This study aimed to determine whether frozen blastocyst transfer can lower the incidence rate of ectopic pregnancy. A total of 1577 in vitro fertilization/ intra-cytoplasmic cycles were analyzed, including 757 fresh D3-ET cycles, 429 frozen D3-ET cycles, and 391 frozen D5-ET cycles. The differences of ages, the proportion of IVF cycles, the proportion of primary infertility, and the proportion of tubal factor between three groups were not significant. The clinical pregnancy rate in Frozen D5-ET Group was 62.15%, which was significantly higher than other two groups (46.10%; 38.93%; P < 0.05). The miscarriage rates of three groups were similar. The ectopic pregnancy rate in Frozen D5-ET Group was 0.41%, which was significantly lower than that in Frozen D3-ET Group (2.99%, P < 0.05) and slightly lower than that in Fresh D3-ET Group (2.29%, P = 0.089). Results from this study show that the transfer of frozen D5 blastocysts can significantly decrease the incidence rate of ectopic pregnancy.