Background: Candida albicans is the causative agent of oral and vaginal candidiasis. Innate host defenses against C. albicans are important against each infection. Among these are oral and vaginal epithelial cells that have anti-Candida activity. The mechanism of action includes a requirement for cell contact with no role for soluble factors, and a putative role for carbohydrates based on the sensitivity of the activity to periodic acid. Methods: Periodic acid treatment of epithelial cells as well as the property of partial resistance of antifungal activity to fixation was used to further dissect the mechanism of action. Results: The results herein effectively now challenge a role for carbohydrates alone. Firstly, the putative carbohydrate(s) released into supernatants of periodic acid-treated epithelial cells could not compete with fresh epithelial cells for activity, and equivalent abrogation of activity was observed by periodic acid-treated cells irrespective of the amount of carbohydrate released. Instead, the similar abrogation of activity following treatment with other acids or when cocultured under acidic conditions suggests that the activity is acid-labile. Finally, while activity requires intact epithelial cells, it does not require live cells; activity was minimally affected by fixing epithelial cells prior to coculture where the majority of cells remained impermeable to Trypan blue but were defined as non-viable by positive nuclear staining with propidium iodide. Conclusion: These results suggest that antifungal activity is dependent on contact by intact, but not necessarily live, epithelial cells through an acid-labile mechanism. © Blackwell Munksgaard, 2005.