Stambaugh Alexandra, Stott Matthew A, Meena Gopikrishnan G, Tamhankar Manasi, Carrion Ricardo, Patterson Jean L, Hawkins Aaron R, Schmidt Holger
School of Engineering, University of California Santa Cruz, Santa Cruz, CA 95064 USA.
Department of Electrical and Computer Engineering, Brigham Young University, Provo UT 84602 USA.
IEEE J Sel Top Quantum Electron. 2021 Jul-Aug;27(4). doi: 10.1109/jstqe.2020.3024239. Epub 2020 Sep 16.
Infectious disease outbreaks such as Ebola and other Viral Hemorrhagic Fevers (VHF) require low-complexity, specific, and differentiated diagnostics as illustrated by the recent outbreak in the Democratic Republic of Congo. Here, we describe amplification-free spectrally multiplex detection of four different VHF total RNA samples using multi-spot excitation on a multimode interference waveguide platform along with combinatorial fluorescence labeling of target nucleic acids. In these experiments, we observed an average of 8-fold greater fluorescence signal amplitudes for the Ebola total RNA sample compared to three other total RNA samples: Lake Victoria Marburg Virus, Ravn Marburg Virus, and Crimean-Congo Hemorrhagic Fever. We have attributed this amplitude amplification to an increased amount of RNA during synthesis of soluble glycoprotein in infection. This hypothesis is confirmed by single molecule detection of the total RNA sample after heat-activated release from the carrier microbeads. From these experiments, we observed at least a 5.3x higher RNA mass loading on the Ebola carrier microbeads compared to the Lake Victoria Marburg carrier microbeads, which is consistent with the known production of soluble glycoprotein during infection.
如埃博拉和其他病毒性出血热（VHF）等传染病爆发需要低复杂度、特异性和差异化的诊断方法，刚果民主共和国最近的疫情就说明了这一点。在此，我们描述了在多模干涉波导平台上使用多点激发以及对靶核酸进行组合荧光标记，对四种不同的VHF总RNA样本进行无扩增光谱多重检测。在这些实验中，我们观察到与其他三种总RNA样本（维多利亚湖马尔堡病毒、拉夫恩马尔堡病毒和克里米亚-刚果出血热）相比，埃博拉总RNA样本的荧光信号幅度平均大8倍。我们将这种幅度放大归因于感染过程中可溶性糖蛋白合成期间RNA量的增加。通过从载体微珠热激活释放后对总RNA样本进行单分子检测，证实了这一假设。从这些实验中，我们观察到与维多利亚湖马尔堡载体微珠相比，埃博拉载体微珠上的RNA质量负载至少高5.3倍，这与感染期间可溶性糖蛋白的已知产生情况一致。
