IMMUNOCHEMICAL AND SUBCELLULAR-LOCALIZATION OF THE 43 KDA GLYCOPROTEIN ANTIGEN OF PARACOCCIDIOIDES-BRASILIENSIS WITH MONOCLONAL-ANTIBODIES

Two monoclonal antibodies ST-7 (IgG1) and ST-8 (IgG2b), directed again st a 43 kDa glycoprotein (gp43) of Paracoccidioides brasiliensis were produced. It was possible to detect the gp43 by ELISA in amounts as lo w as 100 ng per well, and by Western blot about 300 ng were detected. Mild treatment of the gp43 with sodium metaperiodate did not alter its reactivity with ST-7 and ST-8, which suggests that these MAbs recogni ze peptide epitopes. Confirming the periodate oxidation data, the 38 k Da protein resulting from deglycosylation of the gp43 with trifluorome thanesulphonic acid (TMFS), was reactive with ST-7 and ST-8. Immunoele ctron microscopy showed that the gp43 is stored inside large dense cor e vesicles, which flowed into the plasma membrane and extruded from ce ll membrane into the cell wall. Finally the antigen was secreted into the extracellular space as dense membrane-free material. Secretion of the gp43 occurred at scattered sites interspersed along the cell surfa ce.