The invention provides an automatic calibration method of a stepping motor. The automatic calibration method includes: acquiring an actual step count value when a probe reaches a central point of a constant speed area within a current period; on the basis of the actual step count value and preset step number, acquiring step number errors caused by step loss of the stepping motor in an acceleration area and overshoot in a deceleration area within the current period; adopting the step number errors for step number compensation in a next period. By the automatic calibration method, the problem of angle deviation of the stepping motor without a coded disc in the process of quick rotating can be solved, movement position of the probe in the constant speed area can be ensured to be consistent with expectation, the probe can be maintained to collect images in a preset area without angle deviation, and speed of the stepping motor in the constant speed area can be improved while movement accuracy of the probe in the constant speed area is guaranteed.