We have previously reported that a lipophilic yeast, Pityrosporum ovale (P. ovale) produced a high frequency of positive skin prick tests and in vitro histamine-release (HR) tests in patients suffering from atopic dermatitis (AD) of the face, scalp, and neck. In the present study, our aim was to confirm the involvement of P. ovale-specific IgE and to produce a standardized extract for diagnostic tests; 7/20 sera from patients with a positive HR test were positive in RAST. Several IgE-binding proteins could be detected with sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by immunoblotting. Comparison of different extraction methods demonstrated that allergens were not released from P. ovale until after mechanical destruction of the yeast cells. Extraction of cultured P. ovale, obtained from the skin of various individuals suffering from AD of the face, scalp, and neck, resulted in very heterogenous patterns of constituents in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting analysis. Nevertheless, all extracts were recognized by patients' IgE. P. ovale may express varying protein patterns, depending on either source or stage of differentiation. Thus, at present, skin and HR tests may be superior to other available assays for diagnosing type I reactions to P. ovale in AD of the face, neck, and scalp.