We developed a high accuracy thickness measurement system for glass substrates based on the optical design of the astigmatic method. The astigmatic optical system includes a laser diode, a cylindrical lens, a convex lens, and a quadrant detector. This method measures the astigmatic focusing error signal induced from the measured glass placed in the astigmatic optical system. The astigmatic focusing error signal is converted into the thickness of the glass substrate. The proposed glass thickness measurement system is verified by using a coordinate measuring machine (CMM). The accuracy of the proposed system is 0.2 microm, with a standard deviation of 0.7 microm within the thickness measuring range of 1.2 mm.