Generation of monoclonal antibody that distinguishes PrPSc from PrPC and neutralizes prion infectivity
Generation of monoclonal antibody that distinguishes PrPSc from PrPC and neutralizes prion infectivity
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DOI：
10.1016/j.virol.2009.08.025
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发表时间：
2009-11-25
期刊：
影响因子：
3.7
通讯作者：
Shinagawa, Morikazu
中科院分区：
文献类型：
--
作者：
Horiuchi, Motohiro;Karino, Ayako;Shinagawa, Morikazu
为了建立PrPSc特异性mAb，我们用从朊病毒感染的小鼠中纯化的PrPSc免疫Pmp（-/-）小鼠。使用这种方法，我们获得了mAb 6 H10，其与用蛋白酶K处理的PrPSc反应，但不与用超过3 M的GdnHCl预处理的PrPSc反应。与此相反，泛PrP单克隆抗体的反应性随着用于预处理PrPSc的GdnHCl浓度的增加而增加。在组织印迹分析中，mAb 6 H10在非变性组织印迹上显示阳性反应，但当通过高压灭菌预处理组织印迹时，反应性较低。表位分析表明，PrP的极端C-末端可能是mAb 6 H10的表位的一部分。MAb 6 H10免疫沉淀来自感染朊病毒的小鼠、绵羊和牛脑的PrPSc。此外，用mAb 6 H10预处理纯化的PrPSc使感染滴度降低超过1 log。总之，这些结果表明，mAb 6 H10识别PrPSc上与朊病毒感染性相关的构象表位。(C)2009爱思唯尔公司All rights reserved.
To establish PrPSc-specific mAbs, we immunized Pmp(-/-) mice with PrPSc purified from prion-infected mice. Using this approach, we obtained mAb 6H10, which reacted with PrPSc treated with proteinase K, but not with PrPSc pretreated with more than 3 M GdnHCl. In contrast, reactivity of pan-PrP mAbs increased with increasing concentrations of GdnHCl used for pretreatment of PrPSc. In histoblot analysis, mAb 6H10 showed a positive reaction on a non-denatured histoblot but reactivity was lower when the histoblot was pretreated by autoclaving. Epitope analysis suggested that the extreme C-terminus of PrP is likely to be part of the epitope for mAb 6H10. MAb 6H10 immunoprecipitated PrPSc from brains of mice, sheep, and cattle infected with prions. Furthermore, pretreatment of purified PrPSc with mAb 6H10 reduced the infectious titer more than 1 log. Taken together, these results suggest that mAb 6H10 recognizes a conformational epitope on PrPSc that is related to prion infectivity. (C) 2009 Elsevier Inc. All rights reserved.