ESPCI PSL, CBI, IPGG, Paris, France.
Institut Pasteur, Paris Cedex, France.
PLoS One. 2021 Jan 11;16(1):e0243712. doi: 10.1371/journal.pone.0243712. eCollection 2021.
To respond to the urgent need for COVID-19 testing, countries perform nucleic acid amplification tests (NAAT) for the detection of SARS-CoV-2 in centralized laboratories. Real-time RT-PCR (Reverse transcription-Polymerase Chain Reaction), used to amplify and detect the viral RNA., is considered, as the current gold standard for diagnostics. It is an efficient process, but the complex engineering required for automated RNA extraction and temperature cycling makes it incompatible for use in point of care settings [1]. In the present work, by harnessing progress made in the past two decades in isothermal amplification and paper microfluidics, we created a portable test, in which SARS-CoV-2 RNA is extracted, amplified isothermally by RT-LAMP (Loop-mediated Isothermal Amplification), and detected using intercalating dyes or fluorescent probes. Depending on the viral load in the tested samples, the detection takes between twenty minutes and one hour. Using a set of 16 pools of naso-pharyngal swab eluates, we estimated a limit of detection comparable to real-time RT-PCR (i.e. 1 genome copies per microliter of clinical sample) and no cross-reaction with eight major respiratory viruses currently circulating in Europe. We designed and fabricated an easy-to-use portable device called "COVIDISC" to carry out the test at the point of care. The low cost of the materials along with the absence of complex equipment will expedite the widespread dissemination of this device. What is proposed here is a new efficient tool to help managing the pandemics.
为了满足对 COVID-19 检测的紧急需求，各国在集中实验室中进行核酸扩增检测（NAAT）以检测 SARS-CoV-2。实时 RT-PCR（逆转录-聚合酶链反应）用于扩增和检测病毒 RNA，被认为是目前诊断的金标准。它是一种高效的过程，但自动化 RNA 提取和温度循环所需的复杂工程使其不兼容用于即时护理环境[1]。在本工作中，我们利用过去二十年来在等温扩增和纸微流控方面取得的进展，创建了一种便携式测试，其中 SARS-CoV-2 RNA 通过 RT-LAMP（环介导等温扩增）进行等温提取和扩增，并使用嵌入染料或荧光探针进行检测。根据测试样本中的病毒载量，检测时间在二十分钟到一个小时之间。使用一组 16 个鼻咽拭子洗脱液池，我们估计检测限与实时 RT-PCR 相当（即每微升临床样本 1 个基因组拷贝），并且与目前在欧洲流行的八种主要呼吸道病毒没有交叉反应。我们设计并制造了一种名为“COVIDISC”的易于使用的便携式设备，可在即时护理点进行测试。材料成本低，且无需复杂设备，这将加速该设备的广泛传播。这里提出的是一种新的高效工具，有助于管理大流行。
