Plotnikova Marina A, Klotchenko Sergey A, Vasin Andrey V
Research Institute of Influenza, Saint-Petersburg, Russian Federation.
Research Institute of Influenza, Saint-Petersburg, Russian Federation.
J Immunol Methods. 2016 Mar;430:51-5. doi: 10.1016/j.jim.2016.01.005. Epub 2016 Jan 7.
Cytokines are global mediators of cellular communications that are involved in broad array of biological processes, including the immunological and inflammatory mechanisms of virus-host interactions. Measuring the gene expression of simultaneously expressed cytokines is necessary for understanding the pathogenesis of many viral infections, including influenza. We developed a multiplex quantitative real-time PCR (qPCR) method for the detection of the following human cytokines: IL-1B, IL-2, IL-4, IL-6, IL-10, IL-12B, IL-18, IFN-γ and TNF. The assay consisted of three sets of multiple qPCRs; in each qPCR, three target cytokines and reference GAPDH genes were amplified. The assay provided a precise and sensitive quantification of cytokine gene expression with a 20fmol limit of detection and a 1.5% coefficient of variation. This method was successfully applied to cytokine profiling in epithelial A549 cells that were infected with A/California/07/09 (H1N1pdm2009) virus.
细胞因子是细胞通讯的全局介质，参与广泛的生物过程，包括病毒与宿主相互作用的免疫和炎症机制。测量同时表达的细胞因子的基因表达对于理解包括流感在内的许多病毒感染的发病机制是必要的。我们开发了一种多重定量实时PCR（qPCR）方法，用于检测以下人类细胞因子：IL-1B、IL-2、IL-4、IL-6、IL-10、IL-12B、IL-18、IFN-γ和TNF。该检测方法由三组多重qPCR组成；在每个qPCR中，扩增三个目标细胞因子和参考GAPDH基因。该检测方法对细胞因子基因表达进行了精确且灵敏的定量，检测限为20fmol，变异系数为1.5%。该方法成功应用于感染A/California/07/09（H1N1pdm2009）病毒的上皮A549细胞中的细胞因子谱分析。
