A robot fork calibration method and system is provided. At least three non-linear arranged lower sensors are provided on a bottom surface of the fork to detect distances to a fixed detection point. The fixed detection point and a horizontal plane of the detection point define a reference coordinate system. Spatial coordinates of the lower sensors in the reference coordinate system are calculated and a plane equation as well as a tilted angle of the fork are obtained according to the spatial coordinates. Therefore, the height of the fork can be calibrated according to the Z axis coordinates of the lower sensors and the levelness of the fork can be calibrated according to the tilted angle.