Generation of monoclonal antibodies against prion proteins with an unconventional nucleic acid-based immunization strategy
Generation of monoclonal antibodies against prion proteins with an unconventional nucleic acid-based immunization strategy
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DOI：
10.1016/s0168-1656(99)00115-7
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发表时间：
1999-08-20
影响因子：
4.1
通讯作者：
Bodemer, W
中科院分区：
文献类型：
--
作者：
Krasemann, S;Jürgens, T;Bodemer, W
朊病毒病属于影响人类和动物的一组神经退行性疾病。人类疾病包括库鲁病、克雅氏病（CJD）、格斯特曼-施特劳斯勒-沙因克综合征（GSS）和致命性家族性失眠症（FFI）。朊病毒疾病的病理机制尚不清楚。因此，单克隆抗体（mAb）将为这些疾病的诊断和基础研究提供有价值的工具。与传统策略相反，我们开发了一种基于核酸注射到不耐受的 PrP(0/0)-小鼠中的免疫方案。将编码不同人类朊病毒蛋白（包括与 CJD、GSS 和 FFP 相关的突变序列）的 DNA 或 RNA 注射到肌肉组织中。这些小鼠首先接种了编码 PRNP 的 DNA 质粒，并用表达 PRNP 的 DNA、RNA 或重组塞姆利基森林病毒 (SFV) 颗粒进行加强。杂交瘤制备后，获得了不同的针对朊病毒蛋白的单克隆抗体，并通过肽-ELISA、Western blot、免疫荧光和免疫沉淀分析了它们的结合行为。我们的单克隆抗体针对四种不同的线性表位，也可以识别天然朊病毒蛋白的不连续区域。因此，可以证明，用 DNA 和减毒活 SF 病毒对不耐受的小鼠进行免疫是诱导广泛免疫反应的一种有价值的方法，最终导致产生一组用于基础科学和诊断的单克隆抗体。 (C) 1999 Elsevier Science B.V. 保留所有权利。
Prion diseases belong to a group of neurodegenerative disorders affecting humans and animals. The human diseases include kuru, Creutzfeldt-Jakob disease (CJD), Gerstmann-Straussler-Scheinker syndrome (GSS) and fatal familial insomnia (FFI). The pathomechanisms of the prion diseases are not yet understood. Therefore, monoclonal antibodies (mAbs) would provide valuable tools in diagnostics as well as in basic research of these diseases. In contrast to conventional strategies we have developed an immunization protocol based on nucleic acid injection into non tolerant PrP(0/0)-mice. DNA or RNA coding for different human prion proteins including the mutated sequences associated with CJD, GSS and FFP were injected into muscle tissue. The mice were primarily inoculated with DNA-plasmids encoding PRNP and boosted either with DNA, RNA or recombinant Semliki Forest virus (SFV) particles expressing PRNP. After hybridoma preparation, different mAbs against prion proteins were obtained and their binding behaviour was analysed by peptide-ELISA, Western blot, immunofluorescence and immunoprecipitation. Our mAbs are directed against four different linear epitopes and may also recognize discontinuous regions of the native prion protein. It could, therefore, be demonstrated that immunization of non tolerant mice with DNA and live attenuated SF virus is a valuable means to induce a broad immune response leading eventually to the generation of a panel of mAbs for basic science as well as for diagnostics. (C) 1999 Elsevier Science B.V. All rights reserved.