Influence of culture period on the allergenic composition of Pityrosporum orbiculare extracts

Background Previous characterization studies of Pityrosporum orbiculare allergens have led to contradictory results. In immunoblotting studies a range of IgE-bitiding proteins of 10–100kDa have been identified. In another study, however, the IgE-binding structures were claimed to be associated with high-molecular-weight polysaccharides or glycoproteins, presumably mannans or mannoproleins. Objective In the present study the reasons for these discrepancies were investigated. Methods P. orbiculare preparations were compared in IgE ELISA and IgE-inhibilion ELISA, as well as in immunoblotting with sera from atopic dermatitis patients. Results It was inferred that variations in the period of in vitro culture of P. orbiculare constituted the most important factor determining the different compositions of the resulting yeast cell extracts. After 2 days of culture a wide range of allergen if proteins was present but upon more prolonged culture (>4 days) most proteins of 10- 100kDa were lost. Accordingly, the protein concentration of the extracts gradually declined from 40% to 25% between days 4 and 15 of culture. On the other hand, the carbohydrate content remained fairly constant (approximately 30%). Using inhibition ELISA it was demonstrated that the high-molecular-weight glycoproleins or poly-saccharides presumably involved in most of the IgE-binding capacity in extracts from old cultures, were also present in comparable concentrations in all extracts tested, even after culture for only 2 or 4 days. Conclusion Preparations obtained from the exponential phase of yeast cultures (2–4 days old), should preferably be used in studies of the IgE response to P. orbicularc.