Two-Fluorophore Mobile Phone Imaging of Biplexed Real-Time NAATs Overcomes Optical Artifacts in Highly Scattering Porous Media.
Two-Fluorophore Mobile Phone Imaging of Biplexed Real-Time NAATs Overcomes Optical Artifacts in Highly Scattering Porous Media.
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DOI：
10.1021/acs.analchem.0c02000
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发表时间：
2020-10-06
影响因子：
7.4
通讯作者：
Yager, Paul
中科院分区：
文献类型：
--
作者：
Shah, Kamal G.;Kumar, Sujatha;Singh, Vidhi;Hansen, Louise;Heiniger, Erin;Bishop, Joshua D.;Lutz, Barry;Yager, Paul
核酸扩增测试 (NAAT) 在实验室和临床环境中很常见，因为其获得结果的时间短且具有极高的敏感性和特异性。实验室 NAAT 包括用于减少假阴性的板载阳性对照和用于实现实时荧光检测的专用硬件。最近将 NAAT 转化为家庭测试的努力牺牲了实验室 NAAT 的一项或多项优势，例如灵敏度、内部扩增控制 (IAC) 或得出结果的时间。在这篇手稿中，我们在论文中描述了一种基于手机的双工 NAAT 实时成像策略。该策略包括：(1) 使用带有多通道激发和发射滤光片的手机，在闪光灯和相机上对玻璃纤维膜中双工 NAAT 中不同荧光团标记探针类型的信号进行成像； (2) 分析手机图像的红色和绿色通道之间的差异荧光信号，以克服由于折射率变化而在加热玻璃纤维垫中强烈蒸发引起的光学伪影。我们证明，即使在包含 100k 拷贝共扩增 IAC DNA 模板的双工等温链置换扩增反应中，差异荧光成像也能在我们实验室的“MD NAAT”平台中实现低检测限（316 拷贝耐甲氧西林金黄色葡萄球菌 DNA）。这些结果表明，双荧光团手机成像可以将现有的基于实验室的实时 NAAT 的优势转化为护理点。
Nucleic acid amplification tests (NAATs) are common in laboratory and clinical settings because of their low time to result and exquisite sensitivity and specificity. Laboratory NAATs include onboard positive controls to reduce false negatives and specialized hardware to enable real-time fluorescence detection. Recent efforts to translate NAATs into at-home tests sacrifice one or more of the benefits of laboratory NAATs, such as sensitivity, internal amplification controls (IACs), or time to result. In this manuscript, we describe a mobile-phone-based strategy for real-time imaging of biplexed NAATs in paper. The strategy consisted of: (1) using mobile phones with multipass excitation and emission filters on the flash and camera to image the signal from distinct fluorophore-labeled probe types in a biplexed NAAT in a glass fiber membrane; and (2) analyzing the differential fluorescence signal between the red and green color channels of phone images to overcome a strong evaporation-induced optical artifact in heated glass fiber pads due to changes in the refractive index. We demonstrated that differential fluorescence imaging enabled low limits of detection (316 copies of methicillin-resistant Staphylococcus aureus DNA) in our lab’s “MD NAAT” platform, even in biplexed isothermal strand displacement amplification reactions containing 100k copies of coamplifying IAC DNA templates. These results suggest that two-fluorophore mobile phone imaging may enable translating the benefits of extant laboratory-based, real-time NAATs to the point of care.
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影响因子：
7.4
作者：
Rodriguez NM;Linnes JC;Fan A;Ellenson CK;Pollock NR;Klapperich CM
通讯作者：
Klapperich CM
影响因子：
7.4
作者：
Huang S;Abe K;Bennett S;Liang T;Ladd PD;Yokobe L;Anderson CE;Shah K;Bishop J;Purfield M;Kauffman PC;Paul S;Welch AE;Strelitz B;Follmer K;Pullar K;Sanchez-Erebia L;Gerth-Guyette E;Domingo G;Klein E;Englund JA;Fu E;Yager P
通讯作者：
Yager P
影响因子：
17.1
作者：
Rodriguez-Manzano J;Karymov MA;Begolo S;Selck DA;Zhukov DV;Jue E;Ismagilov RF
通讯作者：
Ismagilov RF
影响因子：
4.4
作者：
Hazelton, Briony;Gray, Timothy;Kok, Jen
通讯作者：
Kok, Jen
影响因子：
7.4
作者：
Shah, Kamal G.;Singh, Vidhi;Yager, Paul
通讯作者：
Yager, Paul