基于表达GFP的重组VSV建立安全，快速，方便，低成本的干扰素生物活性抗病毒测定方法。,Journal of Virological Methods

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基于表达GFP的重组VSV建立安全，快速，方便，低成本的干扰素生物活性抗病毒测定方法。
Journal of Virological Methods ( IF 2.2 ) Pub Date : 2017-08-24 , DOI: 10.1016/j.jviromet.2017.08.007
Weiye Chen ₁ , Zhiyuan Wen ₁ , Jialin Zhang ₁ , Cuicui Li ₁ , Kehe Huang ₂ , Zhigao Bu ₁

Affiliation

干扰素（I，II或III型）抗病毒活性的定量测定方法在进行研究期间非常重要，因为它们已经被发现。在这里，使用表达绿色荧光蛋白（GFP）（VSV / GFP）和MDBK细胞的重组水泡性口炎病毒来开发针对IFN的抗病毒测定（AVA）。该方法在96孔细胞培养板上进行，通过检测GFP量化了病毒复制的一半减少。为了定量GFP，将细胞裂解缓冲液直接添加到用VSV / GFP感染的孔中裂解细胞，然后使VSV / GFP失活，并测量GFP的相对荧光单位（RFU）并用于计算抗病毒活性。该方法仅需一个步骤，而无需用萘酚蓝黑染色方法中的三个步骤，可以使用酚红培养基，重现性好。可以将含GFP的样品在4°C的湿箱中保存至少1周，而不会影响测定结果。另外，用这种方法获得的结果与用染色方法获得的结果相似。总之，建立了基于重组VSV / GFP的IFN的安全，快速，方便和低成本的AVA。

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Establishing a safe, rapid, convenient and low-cost antiviral assay of interferon bioactivity based on recombinant VSV expressing GFP.

The methods of the quantitative assay of the antiviral activity of interferons (IFNs) (type I, II or III) are very important during carrying out of the research of them, since they were found. Here a recombinant vesicular stomatitis virus expressing green fluorescent protein (GFP) (VSV/GFP) and MDBK cells were used to develop an antiviral assay (AVA) for IFNs. This method was carried out on a 96-well cell culture plate, and the half reduction of virus replication was quantified by assaying GFP. To quantify GFP, cell lysis buffer was directly added to the wells infected with VSV/GFP to lyse cells, the VSV/GFP was then inactivated, and relative fluorescence unit (RFU) of GFP was measured and used to calculate the antiviral activity. This method needed only one step instead of three steps in the staining method with naphthol blue black, medium with phenol red can be used, and it had good reproducibility. The GFP-containing samples could be stored at 4°C in a wet box for at least 1 week without affecting the assay results. In addition, the results obtained with this method were similar to those obtained with the staining method. In conclusion, a safe, rapid, convenient and low-cost AVA of IFN based on recombinant VSV/GFP was established.

更新日期：2019-11-01

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