Lee B Y, Ahn M H, Kim H C, Min D Y
Department of Parasitology, Institute of Biomedical Science, Hanyang University College of Medicine, Seoul, 133-791, Korea.
Korean J Parasitol. 2001 Mar;39(1):67-75. doi: 10.3347/kjp.2001.39.1.67.
This experiment was focused on the characterization of anti-Toxoplasma monoclonal antibodies (mAbs) and the effect of mAbs on the parasite invasion of mouse peritoneal macrophages. Twenty eight mAbs including M110, M556, R7A6 and M621 were characterized by Ab titer, immunoglobulin isotyping and western blot pattern. Antibody titer (optical density) of 4 mAbs, M110, M556, R7A6 and M621, were 0.53, 0.67, 0.45 and 0.39 (normal mouse serum; 0.19) with the same IgG1 isotypes shown by Enzyme-linked immunosorbent assay (ELISA). Western blot analysis showed that M110, M556, R7A6 and M621 reacted with the 33 kDa (p30), 31 kDa (p28), 43 kDa and 36 kDa protein. Immunogold labelling of mAbs M110, M556, R7A6 and M621 reacted with the surface membrane, dense granules and parasitophorous vacuolar membrane (PVM), rhoptries and cytoplasm of tachyzoite, respectively. For in vitro assay, preincubation of tachyzoites with four mAbs, M110, M556, R7A6 and M621 resulted in the decrease of the number of infected macrophages (p < 0.05) and the suppression of parasite multiplication at 18 h post-infection. Four monoclonal antibodies including M110 (SAG1) were found to have an important role in the inhibition of macrophage invasion and T. gondii multiplication in vitro, and these mAbs may be suitable for vaccine candidates, diagnostic kit and for chemotherapy.
本实验着重于抗弓形虫单克隆抗体（mAb）的特性鉴定以及mAb对寄生虫入侵小鼠腹腔巨噬细胞的影响。通过抗体效价、免疫球蛋白分型和蛋白质印迹图谱对包括M110、M556、R7A6和M621在内的28种mAb进行了特性鉴定。通过酶联免疫吸附测定（ELISA）发现，M110、M556、R7A6和M621这4种mAb的抗体效价（光密度）分别为0.53、0.67、0.45和0.39（正常小鼠血清为0.19），且均为相同的IgG1亚型。蛋白质印迹分析表明，M110、M556、R7A6和M621与33 kDa（p30）、31 kDa（p28）、43 kDa和36 kDa的蛋白发生反应。mAb M110、M556、R7A6和M621的免疫金标记分别与速殖子的表面膜、致密颗粒、寄生泡膜（PVM）、棒状体和细胞质发生反应。在体外试验中，将速殖子与4种mAb（M110、M556、R7A6和M621）预孵育，导致感染巨噬细胞数量减少（p < 0.05），并在感染后18小时抑制了寄生虫的增殖。发现包括M110（SAG1）在内的4种单克隆抗体在体外抑制巨噬细胞入侵和弓形虫增殖方面具有重要作用，这些mAb可能适用于候选疫苗、诊断试剂盒和化疗。
