La2, David F Smith3, Jamie Heimburg-Molinaro3, Richard D Cummings1,3, as well as a Kwame Nyame1,two Department of All-natural Sciences, University of Maryland Eastern Shore, Princess Anne, MD, USA and 3Department of Biochemistry, Emory University College of Medicine, 1510 Clifton Rd #4001, Atlanta, GA 30322, USAepitope. Our results show that F8A1.1 recognizes terminal Lex epitopes and may be used for identification, immunolocalization, immunoprecipitation and purification of Lexcontaining glycoconjugates from schistosomes and mammalian cells. Keyword phrases: glycans / helminth / Lewis x antigen / monoclonal antibody / Schistosoma mansoniReceived on September 19, 2012; revised on March 13, 2013; accepted on March 26,The parasitic blood fluke Schistosoma mansoni synthesizes immunogenic glycans containing the human Lewis x antigen (Lex; Galactose-1-4(Fuc1-3)N-acetylglucosamine–R, also named CD15), but the biological part(s) of this antigen inside the parasites and in humans is poorly understood.Zearalanone web To create IgG-based monoclonal antibodies (mAbs) precise for Lex, we harvested splenocytes from S. mansoni-infected Swiss Webster mice at Week 10 postinfection, when peak IgG responses to glycan antigens take place, and generated a panel of hybridomas secreting anti-glycan IgG that recognize periodate-sensitive epitopes in soluble egg antigens in the parasites, as well as recognizes a neoglycoprotein containing a pentasaccharide with all the Lex sequence. One particular murine mAb, an IgG3 designated F8A1.1, bound to glycoproteins and glycolipids from schistosome adults and human promyelocytic leukemic HL-60 cells that express Lex antigens, as assessed by a wide selection of approaches like immunofluorescence staining, confocal microscopy, flow cytometry and western blotting, as well as overlay assays of glycolipids right after thinlayer chromatography. In contrast, F8A1.1 bound weakly to cercariae, 3-h schistosomula and human Jurkat cells. We also directly compared the glycan specificity of F8A1.1 with commercially obtainable anti-CD15 IgG1 (clone W6D3) working with a defined glycan microarray.Triphenylphosphinechlorogold site The results demonstrated that F8A1.PMID:23074147 1 recognized glycans expressing Lex epitopes within a terminal nonreducing position, whereas anti-CD15 bound to glycans with multiple repeats of Lex epitopes, but to not glycans having a single, terminal LexIntroduction The parasitic blood flukes in the genus Schistosoma synthesize glycoconjugates having a massive range of complicated glycan structures, numerous of that are immunogenic in infected hosts and have already been proposed to play essential roles in the immunobiology of infections (Cummings and Nyame 1996; Van Die and Cummings 2010; Meevissen et al. 2012b; Van Diepen et al. 2012). Schistosome-derived glycans have lots of structural features which are distinct from mammalian cell derived glycans (Nyame et al. 2004; Hokke et al. 2007), including the lack of sialic acid residues, which are popular terminal sugar residues on mammalian cell glycoconjugates (Nyame et al. 1987; Nyame et al. 2004). In addition, many schistosome N-glycans have outer branches containing the LacdiNAc [LDN; GalNAc1-4N-acetylglucosamine (GlcNAc)-R] backbone in lieu of the lactosamine [LN; Galactose (Gal)-1-4GlcNAc-R] structure commonly located on mammalian glycans (Srivatsan et al. 1992b; Srivatsan et al. 1994; Nyame et al. 1999; Van Die and Cummings 2010). Such LDN structures may well be modified additional with fucose (Fuc) residues in very unusual linkages to yield a sizable assortment of novel fucosylated LDN structures (Cummin.
