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Author:
Date:
2020-12-20
[Abstract] The mRNA therapeutics is a new class of medicine to treat many various diseases. However, in vitro transcribed (IVT) mRNA triggers immune responses due to recognition by human endosomal and cytoplasmic RNA sensors, but incorporation of modified nucleosides have been shown to reduce such responses. Therefore, an assay signifying important aspects of the human immune system is still required. Here, we present a simple ex vivo method called ‘RNA ImmunoGenic Assay’ to measure immunogenicity of IVT-mRNAs in human whole blood. Chemically modified and unmodified mRNA are complexed with a transfection reagent (TransIT), and co-incubated in human whole blood. Specific cytokines are measured (TNF-α, INF-α, INF-γ, IL-6 and IL-12p70) using ELISAs. The ...
[摘要] [摘要] mRNA疗法是治疗多种疾病的新型药物。然而，我Ñ体外转录（IVT）的mRNA触发由于人类内体和细胞质RNA传感器，但是修饰的核苷的掺入识别的免疫应答已经示出吨ö减少此类反应。牛逼herefore ，测定标志着重要的环节人体免疫系统仍然需要。这里，我们提出一个简单的离体称为“RNA方法免疫ģ ENIC测定”测量的IVT-mRNA的免疫原性小号在人全血。将化学修饰和未修饰的mRNA与转染试剂（TransIT ）复合，并在人全血中共同孵育。特异性细胞因子测定（TNF- α ，INF- α ，INF- γ ，IL-6和IL-12p70的），使用的ELISA。进行qPCR分析以揭示特异性免疫途径的激活。所述RNA免疫ģ ENIC测定提供小号的简单且快速的方法来检测供体特异性-针对mRNA的治疗剂的免疫应答。
图形摘要：
RNA免疫基因测定的示意图
[背景] mRNA治疗是基因治疗的重要一类（Sahin等，2014 ；Antony等，2015 ...
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Author:
Date:
2020-11-05
[Abstract] Standard diagnostic methods of Coronavirus Disease 2019 (COVID-19) rely on RT-qPCR technique which have limited point-of-care test (POCT) potential due to necessity of dedicated equipment and specialized personnel. LAMP, an isothermal nucleic acid amplification test (NAAT), is a promising technique that may substitute RT-qPCR for POCT of genomic materials. Here, we provide a protocol to perform reverse transcription LAMP targeting SARS-CoV-2. We adopted both real-time fluorescence detection and end-point colorimetric detection approaches. Our protocol would be useful for screening diagnosis of COVID-19 and be a baseline for development of improved POCT NAAT.
[摘要] [摘要] 2019年冠状病毒病的标准诊断方法（COVID-19）依靠RT-qPCR技术，由于需要专用设备和专门人员，因此具有有限的即时检验（POCT）潜力。LAMP是一种等温核酸扩增测试（NAAT），是一种有前途的技术，可以用RT-qPCR代替基因组材料的POCT。在这里，我们提供了一种执行针对SARS-CoV-2的逆转录LAMP的协议。我们同时采用了实时荧光检测和终点比色检测方法。我们的协议对于筛查COVID-19的诊断将是有用的，并且是开发改进的POCT NAAT的基线。
[背景]快速和SARS-CoV的-2，COVID-19的病原的灵敏检测，重要的是要控制电流大流行的情况，因为它使得早期检测，隔离和治疗以及监测筛选。按照世界卫生组织的建议，当前用于检测SARS-CoV-2的标准方法采用RT-qPCR （世界卫生组织，2020年）。然而，RT-qPCR诊断的正确执行需要高水平的设施和专家，通常在医院/采样场所无法获得，因此缺乏POCT适用性。
由于等温扩增方法可以用相对简单的仪器进行检测，因此开发了等温扩增方法以通过在各种现场即时检测核酸来完成诊断（Niemz et ...
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