Objective To observe the changes in estrous cycle and vaginal smears in ovarectomized NOD/SCID mice.Methods To continuously observe the estrous cycle time by vaginal smears of NOD/SCID mice in consecutive nine days, twice daily. After ovariectomy, the changes of estrous cycle were observed by vaginal smears for 7 days.Results The estrous cycle in NOD/SCID mice was 4-6 days. Regular estrous mice accounted for 80%. There was no significant correlation between vaginal opening and estrous cycle status. After ovariectomy, the vaginal smears showed characteristics of metestrus or diestrus. Conclusions Vaginal smear cytology can be used to determine the estrous cycle and characteristics of NOD/SCID female mice.The ovariectomized operation of NOD/SCID female mice is effective.