Md Ahasan Ahamed, Zhikun Zhang, Aneesh Kshirsagar, Anthony J Politza, Usha Sethuraman, Srinivasan Suresh, Steven Hicks, Feng Guo, Weihua Guan
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引用次数: 0
Abstract
While most children with COVID-19 experience mild symptoms or remain asymptomatic, some may develop severe complications. Early identification of children at risk for severe outcomes is essential to ensuring timely and effective intervention. Recent studies have identified alterations in salivary microRNA (miRNA) expression levels as promising biomarkers for predicting severe complications in children. However, there remains a need for a rapid, noninvasive, and quantitative method to detect miRNA expression level changes, as their upregulation or downregulation serves as a hallmark of various diseases, providing an alternative to sequencing-based methods. Here, we developed a highly specific and sensitive ligation-coupled recombinase polymerase amplification (RPA) assay for quantitatively detecting multiplex severe and nonsevere miRNAs on a portable platform. The assay begins with an miRNA-templated annealing and ligation reaction of miR-1273, miR-296, and miR-29, followed by an RPA reaction. We quantified 100 pM to 1 fM, resolving 1 fM, with 100% specificity. Next, we validated portable extraction against benchtop extraction, achieving R2 > 0.85 and r > 0.92 in clinical samples. Finally, testing 154 clinical samples revealed severe miRNA downregulation compared to nonsevere cases. The assay achieved high diagnostic accuracy with an area under the curve (AUC) of 0.98. This platform would empower clinicians to make informed decisions, optimize resource allocation, and improve outcomes, particularly in point-of-care (POC) settings.
期刊介绍:
ACS Sensors is a peer-reviewed research journal that focuses on the dissemination of new and original knowledge in the field of sensor science, particularly those that selectively sense chemical or biological species or processes. The journal covers a broad range of topics, including but not limited to biosensors, chemical sensors, gas sensors, intracellular sensors, single molecule sensors, cell chips, and microfluidic devices. It aims to publish articles that address conceptual advances in sensing technology applicable to various types of analytes or application papers that report on the use of existing sensing concepts in new ways or for new analytes.