Role of the rhoptry proteins of sporozoites during hepatocyte invasion by Plasmodium

所有侵袭型的尖端复合体都有位于寄生虫前端的细胞器复合体。细胞器由棒状体、致密颗粒和微线体组成。棒状体是梨形、电子致密、膜结合的细胞器，被认为在寄主细胞的附着、入侵和寄生液泡膜(PVM)的形成中起作用。在入侵过程中，膜物质从棒状体中分泌出来。这些物质被认为被整合到扩展的PVM中。已在恶性疟原虫裂殖子棒状体中发现了几种不同分子量的蛋白质，但尚未对子孢子顶端细胞器进行研究。为了分析子孢子的棒状蛋白，我们试图制备抗鼠疟原虫子孢子棒状蛋白的单抗。伯氏肺孢子虫的子孢子被浓缩，并用氮气减压的方法破坏，以便在温和的条件下释放细胞器。然后用蔗糖密度梯度离心法纯化细胞器。电子显微镜下可观察到亚细胞组分。我们用这些组分免疫BALB/c小鼠或破坏自然子孢子。我们试图用ELISA或IFA试验来克隆产生抗伯氏肺孢子虫子孢子顶器的单抗的杂交瘤细胞。

All invasive forms of Apicomplexa possess a complex of organelles located at the anterior end of the parasite. The organelles comprise rhoptries, dense granules and micronemes. The rhoptries are pear shaped, electron-dense, membrane bound organdies and are considered to have roles in host cell attachment, invasion and formation of parasitophorous vacuole membrane (PVM). Membranous materials are secreted from the rhoptries during the invasion. These materials are thought to be incorporated into the expanding PVM.Several proteins of varying molecular weight have been identified in P.falciparum merozoite rhoptries, but apical organelles of sporozoite have not been investigated. To analyze the rhoptry proteins of sporozoites, we try to produce monoclonal antibodies against sporozoite rhoptry proteins of rodent malaria. P.berghei sporozoites were enriched and were disrupted by nitrogen decompression method in order to release the organelles in a mild condition. Organelles were then, purified by sucrose density gradient centrifugation. Subcellular fractions were observed by the electron microscopy. We immunized BALB/c mice with these fractions or disrupted native sporozoites. We are trying to clone the hybridoma which produce the monoclonal antibody against P.berghei sporozoite apical organelles by using ELISA or IFA test.