IDENTIFICATION OF ALLERGEN COMPONENTS OF THE OPPORTUNISTIC YEAST PITYROSPORUM ORBICULARE BY MONOCLONAL-ANTIBODIES

The yeast Pityrosporum orbiculare (P. orbiculare) is a member of the n ormal human cutaneous flora, but it is also associated with several cl inical manifestations of the skin. We have previously observed IgE-bin ding components in P. orbiculare extracts, using sera from patients wi th atopic dermatitis. In the present study, we raised several monoclon al antibodies (MoAbs) against P. orbiculare to characterize some of it s antigens, and used Candida albicans (C. albicans) as a control. We o btained several IgG1 MoAbs which specifically recognized P. orbiculare in ELISA. Two of these were selected for immunoblotting studies on P. orbiculare, and two patterns of reactivity emerged. Firstly, one MoAb showed a distinct band at a molecular mass of 67 kDa. In the second p attern, a sharp band at about 37 kDa appeared. In contrast, the IgM an tibodies raised reacted with a 14-kDa component; but they reacted with C. albicans in addition to P. orbiculare. The IgG1 antibodies seemed to react with proteins, as their ability to react in ELISA with extrac t pretreated with protease was greatly reduced. In contrast, IgM MoAbs were much less affected, suggesting that they recognized nonprotein c omponents. To determine whether these MoAbs-binding components were al so recognized by human IgE, we adopted a radioimmunoassay (RIA) using the MoAbs as catcher antibodies. Both the 67-kDa and the 37-kDa compon ents were IgE-binding proteins. P. orbiculare RAST positive sera were scored as positive in the RIA, whereas the control serum was not.