Ivanovova Natalia, Handzusova Martina, Hanes Jozef, Kontsekova Eva, Novak Michal
Institute of Neuroimmunology, Slovak Academy of Sciences, Bratislava, Slovak Republic.
J Immunol Methods. 2008 Nov 30;339(1):17-22. doi: 10.1016/j.jim.2008.07.014. Epub 2008 Aug 17.
The fetal type of tau phosphorylation always re-appears during pathogenesis of Alzheimer's disease and related tauopathies. The major obstacle in the study of the fetal tau phosphorylation has been the lack of a simple and reproducible purification method yielding fetal tau with high purity and unmodified phosphorylation pattern. We have developed a two-step, highly efficient purification procedure of perchloric acid-extracted fetal tau by immunoaffinity chromatography and trichloroacetic acid (TCA) precipitation. The method yielded tau with more than 90% purity. Most importantly, purified fetal tau exhibited unmodified phosphorylation pattern as confirmed by phosphorylation-dependent antibodies. In summary, this purification process preserves and protects unstable phosphoresidues from dephosphorylation and allows their detailed molecular analysis especially in the pathogenesis of Alzheimer's disease and related tauopathies.
胎儿型tau蛋白磷酸化在阿尔茨海默病及相关tau蛋白病的发病过程中总是会再次出现。胎儿tau蛋白磷酸化研究中的主要障碍一直是缺乏一种简单且可重复的纯化方法，无法获得高纯度且磷酸化模式未改变的胎儿tau蛋白。我们通过免疫亲和色谱和三氯乙酸（TCA）沉淀开发了一种两步高效纯化程序，用于纯化高氯酸提取的胎儿tau蛋白。该方法产生的tau蛋白纯度超过90%。最重要的是，经磷酸化依赖性抗体证实，纯化的胎儿tau蛋白呈现未改变的磷酸化模式。总之，这种纯化过程可保留并保护不稳定的磷酸化残基不被去磷酸化，并允许对其进行详细的分子分析，尤其是在阿尔茨海默病及相关tau蛋白病的发病机制研究中。
