This study aimed to examine the influence of sub-microscopic media animation on students' critical thinking skills in terms of gender. The study used pre-test and post-test control group design. The sample consisted of experimental group and control group, each of which consisted of 27 students (12 males and 15 females). The sampling technique used was purposive sampling. Data on critical thinking skills was taken using a description test on buffer concepts. Further data were analyzed using t-test at 95% significant level. The results showed that sub-microscopic media animation influence students' critical thinking skills, both male and female. In the experimental group, students' critical thinking skills are higher than male students, which means that sub-microscopic animation media influence female students' critical thinking skills better than male students.