Aim: The current study was designed to isolate and characterize Toxocara vitulorum glycoprotein antigens and then to evaluate its potency in accurate diagnosis of toxocariasis.
Materials and methods: T. vitulorum glycoprotein fractions were isolated using Con-A affinity chromatography. The fractions characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and immunoblot assay. Mass spectrometric analysis was used for identification of proposed structure of the N-acetylglucosamine (GlcNAc) fraction. Enzyme-linked immunosorbent assay (ELISA) was used to assess the diagnostic potential of the isolated fractions.
Results: Surface of T. vitulorum adult worm revealed two glycoprotein fractions rich in glucose (Glc) and GlcNAc. Three bands of molecular weight 212kDa, 107 kDa, and 93 kDa were detected in Glc fraction by SDS-PAGE. These bands were also detected in GlcNAc fraction with an additional band of 49 kDa. GlcNAc fraction showed more diagnostic potency of calves' toxocariasis; 79% than Glc fraction; 46.9% by indirect ELISA. The additional band of 49 kDa in GlcNAc fraction is probably responsible for its higher diagnostic potentials. Western blotting verified the immunoreactivity of the Glc and GlcNAc isolated fraction as they reacted with calves sera infected with toxocariasis. The proposed structure of GlcNAc fraction was Ser-Meth-Arg-O-methylated GlcNAc.
Conclusion: GlcNAc-rich fraction of T. vitulorum can be successfully utilized in the diagnosis of calves' toxocariasis.
Keywords: Con A; N-acetylglucosamine; affinity chromatography; calves toxocariasis; enzyme-linked immunosorbent assay; mass spectrometric analysis.