Cell wound healing assay is an important experimental technique for the detection of cell migration in vitro. At present, scratch on monolayer cells using a pipette tip is commonly used. However, it is difficult to guarantee the scratch with same width, and only the migration distance of a certain part is calculated. Therefore, the experimental method needs to be optimized. ATRA was used to treat hepa1-6 mouse hepatoma cells. Circular wound with diameter of 0.2 cm were formed by a circular gap forming device. The whole cell wound region could be captured under the microscope to observe cell migration. There are almost no crawling cells in the wound region. The migration capacity of hepa1-6 cells was evaluated by calculating the healing area. ATRA could significantly inhibit the migration of hepa1-6 cells. Compared with linear wound, the standard deviation of wound healing rate in the circular cell wound method is smaller. The circular cell wound method can ensure the dynamic observation of the same wound region, and calculate the healing area at the two-dimensional level with small error and high repetition rate. It is reliable and easy to operate, can be widely used in laboratory.