Secretory activity, particularly of antibacterial agents such as lysozyme and lactoferrin, is an important aspect of the mucosal defense mechanism, and the development of these agents may have a direct bearing on the susceptibility of the ear to infection. In this study, the secretory cells of the murine tubal epithelium were first observed at gestational day 17. Although tubal glands began to develop on gestational day 18, their secretory activity was first shown on postnatal day 3. The number of secretory cells of the tubal epithelium increased rapidly immediately after birth, while that of the tubal glands showed a gradual increase from postnatal day 3. The epithelial secretory cells in the tubotympanum matured at birth, but the tubal glands matured gradually after birth. Lysozyme was first recognized in the epithelial secretory cells on postnatal day 1, while lactoferrin was first detected in the tubal glands on postnatal day 3. Both lysozyme and lactoferrin were co-localized in the serous cells of the tubal glands. The secretion of lysozyme and lactoferrin seemed to reflect the maturation of the secretory cells in the murine tubotympanum.