Internal defects in the glass, like-as micro-voids, micro-cracks, or inclusions, easily cause the failure when the glass is exposed to the shock or the thermal variation. In order to produce the highly reliable glass product, the precision inspection of the defect in the glass is required. For this purpose, this paper proposes a machine vision technique based on the auto-focusing method, which searches the defect and measures the location under the fact that the edge image of defect must be the most clear when the focal plane of CCD camera is coincided with the defect. As for the search index, the gradient indicator is presented. The basic principles are verified through the simulations for the computer-generated defect images, where the affects of defect shape, gray level of background, and the brightness of the defect image are also analyzed. Finally, experimental results for actual glass specimens are shown to confirm the applicability of this method to the actual field.