The potential of microRNAs (miRNAs) as biomarker candidates in clinical practice for diagnosis, prognosis and treatment response prediction, especially in liquid biopsies, has led to a tremendous demand for techniques that can detect these molecules rapidly and accurately. Hence, numerous achievements have been reported recently in miRNA research. In this review, we discuss the challenges associated with the emerging field of miRNA detection, which are linked to the intrinsic properties of miRNAs, advantages and drawbacks of the currently available technologies and their potential applications in clinical research. We summarize the most promising nucleic acid amplification techniques applied to the in vitro detection of miRNAs, with a particular emphasis on the state of the art for isothermal alternatives to RT-qPCR. We detail the sensitivity, specificity and quantitativity of these approaches, as well as their potential for multiplexing. We also review the different detection formats to which these chemistries have been adapted, including analog readouts such as real-time monitoring, digital counting based on single-molecule amplification in compartments, and surface-based strategies.

在临床实践中，尤其是在液体活检中，microRNA（miRNA）作为生物标志物候选物的潜力可用于诊断，预后和治疗反应预测，因此，人们对能够快速，准确地检测这些分子的技术提出了巨大的需求。因此，最近在miRNA研究中已报告了许多成就。在这篇综述中，我们讨论了与miRNA检测新兴领域相关的挑战，这些挑战与miRNA的固有特性，当前可用技术的优缺点及其在临床研究中的潜在应用有关。我们总结了应用于体外的最有希望的核酸扩增技术检测miRNA，特别强调RT-qPCR的等温替代技术。我们详细介绍了这些方法的敏感性，特异性和定量性，以及它们在多路复用中的潜力。我们还将回顾这些化学方法已适应的不同检测格式，包括模拟读数，如实时监控，基于隔室中单分子扩增的数字计数以及基于表面的策略。