细粒棘球蚴生发层细胞系的建立

ESTABLISHMENT OF GERMINAL CELL LINE OF ECHINOCOCCUS GRANULOSUS

目的：为获得能在体外稳定繁殖的细粒棘球蚴细胞系。方法：用研磨法将绵羊源细粒棘球蚴生发层处理得分散的生发细胞，将其在含１０％—２０％小牛血清的ＲＰＭＩ１６４０培养基在２４孔培养板和包被胶原的２４孔细胞培养板上交替培养至１４代，随后在常规２４孔培养板上连续传代培养；用倒置显微镜观察细胞形态及生长繁殖情况；将培养细胞接种ＢＡＬＢ／ｃ小鼠以观察其形成继发性包囊的能力；用ＥＬＩＳＡ法研究细胞系抗原的免疫学特征。结果：生发层细胞已在体外连续培养７５代以上，生长良好；细胞系细胞呈圆形，具有形成合胞体乃至类组织块的趋势，在４℃冰箱中至少可保存半个月，在液氮中至少可保存１０个月；将细胞接种ＢＡＬＢ／ｃ小鼠后３个月至７个月，解剖未见细粒棘球蚴包囊生长；细胞系抗原可和抗生发层体抗原、原头节可溶性抗原、囊液抗原的小鼠血清及人工感染原头节的阳性小鼠血清起反应。结论：细粒棘球蚴生发层细胞系已经建立。

AIM:To establish a cell line of Echinococcus granulosus. METHODS:The proliferating membranes striped from liver cysts in a naturally infected sheep from Urumq were released to monodispersed germinal cells by grinding. The germinal cells were cultivated in the RPMI 1640 medium supplemented with 10%—20% calf serum on collagen coated 24 well culture plate alternately from passage 1 to passage 14,and then continued. The morphological feature and growth situation were observed by light microscopy.The cultured cells were inoculated into BALB/c mice to identify the infectivity . ELISA was used to determine the immunogenicity of the cells . RESULTS: The germinal cells have been cultivated continuously up to passage 75. The subcultured cells were circular in shape with smooth surface and had the tendency to form syncytia and tissue like masses. The cells from this cell line could be stored for at least 15 days in refrigerator at 4℃ and 10 months in liquid nitrogen. No cyst materials were detected in the mice inoculated with cells. The antigens from cell line could react with positive sera from mice infected with protoscoleces and sera against secreted antigens of cyst membrane ,soluble antigens from cyst membrane, soluble antigens from protoscoleces and SHF . CONCLUSION: A germinal cell line of Echinococcus granulosus was successfully established.