To provide a method for manufacturing an SOI substrate having a single crystal semiconductor layer having a small and uniform thickness over an insulating film. Further, time of adding hydrogen ions is reduced and time of manufacture per SOI substrate is reduced. A bond layer is formed over a surface of a first semiconductor wafer and a separation layer is formed below the bond layer by irradiating the first semiconductor wafer with H3+ ions by an ion doping apparatus. H3+ ions accelerated by high voltage are separated to be three H+ ions at a semiconductor wafer surface, and the H+ ions cannot enter deeply. Therefore, H+ ions are added into a shallower region in the semiconductor wafer at a higher concentration than the case of using a conventional ion implantation method.