Borrego Belén, Martinez-Sobrido Luis, Brun Alejandro, Nogales Aitor
Animal Health Research Centre (CISA), Centro Nacional Instituto de Investigación y Tecnología Agraria y Alimentaria (INIA, CSIC), Madrid, Spain.
Texas Biomedical Research Institute, San Antonio, TX, USA.
Methods Mol Biol. 2025;2893:11-23. doi: 10.1007/978-1-0716-4338-9_2.
Rift Valley fever (RVF) is an arboviral zoonotic disease affecting many African countries with the potential to spread to other geographical areas. In this chapter we describe the use of a replication-competent recombinant (r)RVFV expressing NanoLuc Luciferase (Nluc) for in vitro studies. The determination of parameters such as neutralizing antibodies in serum samples, or the antiviral activity of drugs is usually carried out using standard assays based on the assessment of cytopathic effect on cell cultures. The use of a virus encoding a traceable reporter protein allows to correlate the presence or absence of infection with the detection of the product in the infected cultures, thus tracking the level of RVFV infection in an objective, quantitative manner. In addition to this quantitative measurement of results, our protocol offers two other advantages, such as a shorter time to read, given that 48 h post-infection the production of the reporter protein is enough to give an accurate result, and the use of an attenuated virus, which reduces the risk of exposure.
裂谷热（RVF）是一种虫媒病毒人畜共患病，影响许多非洲国家，并有可能传播到其他地理区域。在本章中，我们描述了使用表达纳米荧光素酶（Nluc）的具有复制能力的重组（r）RVFV进行体外研究。血清样本中中和抗体等参数的测定，或药物的抗病毒活性，通常使用基于评估细胞培养中细胞病变效应的标准检测方法来进行。使用编码可追踪报告蛋白的病毒能够将感染的存在与否与感染培养物中产物的检测相关联，从而以客观、定量的方式追踪RVFV感染水平。除了对结果进行这种定量测量外，我们的方案还有另外两个优点，比如读取时间更短，因为感染后48小时报告蛋白的产生就足以给出准确结果，以及使用减毒病毒，这降低了暴露风险。
