Objective To explore the feasibility of establishing a stable mouse model of thin endometrium by intrauterine injection of 95% ethanol, characterized by immunohistochemistry and fertility assessment, and provide an ideal animal model for the study of pathological features and repair mechanisms of thin endometrium. Methods Seventy-eight female unmated C57BL/6J mice with sexual maturity and regular estrus were used in this study. Thirty mice were randomly divided into three groups: the blank group, control group, and experimental group. A mouse model of thin endometrium was established by intrauterine infusion of 95% ethanol injured endometrium. The experimental mice were euthanized by cervical dislocation in the third estrus period after modeling, and uterine samples were collected. The regeneration of endometrial cells was evaluated by immunohistochemistry for cytokeratin, vimentin, vascular endothelial growth factor(VEGF) and estrogen receptor α(ERα). Another group of 48 mice were randomly divided into the blank group, control group, unilateral injury group, and the bilateral injury group. All surviving mice were simultaneously caged after modeling to analyze the effect of thin endometrium on the fertility of mice and to evaluate whether the mouse model of thin endometrium was successfully established. Results The results of the immunohistochemistry showed that the expressions of cytokeratin, vimentin, VEGF and ERα in the endometrial cells of the experimental group were significantly lower than those in the blank and control groups. Fertility assessment showed that mice in the blank and control groups were normally conceived after mating and that no mice in the bilateral injury group were conceived. In addition, no mice were conceived in the injured side of the uterus in the unilateral injury group, whereas mice in the uninjured side of the uterus were conceived normally. The mean pregnancy rate of the injured side of the uterus in the experimental group was significantly lower than that on the uninjured side ( P < 0. 05), indicating that intrauterine injection of 95% ethanol damaged the endometrium, resultsing in reduced fertility. Conclusions We have successfully established a mouse model of thin endometrium, which can be used to study the repair mechanisms of the thin endometrium.