Objective: To establish a reporter gene assay (RGA) for detecting the biological activity of recombinant human interleukin-1 receptor antagonists(IL-1Ra). Methods: D10G4.1-NFκB-Luc cells were stimulated by IL-β to produce fluorescence, the IL-1Ra inhibited this reaction, and the effect of inhibition had a linear relationship with its dosage to determine its biological activity. The method was established after exploring cell density, the amount of irritant IL-1β, the amount of antagonist IL-1Ra, dilution gradient and incubation time, and the accuracy, linear range, specificity and durability of the method were investigated. The established method was compared with the A375.S2 cell viability method. Results: The biological activity of IL-1Ra was successfully established by RGA, and the relative deviation of the detection results in the range of 50%-160% in the titer level range was within the range of ±12%, the slope of the regression equation was 1.001 3, the correlation coefficient was 0.9998, and the geometric variation (GCV) of durability for different cell generations, cell densities and culture time was 4.0%. The method took a short time, the detection results were consistent with the A375.S2 cell viability method, and the precision and accuracy were significantly improved. Conclusion: The reporter gene assay has been successfully established to detect the biological activity of IL-1Ra, and has been validated, which can replace the A375.S2 cell activity method for product quality control.
YU Lu, LIU Yu-lin, ZHENG Yi-han, PENG Bing-huai, LIU Jing-hui
. Study on the biological activity of IL-1Ra by reporter gene assay[J]. Chinese Journal of Pharmaceutical Analysis, 2023
, 43(12)
: 2025
-2029
.
DOI: 10.16155/j.0254-1793.2023.12.05
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