Membrane proteins and their antigenicity of Toxoplasma gondii

Surface membrane proteins of virulent RH strain and tissue cyst-forming Fukaya strain of Toxoplasma gondii were analyzed by SDS-polyacrylamide gel electrophoresis after LPO-catalyzed surface iodination and lectin blotting, then identified the zoite-specific antigens. Prior to the analyses, purification of RH tachyzoites from mouse peritoneal exudate and of Fukaya bradyzoites from mouse brain tissues were performed by centrifugation on the discontinuous Percoll density-gradient. Tachyzoites were obtained at the interface of 50 per cent and 60 per cent Percoll solution and brain cysts were harvested at the interfaces of 40-50 per cent and 50-60 per cent, then bradyzoites were obtained by treating the cysts with hypertonic solution. The LPO-catalyzed iodination detected 15 KDa and 14 KDa proteins of bradyzoites and 30 KDa protein of tachyzoites as major bands with several other minor bands. But Con A blotting revealed some bands of 200 K-50 KDa glycoproteins of bradyzoites and 52 KDa band as major and minor bands of 33 K-20 KDa of tachyzoites. Phytohemagglutinin did not detect any band in the two forms. EITB with anti-Fukaya antibody and anti-RH antibody revealed cross-reactivities between the two forms. Despite the cross-reactivity, anti-Fukaya antibody reacted with 15 KDa band of bradyzoites specifically and, anti-RH antibody with 52 KDa, 30 KDa, and 25 KDa bands of tachyzoites, respectively. It was identified that 15 KDa protein in bradyzoite, which was not a glycoprotein, was a major membrane protein with sufficient antigenicity, and in the case of tachyzoite, 52 KDa surface glycoprotein (gp52) with specific antigenicity might be added to the major surface protein, p30.