[Application of NF-kappaB reporter and Dual-Luciferase assays in the measure of bioactivity of interleukin-1beta and interleukin-1 receptor antagonist].
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.


CONCLUSIONS
We have established a new method to detect the bioactivity of IL-1beta and IL-1 receptor antagonist, which can give repeatable results.
Zhang
Ying-mei
YM
Department of Immunology, Peking University School of Basic Medical Science, Peking University, Beijing 100083, China.
Wang
Ying
Y
Xu
Qian-mei
QM
Lv
Bing-feng
BF
Ma
Da-long
DL
D000818:Animals; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D053590:Interleukin 1 Receptor Antagonist Protein / Q000494:pharmacology*; D053583:Interleukin-1beta / Q000494:pharmacology*; D008156:Luciferases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D017472:Receptors, Interleukin-1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D013945:Thymoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D013953:Thymus Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D014162:Transfection
D053590:Interleukin 1 Receptor Antagonist Protein; D053583:Interleukin-1beta; D016328:NF-kappa B; D017472:Receptors, Interleukin-1; D011993:Recombinant Fusion Proteins; D008156:Luciferases