亚马逊利什曼原虫无鞭毛体的体外培养及其抗原性分析  

Axenic cultivation and indirect immunofluorescent analyasis of Leishmania anazonesis amastigotes

作　　者：王华民[1] Soong Lynn 

机构地区：[1]海南医学院微生物学与免疫学教研室,海南海口571101 [2]Dept.of Microbiology & Immunology,University of Texas Medical Branch,Galve-ston,TX.,USA.

出　　处：《中国热带医学》2006年第5期755-756,810,共3页China Tropical Medicine

摘　　要：目的建立亚马逊利什曼原虫无鞭毛体体外培养方法,并用间接免疫荧光法分析其抗原特异性。方法小鼠皮损组织处获得的无鞭毛体(Al)、由前鞭毛体转化来的无鞭毛体(Ap)、J774.G8巨噬细胞来源的无鞭毛体(Aj)置于改良后的Schneider’sDrosophila培养基,pH4.6、33℃条件下培养,取培养物光镜下观察并用间接免疫荧光法分析。结果三种不同来源无鞭毛体体外培养物光镜检查形态相似,均与纯培养获得的无鞭毛体一样。与无鞭毛体特异性蛋白P-8和P-4特异性单克隆抗体进行间接免疫荧光分析显示,三种不同来源的无鞭毛体均出现阳性,且浓度与部位相似。而前鞭毛体与抗P-8单克隆抗体(mAb)染色呈弱阳性反应,与抗P-4单克隆抗体呈阴性反应。相反,GP-46/M-2McAb与前鞭毛体染色呈强阳性反应,与无鞭毛体仅呈弱阳性反应。结论在33℃条件下,用改良的Schneider’sDrosophila培养基进行无鞭毛体体外培养获得成功。用阶段特异性单克隆抗体,进行间接免疫荧光分析法可鉴定亚马逊利什曼原虫的无鞭毛体。Objective To establish a method for culture of Leishmania amazonensis amastigotes in vitro and identify the antigenie specificity of amastigotes with indirect immunofluorescent assay. Methods Amastigotes derived from mouse lesions, infected J774. G8 macrophages, or promastigotes, were cultured in modified Schneider＇s Drosophila medium under pH 4.6 and 33℃. The cultured amastigotes were observed by light microscope and analyzed with indirect immunofluoreseent assay. Resets Three different sources of amastigotes analyzed by light microscope showed similar morphological features. Indirect immunofluoreseent analyses employing mAbs specific to amastigote proteins P - 4 and P - 8 indicated positive staining for all three types of amastigotes with similar intensity and localization pattern, Anti - P - 8 but anti - P - 4 mAb showed weak staining for promastigotes. In eontraest, GP - 46/ M - 2 mAb was reactive strongly to promistigotes but weak staining for amistigotes. Conclusion Under 33℃, amastigotes were cultured successfully using modified conditions, and Leishmania amazonensis amastigotes can be identified with indirect immunofluoreseent analysis by employing stage- specific mAbs.

关 键 词：亚马逊利什曼原虫 无鞭毛体 体外培养 间接免疫荧光 

分 类 号：R382.2[医药卫生—医学寄生虫学]