Beijing da xue xue bao. Yi xue ban = Journal of Peking University. Health sciences · 2006
[Application of NF-kappaB reporter and Dual-Luciferase assays in the measure of bioactivity of interleukin-1beta and interleukin-1 receptor antagonist].
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Zhang Ying-mei, Wang Ying, Xu Qian-mei, Lv Bing-feng, Ma Da-long
Аннотация
OBJECTIVE: To develop a reporter gene system based on transient transfections with a NF-kappaB responsive reporter gene to detect the bioactivity of IL-1beta and IL-1 receptor antagonist.
METHODS: NF-kappaB reporter and Dual-Luciferase assays were applied to measure the bioactivity of IL-1beta and IL-1 receptor antagonist in mouse EL4 cells (some subclones of EL4 cells expressed high level of IL-1 receptor on cell surface). pNF-kappaB-luc and pRL-TK, used as an internal control, were co-transfected into EL4 cells and then the IL-1beta was added.
RESULTS: The results indicated that IL-1beta was able to induce the expression of this luciferase, which could be blocked by IL-1 receptor antagonist. The optimal dose of IL-1beta was 5 microg/L in Dual-Luciferase assay, whose bioactivity can be effectively inhibited by IL-1ra at 50 microg/L.
CONCLUSION: We have established a new method to detect the bioactivity of IL-1beta and IL-1 receptor antagonist, which can give repeatable results.
METHODS: NF-kappaB reporter and Dual-Luciferase assays were applied to measure the bioactivity of IL-1beta and IL-1 receptor antagonist in mouse EL4 cells (some subclones of EL4 cells expressed high level of IL-1 receptor on cell surface). pNF-kappaB-luc and pRL-TK, used as an internal control, were co-transfected into EL4 cells and then the IL-1beta was added.
RESULTS: The results indicated that IL-1beta was able to induce the expression of this luciferase, which could be blocked by IL-1 receptor antagonist. The optimal dose of IL-1beta was 5 microg/L in Dual-Luciferase assay, whose bioactivity can be effectively inhibited by IL-1ra at 50 microg/L.
CONCLUSION: We have established a new method to detect the bioactivity of IL-1beta and IL-1 receptor antagonist, which can give repeatable results.