A measurement method of an overlay mark is provided. An overlay mark on a wafer is measured with a plurality of different wavelength regions of an optical measurement tool, so as to obtain a plurality of overlay values corresponding to the wavelength regions. The overlay mark on the wafer is measured with an electrical measurement tool to obtain a reference overlay value. The wavelength region that corresponds to the overlay value closest to the reference overlay value is determined as a correct wavelength region for the overlay mark.