Objective: To explore the potential effect of melatonin on the in-vitro maturation of mouse oocytes under heat shock condition.Methods: This study used a heat shock model of mouse oocyte maturation. The oocytes were randomly divided into three groups: control group, heat shock group and heat shock + melatonin group, in order to evaluate the effect of 1×10−9 mol/L melatonin on the quality of oocytes after heat shock.Results: In comparison with the control group, the maturation rate of mouse oocytes in heat shock group was significantly decreased [(33.00 ± 0.07)% vs. (85.00 ± 0.03)%, p < .01], with abnormal spindle assembly, and the early apoptosis rate was significantly increased [(59.7 ± 4.5)% vs. (22.0 ± 3.5)%, p < .01]. Compared with heat shock group, the maturation rate ofoocytes was significantly increased in heat shock + melatonin group [(70.00 ± 0.05)% vs. (33.00 ± 0.07)%, p < .01], meanwhile, the spindle abnormality rate and the early apoptosis rate were significantly decreased accordingly [(37.3 ± 6.1)% vs. (59.7 ± 4.5)%, p < .01]. The expression level of heat shock protein 70 was significantly up-regulated in heat shock + melatonin group in comparison with other two groups (p < .01).Conclusions: By regulating the over-expression of heat shock protein 70, melatonin can improve the declined maturation rate of oocytes and the increased rates of spindle assembly abnormality and early apoptosis caused by heat shock.