摘要
目的:α-突触核蛋白(α-Synuclein,α-Syn)是与帕金森病的发病密切相关的蛋白质。关于这种蛋白质在脑内神经元的亚细胞分布迄今尚无定论。制备α-Syn抗体探讨α-Syn在亚细胞的定位。方法:实验于2004-01/05在北京老年医学研究所神经生物研究室完成。采用重组的人α-Syn免疫小鼠,并将其脾细胞与骨髓瘤融合制备出杂交瘤,利用免疫组化和Westernblot法筛选阳性克隆,获得一特异性抗α-Syn单克隆抗体。结果:噬菌体多肽展示分析表明,该抗体识别人α-SynC-末端的一段特异序列,但该序列与大鼠和小鼠的相应序列差一个氨基酸。免疫印记分析表明,该抗体能够检测人、大鼠与小鼠脑组织中的α-Syn。用该抗体进行免疫组化染色结果显示,α-Syn免疫阳性物质主要存在于神经元末梢与核中。结论:所制备的单克隆抗体对α-Syn具有特异性,可用于人、大鼠和小鼠的α-Syn研究。
AIM:Alpha Synuclein(α Syn) is a protein closely linking to the pathogen esis of Parkinson disease.However,its subcellular localization in brain neuron s remains unconfirmed.The aim of this paper is to produce a monoclonal antibody applicable to subcellular localization of α Syn in neurons. METHODS:The experiment was conducted in Department of Neurobiology,Beijing Ins titute of Geriatrics from January to May 2004.Recombinant human α Syn was used to immunize a Balb/c mouse and hybridoma cells were produced by fusion of the m ouse spleen cells with myeloma cells. A positive clone was selected by screening the hybridoma cells using immunohistochemical staining and Western blot analysi s. RESULTS:Phase polypeptide display assay showed that the antibody obtained reco gnized a C terminal amino acid sequence of human α Syn,which was different in one amino acid from the same part of mouse and rat α Syn.As shown in Western blot analysis,the antibody detected the α Syn of either human or mouse and rat brains.Immunohistochemical study using this antibody revealed that α Syn immu noreactivity was localized in both nerve terminals and the nuclei of many brain neurons. CONCLUSION:The above results suggest that the monoclonal antibody produced in this study is specific to α Syn and can be used for α Syn study in human,rat and mouse brains.
出处
《中国临床康复》
CSCD
北大核心
2005年第1期44-46,i001,共4页
Chinese Journal of Clinical Rehabilitation
基金
科技部"九七三项目(G2000057005)
国家自然科学基金项目(30271437
30270482)
北京市自然科学基金项目(7022011)~~
