Abstract: An object of the present invention is to provide a method for producing an ADM suitable as a scaffold for a cultured skin, that is, a separation and decellularization method that enables various types of extracellular matrixes such as a basement membrane to be preserved, allows the epidermal layer to be peeled off easily, and does not cause any damage to the dermal matrix. The present invention relates to a skin separation and decellularization method comprising a step of freeze thawing harvested skin and then separating the skin into epidermis and dermis by a treatment with hypertonic saline, and a step of washing the separated dermis.