,the licking/biting time of male Kunming mice （23.25± 17.27）s was significantly less than that of C57BL/6 （63.33± 18. 20）s, and the licking/biting time of female Kunruing mice （64.63± 52.71）s was significantly longer than that of BALB/c （17.00± 22.34）s or CATBL/6 （32. 17±27, 42）s at 20-25 min. The licking/biting time of female C57BL/6 mice at 50-55 min （0. 83± 1.33）s was significantly less than that of Kunming mice （47.50±51.03）s. （2）The licking/biting time of male Kunming mice （23. 25±17.27）s was significantly less than that of female mice （64.63±52. 71）s. The licking/biting time of male C57BL/6 mice （63.33 ±18.20）s was significantly more than that of fe- male mice （22. 83±118. 41）s. Conclusions Gender and strain could significantly affect the licking/biting time of mice in formalin test. To increase the reliability of the test,male mice should be chosen.