Paragonimus ohirai: immunobiochemical characterization on the tegumental glycocalyx of excysted juvenile recognized by a monoclonal antibody.
The tegumental glycocalyx of excysted juvenile (EJ) of Paragonimus ohirai was immunobiochemically characterized using a monoclonal antibody (MS-Mab). HPLC gel filtration showed that the antigens detected by two-site ELISA had a molecular weight of greater than or equal to 2 x 10(6) Da (dextran marker). On reduced SDS-PAGE, the glycocalyx antigen retained in the stacking gel was cleaved into several much smaller antigens after pronase treatment. The antigenic activity of the glycocalyx was stable in two-site ELISA to heat and acid treatments, but sensitive to alkali, periodate, base/borohydride, and pronase treatments. Precipitin formation in immunodouble diffusion between MS-Mab and EJ crude antigen was inhibited only by two monosaccharides: galactose and N-acetylgalactosamine. The purified glycocalyx bound strongly to PNA lectin, fairly well to RCA120 lectin, and slightly to SBA lectin, but not to Con A, WGA, UEA-1, DBA, or LFA lectins. Exo-beta-galactosidase treatment increased SBA binding, whereas it decreased PNA binding. PNA was observed to strongly bind to the body surface of living EJ. The antigenic activity of the glycocalyx was remarkably lost by incubation with exo-beta-galactosidase and O-glycanase. The glycocalyx was reactive with sera of P. ohirai-infected rats, and its reactivity was remarkably reduced by O-glycanase treatment. The ELISA level was higher in sera at an early stage of infection than in a late one. These studies show that the EJ tegumental glycocalyx is antigenic in infection, a marked, high molecular weight glycoprotein containing antigenic O-linked sugars, and that the sugar epitope is at the nonreducing terminal of the O-linked sugars and is composed of galactose and N-acetylgalactosamine.
Ikeda
T
T
Department of Medical Zoology, Kanazawa Medical University, Ishikawa, Japan.
Oikawa
Y
Y
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000947:Antigens, Helminth / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D000954:Antigens, Surface / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000276:immunology*; D005779:Immunodiffusion; D008970:Molecular Weight; D010238:Paragonimus / Q000276:immunology*; D011134:Polysaccharides / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000947:Antigens, Helminth; D000954:Antigens, Surface; D006023:Glycoproteins; D011134:Polysaccharides