通过微流控震荡诱导转换(Micro-QuIC)对折叠错误的蛋白质进行现场快速扩增和可视检测

Dong Jun Lee, Peter R. Christenson, Gage Rowden, Nathan C. Lindquist, Peter A. Larsen, Sang-Hyun Oh
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摘要

蛋白质错误折叠疾病,如朊病毒病、阿尔茨海默氏症和帕金森氏症,都有一个共同的分子机制,涉及特定蛋白质的错误折叠和聚集。目前迫切需要能准确检测这些错误折叠蛋白质的床旁诊断(POC)技术,以促进早期诊断和干预。在这里,我们介绍了微流体震荡诱导转换(Micro-QuIC),这是一种新型声学流体平台,可用于快速、灵敏地检测蛋白质错误折叠疾病。我们以慢性消耗性疾病(CWD)为模型系统展示了我们技术的实用性,因为野生白尾鹿的样本很容易获得,而且慢性消耗性疾病与人类蛋白质错误折叠疾病有相似之处。声流体混合技术能使试剂在高雷诺数条件下均匀混合,从而大大加快了 CWD 诊断的周转时间。我们的 Micro-QuIC 分析法扩增朊病毒的速度明显快于目前的黄金标准--实时震荡诱导转换(RT-QuIC)。此外,我们还将 Micro-QuIC 与基于金纳米粒子的裸眼检测方法整合在一起,这样就可以目测区分 CWD 阳性样本和 CWD 阴性样本,而无需使用笨重的荧光检测模块。这种集成创建了一个快速的 POC 检测平台,能够检测与各种蛋白质折叠错误疾病相关的折叠错误蛋白质。
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Rapid on-site amplification and visual detection of misfolded proteins via microfluidic quaking-induced conversion (Micro-QuIC)
Protein misfolding diseases, such as prion diseases, Alzheimer’s, and Parkinson’s, share a common molecular mechanism involving the misfolding and aggregation of specific proteins. There is an urgent need for point-of-care (POC) diagnostic technologies that can accurately detect these misfolded proteins, facilitating early diagnosis and intervention. Here, we introduce the microfluidic quaking-induced conversion (Micro-QuIC), a novel acoustofluidic platform for the rapid and sensitive detection of protein misfolding diseases. We demonstrate the utility of our technology using chronic wasting disease (CWD) as a model system, since samples from wild white-tailed deer are readily accessible, and CWD shares similarities with human protein misfolding diseases. Acoustofluidic mixing enables homogeneous mixing of reagents in a high-Reynolds-number regime, significantly accelerating the turnaround time for CWD diagnosis. Our Micro-QuIC assay amplifies prions significantly faster than the current gold standard, real-time quaking-induced conversion (RT-QuIC). Furthermore, we integrated Micro-QuIC with a gold nanoparticle-based, naked-eye detection method, which enables visual discrimination between CWD-positive and CWD-negative samples without the need for a bulky fluorescence detection module. This integration creates a rapid, POC testing platform capable of detecting misfolded proteins associated with a variety of protein misfolding diseases.
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