Immunoelectronmicroscopic characterization of monoclonal antibodies (MAbs) against Cryptococcus neoformans.
Three monoclonal antibodies (MAbs) (BA4, BD1, CD6) reacted with Cryptococcus neoformans capsular glucuronoxylomannan (GXM) polysaccharide showing distinctive patterns against four serotypes as revealed by enzyme immunoassay (EIA), dot EIA, and immunofluorescence. Immunoelectron microscopy (IEM) was used to characterize binding sites for the MAbs on the C. neoformans capsule. All three MAbs bound to the capsule of serotype A strains 9104 and 9759. Differences in the intensity of binding to the two serotype A strains could not be explained by capsule diameter. The MAb BA-4 IgM bound well to 9759 (large capsule) and poorly to 9104 (small capsule), whereas MAb BD-1 (IgG-1) bound well to strain 9104 and poorly to strain 9759. Spurr's embedment inactivated the BA-4-binding epitopes in the C. neoformans 9759 capsule, but did not inactivate the ones that bound to BD-1. The epitopes recognized by BA-4 were different than the BD-1-binding determinants. The MAb CD-6 bound to a cytoplasmic precursor of capsular GXM. CD-6 (IgG) stained the capsule, cell wall, and cytoplasm of both C. neoformans tester strains. Competitive binding experiments were conducted. Single immunogold labelling showed that BD-1 inhibited the binding of BA-4, but not vice versa. The interaction between CD-6 and BA-4 resulted in a reciprocal inhibition. Double-labelling experiments showed reciprocal inhibition between BA-4 and each of the IgG MAbs. These MAbs are directed against capsular polysaccharide or its intracellular precursor. None of the MAbs stained C. neoformans cap 67, an acapsular mutant that does not contain GXM.
Todaro-Luck
F
F
Institute of Microbiology, University of Messina, Italy.
White
E H
EH
Reiss
E
E
Cherniak
R
R
D000908:Antibodies, Fungal / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D000946:Antigens, Fungal / Q000276:immunology*; D001667:Binding, Competitive; D003454:Cryptococcus / Q000276:immunology*; D003455:Cryptococcus neoformans / Q000276:immunology* / Q000648:ultrastructure; D000939:Epitopes / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D007075:Immunoglobulin M / Q000276:immunology; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D011134:Polysaccharides; D011135:Polysaccharides, Bacterial / Q000276:immunology*; D013048:Specimen Handling
D000908:Antibodies, Fungal; D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal; D000939:Epitopes; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011134:Polysaccharides; D011135:Polysaccharides, Bacterial; C027478:glucuronoxylomannan