利用SIE转录活性荧光素酶报告基因系统检测白细胞介素6受体拮抗剂的生物活性

目的：利用sis诱导元素（sis-inducible element，SIE）转录活性荧光素酶报告基因系统检测白细胞介素6（IL-6）受体拮抗剂的生物活性。方法：采用脂质体转染法将含有SIE转录因子组件及荧光素酶报告基因的质粒pGL 4.47转入293T细胞，并通过潮霉素压力筛选获得稳定转染细胞株。加入IL-6激活稳转细胞株中的荧光素酶报告基因，构建SIE转录活性荧光素酶报告基因系统，并进一步建立基于此系统的IL-6受体拮抗剂药物的生物学活性定量检测方法。结果：IL-6可激活293T-SIE稳定转染细胞中的荧光素酶报告基因，且具有量效关系。对此系统检测IL-6受体拮抗剂药物的生物学活性进行方法学验证，显示本检测方法准确性及重复性均较好。结论：建立了一种基于荧光素酶报告基因系统的IL-6受体拮抗剂药物生物学活性检测方法，与现有方法相比，具有耗时少、准确性高、重复性好的优势。

Objective:To determine bioactivity of interleukin-6(IL-6) receptor antagonists using sis-inducible element(SIE) transcriptionally active luciferase reporter gene system.Methods:Plasmid pGL 4.47 containing the SIE transcription factor and the luciferase reporter gene was transferred into 293T cells by liposome transfection, and the stable transfected cell line(293T-SIE) was obtained by hygromycin pressure screening.Being activated by IL-6, the luciferase reporter gene system was constructed and the bioactivity detection method of IL-6 receptor antagonist based on this system was further established.Results:In 293T-SIE, the luciferase reporter gene could be activated by IL-6 with a dose-effect relationship.The methodological validation of this system for the detection of biological activity of IL-6 receptor antagonists showed that the method had good accuracy and reproducibity.Conclusion:A novel method was established to detect the bioactivity of IL-6 receptor antagonist based on the luciferase reporter gene system which had the advantages of being less time-consuming, having higher accuracy and better reproducibility compared with the existing methods.