Cultured epidermis (JACE®) (Japan Tissue Engineering Co., Ltd.)

3T3-J2 cells (2×10⁴ cells/cm²), treated with mitomycin c for proliferation suppression, are used as a feeder layer for the epithelial cells (1×10⁵ cells/cm²) isolated from patient skin. As a culture medium, a 3:1 mixture of Dulbecco’s modified Eagles medium (DMEM) and HAM’s F-12 are supplemented with fetal bovine serum (5% FBS), cholera toxin (10 ng/ml), hydrocortisone (0.5 μg/ml), insulin (5 μg/ml), transferrin (5 μg/ml), triiodothronin (2×10⁻⁹ M ) and epithelial growth factor (10 ng/ml, EGF), along with antibiotic antimycotic reagent (1%). The cells are kept in an atmosphere of 10% CO₂ at 37 °C.