Cryptococcus neoformans is an encapsulated fungus that is a major cause of meningitis in patients with AIDS. In immunocompetent mice, administration of IgG1 mAb protects against cryptococcal infection, whereas administration of IgG3 is not protective and can accelerate the infection. In beige mice with impaired natural killer cell function, the effects of
The biosynthesis of the 0 antigen of Citrobacter 139 (Escherichia coli 3 Zurich 4,5,12:z2o) was shown to proceed through a series of lipid-linked intermediates, similar to those involved in 0-antigen synthesis in Salmonella. Galactose was the first sugar incorporated, followed by rhamnose and mannose. Abequose was incorporated from cytidine diphosphate (CDP)-abequose only when all three of the other nucleotide sugars (uridine diphosphate galactose, guanosine diphosphate mannose, and thymidine diphosphate rhamnose) were present. Rhamnosyl-galactosyl 1-phosphate and mannosyl-rhamnosyl-galactosyl 1-phosphate were identified as the products of mild alkaline hydrolysis of the lipid-linked intermediates. The incorporation of galactose, mannose, and rhamnose into 0-antigen repeating units has been reported for a guanosine diphosphate (GDP)mannose-deficient mutant of Salmonella typhimurium by Zeleznick et al. (12), for a thymidine diphosphate (TDP)-rhamnose-deficient mutant of S. typhimurium by Nikaido (5), and for wild-type S. anatum by Robbins (7). Weiner and co-workers (8, 9) and Wright et al. (11) have obtained evidence for a series of lipid-linked oligosaccharide intermediates in the formation of the 0-antigen repeating units in both S. typhimurium and S. newington. Anderson et al. (1, 2) have shown that similar lipid-linked intermediates are formed in the biosynthesis of cell wall glycopeptide in Staphylococcus aureus and Micrococcus lysodeikticus.
Allelic exclusion normally results in the expression of only one light and one heavy chain gene. However, some hybridomas have been reported to express two different heavy chain genes. Here we report that the IgG3 hybridoma 4H3.C8B expresses both kappa and lambda light chains. 4H3.C8B was originally recovered by screening for antibody binding to Cryptococcus neoformans polysaccharide antigen and characterized as gamma 3 lambda. The gamma 3 lambda but not the gamma 3 kappa binds to polysaccharide antigen. Some of the antibody molecules were heterodimers composed of both lambda and kappa. IgG1 and IgG2b isotype switch variants were identified and isolated by the technique of sib selection, using the ELISA spot assay. Like the parent IgG3 line, the switch variants continued to express both kappa and lambda light chains but only the lambda-containing antibodies bound to the antigen. Our experience suggests that hybridomas recovered by assays that are antigen dependent should also be tested for the expression of other isotypes and light chains in a non-antigen-binding immunoassay.
Passively administered mAbs to Cryptococcus neoformans capsular polysaccharide can alter the course of infection in mouse models. In preliminary studies of passive Ab efficacy, most IgM, IgA, and IgG1 mAbs were protective, but the few IgG3 mAbs tested did not confer significant protection. Because IgG3 is effective in pneumococcal infections, this phenomenon was examined more rigorously by generating an IgG1 switch variant from the non-protective IgG3 mAb 3E5 and comparing its protective efficacy in a murine model of i.v. infection by using strains of both the A and D serotypes. The 3E5 IgG3 mAb did not prolong survival or reduce organ fungal burden. Rather, the IgG3 decreased survival relative to controls. In contrast, the IgG1 switch variant of 3E5 significantly prolonged survival, reduced organ colony-forming units, and reduced serum polysaccharide Ag level in infected mice. The results establish that isotype is important for Ab efficacy against C. neoformans.
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