Department of Biosystems Engineering, The University of Arizona, Tucson, Arizona, 85721, USA.
Department of Biomedical Engineering, The University of Arizona, Tucson, Arizona, 85721, USA.
Sci Rep. 2018 Aug 20;8(1):12438. doi: 10.1038/s41598-018-30797-9.
The recent Zika virus (ZIKV) outbreak has prompted the need for field-ready diagnostics that are rapid, easy-to-use, handheld, and disposable while providing extreme sensitivity and specificity. To meet this demand, we developed a wax-printed paper microfluidic chip utilizing reverse transcription loop-mediated isothermal amplification (RT-LAMP). The developed simple and sensitive ZIKV assay was demonstrated using undiluted tap water, human urine, and diluted (10%) human blood plasma. Paper type, pore size, and channel dimension of various paper microfluidic chips were investigated and optimized to ensure proper filtration of direct-use biological samples (tap water, urine, and plasma) during capillary action-driven flow. Once ZIKV RNA has flowed and reached to a detection area of the paper microfluidic chip, it was excised for the addition of an RT-LAMP mixture with a pH indicator, then placed on a hot plate at 68 °C. Visible color changes from successful amplification were observed in 15 minutes and quantified by smartphone imaging. The limit of detection was as low as 1 copy/μL. The developed platform can also be used for identifying other flaviviruses, such as Chikungunya virus (CHIKV) and Dengue virus (DENV), and potentially other quickly transmitted virus pathogens, towards field-based diagnostics.
最近的寨卡病毒（ZIKV）爆发促使人们需要现场准备的诊断方法，这些方法应快速、易于使用、手持式和一次性使用，同时提供极高的灵敏度和特异性。为了满足这一需求，我们开发了一种利用逆转录环介导等温扩增（RT-LAMP）的蜡印纸微流控芯片。该简单而灵敏的 ZIKV 检测方法已使用未经稀释的自来水、人体尿液和稀释（10%）的人体血浆进行了验证。研究和优化了各种纸微流控芯片的纸张类型、孔径和通道尺寸，以确保在毛细作用驱动的流动过程中适当过滤直接使用的生物样品（自来水、尿液和血浆）。一旦 ZIKV RNA 流动并到达纸微流控芯片的检测区域，就可以将其切除以加入带有 pH 指示剂的 RT-LAMP 混合物，然后将其放在 68°C 的热板上。成功扩增后可在 15 分钟内观察到可见的颜色变化，并通过智能手机成像进行定量。检测限低至 1 拷贝/μL。该开发平台还可用于鉴定其他黄病毒，如基孔肯雅病毒（CHIKV）和登革热病毒（DENV），并可能用于其他快速传播的病毒病原体，以实现现场诊断。
