[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.

CONCLUSION

We have established a new method to detect the bioactivity of IL-1beta and IL-1 receptor antagonist, which can give repeatable results.