Quantification of virus-infected cells using RNA FISH-Flow.

Quantification of virus-infected cells using RNA FISH-Flow.
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DOI:
10.1016/j.xpro.2023.102291
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发表时间:
2023-05-18
期刊:
影响因子:
--
通讯作者:
Sawyer, Sara L.
Sawyer, Sara L.
中科院分区:
其他
文献类型:
--
作者:
Warren, Cody J.;Barbachano-Guerrero, Arturo;Huey, Devra;Yang, Qing;Worden-Sapper, Emma R.;Kuhn, Jens H.;Sawyer, Sara L.

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我们提出了一个协议来检测已被RNA病毒感染的细胞。该方法,RNA荧光原位杂交流式细胞术(RNA FISH-Flow),使用48个荧光标记的DNA探针,串联杂交病毒RNA。RNA FISH-Flow探针可以合成以匹配任何RNA病毒基因组,无论是正义还是反义,从而能够检测细胞内的基因组或复制中间体。流式细胞术能够在单细胞水平上对群体内的感染动力学进行高通量分析。有关本方案使用和执行的完整详细信息,请参见Warren等人。(2022年)。病毒特异性RNA FISH-流式探针生成的分步程序RNA FISH-流式探针与病毒暴露细胞杂交的详细程序通过流式细胞术分析病毒RNA阳性细胞出版商说明:进行任何实验方案都需要遵守当地机构的实验室安全和伦理指南。我们提出了一个协议来检测已被RNA病毒感染的细胞。该方法,RNA荧光原位杂交流式细胞术(RNA FISH-Flow),使用48个荧光标记的DNA探针,串联杂交病毒RNA。RNA FISH-Flow探针可以合成以匹配任何RNA病毒基因组,无论是正义还是反义,从而能够检测细胞内的基因组或复制中间体。流式细胞术能够在单细胞水平上对群体内的感染动力学进行高通量分析。
We present a protocol to detect cells that have been infected by RNA viruses. The method, RNA fluorescence in situ hybridization flow cytometry (RNA FISH-Flow), uses 48 fluorescently labeled DNA probes that hybridize in tandem to viral RNA. RNA FISH-Flow probes can be synthesized to match any RNA virus genome, in either sense or anti-sense, enabling detection of genomes or replication intermediates within cells. Flow cytometry enables high-throughput analysis of infection dynamics within a population at the single cell level. For complete details on the use and execution of this protocol, please refer to Warren et al. (2022). Step-by-step procedure for virus-specific RNA FISH-Flow probe generation Detailed procedure for RNA FISH-Flow probe hybridization to virus-exposed cells Analysis of viral RNA-positive cells by flow cytometry Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. We present a protocol to detect cells that have been infected by RNA viruses. The method, RNA fluorescence in situ hybridization flow cytometry (RNA FISH-Flow), uses 48 fluorescently labeled DNA probes that hybridize in tandem to viral RNA. RNA FISH-Flow probes can be synthesized to match any RNA virus genome, in either sense or anti-sense, enabling detection of genomes or replication intermediates within cells. Flow cytometry enables high-throughput analysis of infection dynamics within a population at the single cell level.
DOI: 10.1016/j.cell.2022.09.022
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