Epidermal stem cells accelerate their repopulation rate during fractionated irradiation. To study the changes in keratinocyte differentiation associated with radiation-induced repopulation, we investigated the expression of a panel of 14 monoclonal or polyclonal antibodies against keratins, involucrin and others, as well as the expression of 15 lectins in the irradiated mouse leg skin. Tissue samples were collected after 1, 2, and 3 weeks of daily irradiation with 3 Gy per fraction. Abnormal morphological appearance of the irradiated epidermis suggested disturbed terminal differentiation. Keratin 16 (K16) was negative in normal epidermis but intense staining was observed in the irradiated epidermis. Involucrin was expressed in the outmost suprabasal layers only in the normal epidermis but extended to the lower layers in the irradiated epidermis. The lectin binding patterns for agglutinins from Soybean, Dolichos biflorus, and Helix pomatia showed differences between the normal and the irradiated epidermis. From these characteristic changes in staining patterns we concluded that accelerated repopulation of the epidermis during fractionated irradiation is associated with a deficiency in terminal squamous differentiation.