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一种用于检测重组真实tau原纤维的优化过滤捕获分析方法。

An optimized filter trap assay for detecting recombinant authentic tau fibrils.

作者信息

Balaj Allison R, Rothstein Thomas L, Kaku Hiroaki

机构信息

Department of Investigative Medicine, Western Michigan University Homer Stryker M.D. School of Medicine, Kalamazoo, MI, USA.

出版信息

Free Neuropathol. 2024 Aug 2;5:15. doi: 10.17879/freeneuropathology-2024-5497. eCollection 2024 Jan.

Abstract

The development and optimization of the Filter Trap Assay (FTA) for the detection of authentic tau fibrils mark a pivotal advancement in the realm of tauopathy research, particularly by addressing the limitations of using polyanion-induced tau fibrils, which structurally differ from those isolated from tauopathy patients. Recently it has been shown that truncated tau fragment (297-391), also termed dGAE, can form authentic tau fibrils in the absence of polyanions. This study introduces a refined protocol that reliably detects authentic tau fibrils in a physiologically relevant framework, utilizing nitrocellulose membranes to achieve heightened sensitivity. Our investigation highlights the superior efficacy of sarkosyl, an anionic surfactant traditionally used to prepare protein lysates from brains and cultured neurons, in preserving the aggregated state of tau dGAE fibrils , underscoring its potential for further exploratory studies. By offering a user-friendly and economically feasible approach, this technique enables a broad range of laboratories to measure the presence of authentic tau fibrils. This methodological enhancement propels our understanding of tauopathies forward and bridges the gap between basic research and advanced structural analyses, enriching the scientific community's methodologies for studying neurodegenerative disorders.

摘要

用于检测真实tau原纤维的滤膜捕获分析(FTA)的开发和优化标志着tau蛋白病研究领域的一项关键进展,特别是通过解决使用聚阴离子诱导的tau原纤维的局限性,后者在结构上与从tau蛋白病患者分离出的原纤维不同。最近有研究表明,截短的tau片段(297-391),也称为dGAE,在没有聚阴离子的情况下可以形成真实的tau原纤维。本研究引入了一种优化方案,该方案在生理相关框架中可靠地检测真实的tau原纤维,利用硝酸纤维素膜提高灵敏度。我们的研究强调了十二烷基肌氨酸钠(一种传统上用于从大脑和培养神经元制备蛋白质裂解物的阴离子表面活性剂)在保持tau dGAE原纤维聚集状态方面的卓越功效,强调了其在进一步探索性研究中的潜力。通过提供一种用户友好且经济可行的方法,该技术使广泛的实验室能够测量真实tau原纤维的存在。这种方法学的改进推动了我们对tau蛋白病的理解,并弥合了基础研究与高级结构分析之间的差距,丰富了科学界研究神经退行性疾病的方法。

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