Background We have previously identified two major allergens of Pityrosporum orbiculare and characterized these as 37 kDa and 67 kDa proteins.
Objective In the present study we have investigated the presence and subcellular location of the 37 kDa and 67 kDa allergen components in various members of the genus Pityrosporum as well as in Candida albicans. Candida parapsilosis and Saccharomyces cerevisiae. Methods To detect both cell surface and intracellular expression of the allergens, flow cytometry and confocal laser scanning microscopy (CLSM) were used. The cells were stained with indirect immunofluorescent (IIP) or alkaline phosphatase antialkaline phosphatase (APAAP) methods using mouse monoclonal antibodies (MoAbs)
Results Ninety-five per cent of the P. orbiculare (P. ovale) cells cultured for 4 days showed cell surface-binding of the anti-37 kDa MoAb and 88% of the cells bound the anti- 67 kDa MoAb when analysed with IIF and flow cytometry. It was found that the members of the genus Pityrosporum (Malassezia). P. pachydermatis and M. sympodialis, expressed the 37 kDa and 67 kDa allergens to a similar extent as did P. orbiculare. Less than 5% of the cells of the genus Candida and S. cerevisiae showed positive staining with the MoAbs, The CLSM revealed that the 37 kDa and the 67 kDa components were located to the cell wall and could not be detected inside the acetone fixed and APAAP stained yeast cells of the genus Pityrosporum. When the yeast cells were cultured for more than 4 days the expression of both allergens decreased significantly.
Conclusion All three members of the genus Pityrosporum express the 37 kDa and 67 kDa major allergens on the cell surface, whereas these proteins could virtually not be detected in the Candida genus and S. cerevisiae.