Preparation of species-specific murine monoclonal antibodies against the yeast phase of Paracoccidioides brasiliensis

Abstract

A panel of four murine monoclonal antibodies showing species specificity for the yeast phase of the pathogenic dimorphic fungus Paracoccidioides brasiliensis was produced by using a modification of the standard monoclonal antibody technology. This involved the use of the immunosuppressive drug cyclophosphamide to suppress the immune response of test animals to fungi showing cross-reactivity, i.e., to Histoplasma capsulatum. One monoclonal antibody, P4, which had a high titer by enzyme-linked immunosorbent assay, was shown to recognize a linear antigenic epitope of P. brasiliensis at a molecular size of 70,000 to 75,000 daltons by Western blot (immunoblot) analysis. The potential use of these monoclonal antibodies, which are the first species-specific probes to P. brasiliensis that have been produced, in the field of serodiagnosis is discussed.

Development of a novel antigen detection test for histoplasmosis

“…Fungal isolates and antigen preparations. The following pathogenic fungi were cultured and harvested as previously described ( (4,11).…”

“…MAb production. The modification by Hamilton et al (4,11) of the Matthew and Sandrock protocol (14) was used. Briefly, 10 BALB/c mice were inoculated intraperitoneally (i.p.)…”

“…8 days later. After a further 3 days, mice were bled and sera were used in an enzyme-linked immunosorbent assay (ELISA) test (4,11) to determine the individual with the highest differential response to H. capsulatum antigens. This mouse received a further intravenous dose of 50 g of the H. capsulatum antigen mixture in sterile phosphate-buffered saline (PBS) and was used for the fusion protocol 3 days later.…”

“…This mouse received a further intravenous dose of 50 g of the H. capsulatum antigen mixture in sterile phosphate-buffered saline (PBS) and was used for the fusion protocol 3 days later. Spleen cells from the donor mouse were fused with Sp2/0 murine myeloma cells, and hybridomas were produced as previously described (4).…”

Paracoccidioidomycosis: an update.

“…Puccia and Travassos produced antibodies against the 43-kDa glycoprotein to foster their experimental studies on its excretion and proteolysis (236). Figueroa et al prepared monoclonal antibodies to define species-specific epitopes, and they discussed the potential use of monoclonal antibodies in serodiagnosis and the development of an ELISA for the detection of circulating antigen (107).…”