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含盐酸胍的样本缓冲液结合生物安全性与RNA保存用于新冠病毒分子诊断

Sample Buffer Containing Guanidine-Hydrochloride Combines Biological Safety and RNA Preservation for SARS-CoV-2 Molecular Diagnostics.

作者信息

Weidner Lisa, Laner-Plamberger Sandra, Horner David, Pistorius Charlotte, Jurkin Jennifer, Karbiener Michael, Schistal Elisabeth, Kreil Thomas R, Jungbauer Christof

机构信息

Austrian Red Cross, Blood Service for Vienna, Lower Austria and Burgenland, Wiedner Hauptstraße 32, 1040 Vienna, Austria.

Department for Transfusion Medicine, University Hospital of Salzburg (SALK), Paracelsus Medical University (PMU), Müllner-Hauptstraße 48, 5020 Salzburg, Austria.

出版信息

Diagnostics (Basel). 2022 May 10;12(5):1186. doi: 10.3390/diagnostics12051186.

Abstract

The COVID-19 pandemic has elicited the need to analyse and store large amounts of infectious samples for laboratory diagnostics. Therefore, there has been a demand for sample storage buffers that effectively inactivate infectious viral particles while simultaneously preserving the viral RNA. Here, we present a storage buffer containing guanidine-hydrochloride that fulfils both requirements. Its ability to preserve RNA stability was confirmed by RT-qPCR, and virus-inactivating properties were tested by tissue culture infectious dose assay. Our data revealed that RNA from samples diluted in this storage buffer was efficiently preserved. Spiking samples with RNase A resulted in RNAse concentrations up to 100 ng/mL being efficiently inhibited, whereas spiking samples with infectious SARS-CoV-2 particles demonstrated rapid virus inactivation. In addition, our buffer demonstrated good compatibility with several commercially available RNA extraction platforms. The presented guanidine-hydrochloride-based storage buffer efficiently inactivates infectious SARS-CoV-2 particles and supports viral RNA stability, leading to a reduced infection risk during sample analysis and an increased period for follow-up analysis, such as sequencing for virus variants. Because the presented buffer is uncomplicated to manufacture and compatible with a variety of commercially available test systems, its application can support and improve SARS-CoV-2 laboratory diagnostics worldwide.

摘要

新冠疫情引发了对用于实验室诊断的大量感染性样本进行分析和储存的需求。因此,人们需要一种能有效灭活感染性病毒颗粒,同时保存病毒RNA的样本储存缓冲液。在此,我们展示了一种含盐酸胍的储存缓冲液,它能满足这两个要求。通过逆转录定量聚合酶链反应(RT-qPCR)证实了其保存RNA稳定性的能力,并通过组织培养感染剂量试验测试了其病毒灭活特性。我们的数据显示,用这种储存缓冲液稀释的样本中的RNA得到了有效保存。向样本中加入核糖核酸酶A(RNase A),高达100 ng/mL的核糖核酸酶浓度被有效抑制,而向样本中加入感染性严重急性呼吸综合征冠状病毒2(SARS-CoV-2)颗粒则显示出病毒的快速灭活。此外,我们的缓冲液与几种市售的RNA提取平台表现出良好的兼容性。所展示的基于盐酸胍的储存缓冲液能有效灭活感染性SARS-CoV-2颗粒,并维持病毒RNA的稳定性,从而降低样本分析期间的感染风险,并延长后续分析(如病毒变异体测序)的时间。由于所展示的缓冲液制备简单,且与多种市售测试系统兼容,其应用能够支持并改善全球范围内的SARS-CoV-2实验室诊断。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/81e1/9139951/4ee9b940e9f7/diagnostics-12-01186-g001.jpg

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