School of Molecular Sciences, Arizona State University, Tempe, Arizona 85287, United States.
Center for Applied Structural Discovery, The Biodesign Institute, Arizona State University, Tempe, Arizona 85287, United States.
Anal Chem. 2023 Apr 4;95(13):5522-5531. doi: 10.1021/acs.analchem.2c03825. Epub 2023 Mar 9.
Alzheimer's disease (AD) currently affects more than 30 million people worldwide. The lack of understanding of AD's physiopathology limits the development of therapeutic and diagnostic tools. Soluble amyloid-β peptide (Aβ) oligomers that appear as intermediates along the Aβ aggregation into plaques are considered among the main AD neurotoxic species. Although a wealth of data are available about Aβ from in vitro and animal models, there is little known about intracellular Aβ in human brain cells, mainly due to the lack of technology to assess the intracellular protein content. The elucidation of the Aβ species in specific brain cell subpopulations can provide insight into the role of Aβ in AD and the neurotoxic mechanism involved. Here, we report a microfluidic immunoassay for in situ mass spectrometry analysis of intracellular Aβ species from archived human brain tissue. This approach comprises the selective laser dissection of individual pyramidal cell bodies from tissues, their transfer to the microfluidic platform for sample processing on-chip, and mass spectrometric characterization. As a proof-of-principle, we demonstrate the detection of intracellular Aβ species from as few as 20 human brain cells.
阿尔茨海默病（AD）目前影响着全球超过 3000 万人。对 AD 病理生理学的缺乏了解限制了治疗和诊断工具的发展。可溶性淀粉样蛋白-β 肽（Aβ）寡聚体作为 Aβ 聚集形成斑块的中间产物，被认为是主要的 AD 神经毒性物质之一。尽管有大量关于体外和动物模型中 Aβ 的数据，但对人类脑细胞内的 Aβ知之甚少，主要是因为缺乏评估细胞内蛋白质含量的技术。阐明特定脑细胞亚群中的 Aβ 种类可以深入了解 Aβ 在 AD 中的作用以及涉及的神经毒性机制。在这里，我们报告了一种用于原位质谱分析存档人脑组织内细胞内 Aβ 种类的微流控免疫测定法。该方法包括从组织中选择性激光切割单个锥体细胞体，将其转移到微流控平台上进行芯片内样品处理，以及质谱特征分析。作为原理验证，我们证明了从少至 20 个人脑细胞中检测到细胞内 Aβ 种类。
