Matsuda H, Mitsuda H, Nakamura N, Furusawa S, Mohri S, Kitamoto T
Department of Immunobiology, Faculty of Applied Biological Science, Hiroshima University, Japan.
FEMS Immunol Med Microbiol. 1999 Mar;23(3):189-94. doi: 10.1111/j.1574-695X.1999.tb01238.x.
Chickens were immunized with human prion protein (PrP) peptide H25 (amino acid residues 25-49) coupled to keyhole limpet hemocyanin. From a fusion experiment using the chicken fusion partner cell line MuH1 and immune spleen cells, one mAb, HUC2-13, was generated which reacted with the peptide. HUC2-13 was specific for a pentapeptide (RPKPG) of the N-terminal of the peptide H25. In Western blotting analysis, the mAb reacted with PrP materials from a human Creutzfeldt-Jakob disease (CJD) case and the membrane fraction from normal murine brain, but not with the same materials pretreated with proteinase K. When compared with the HUC2-13 and the conventional mouse mAb 3F4, the background stainings using the HUC2-13 were minimal. In immunohistochemistry, the HUC2-13 stained positively with kuru plaques in brain sections from patients with Gerstmann-Straussler syndrome (GSS), and also reacted with synaptic structures of the CJD patients. However, any immunolabelings using the HUC2-13 were not observed in the section from a patient with amyotrophic lateral sclerosis (ALS) as CJD-negative control. These results indicate that the mAb HUC2-13 is a suitable tool for immunological and diagnostic analyses of prion disease in humans and other mammals.
用与人朊病毒蛋白（PrP）肽H25（氨基酸残基25 - 49）偶联的钥孔血蓝蛋白免疫鸡。通过使用鸡融合伴侣细胞系MuH1和免疫脾细胞进行融合实验，产生了一种与该肽反应的单克隆抗体HUC2 - 13。HUC2 - 13对肽H25 N端的一个五肽（RPKPG）具有特异性。在蛋白质印迹分析中，该单克隆抗体与来自一名人类克雅氏病（CJD）病例的PrP物质以及正常鼠脑的膜组分反应，但不与用蛋白酶K预处理的相同物质反应。与HUC2 - 13和传统小鼠单克隆抗体3F4相比，使用HUC2 - 13的背景染色最少。在免疫组织化学中，HUC2 - 13在格斯特曼 - 施特劳斯勒综合征（GSS）患者脑切片中的库鲁斑块上呈阳性染色，并且也与CJD患者的突触结构反应。然而，在作为CJD阴性对照的肌萎缩侧索硬化症（ALS）患者的切片中未观察到使用HUC2 - 13的任何免疫标记。这些结果表明单克隆抗体HUC2 - 13是用于人类和其他哺乳动物朊病毒病免疫和诊断分析的合适工具。
