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Validation of a HeLa Mx2/Luc reporter cell line for the quantification of human type i interferons

Title
Validation of a HeLa Mx2/Luc reporter cell line for the quantification of human type i interferons
Authors
Seo Y.-J.Kim G.-H.Kwak H.-J.Nam J.-S.Lee H.J.Suh S.-K.Baek K.-M.Sohn Y.-W.Hong S.-H.
Ewha Authors
이화정
SCOPUS Author ID
이화정scopus
Issue Date
2009
Journal Title
Pharmacology
ISSN
0031-7012JCR Link
Citation
vol. 84, no. 3, pp. 135 - 144
Indexed
SCI; SCIE; SCOPUS WOS scopus
Abstract
Although antiviral assays have been the most widely available biological assays for interferons (IFNs), they are less sensitive and provide considerable interassay variation. In this study, we demonstrate a new reporter cell line, which is based on HeLa cells transfected with a plasmid containing a human Mx2 promoter driving a luciferase (Luc) cDNA. To characterize the specific gene expression profiles induced by interferon alpha, we analyzed the microarray results of interferon response gene expression induced by IFN-α2a or IFN-α2b treatment with HeLa cells. We found that the Mx2 gene increased the most by treatment with both IFN-α2a and IFN-α2b. Based on this result, we designed a reporter cell line, HeLa-Mx2, suitable for determination of IFN-α. HeLa cells were stably transfected with the luciferase gene under the control of Mx2 promoter. The expression of luciferase can be easily measured for luminescence using a 96-well luminometer and has been correlated with the concentration of added IFN and cell density. In the validation results, our reporter cell line had specificity for type I IFN, but the significant effects of a number of other cytokines such as tumor necrosis factor-α, interleukin (IL)-1β, IL-2, IL-5, IL-6 and GM-CSF, or type II interferon (IFN-γ) were not observed. Moreover, the robustness of our cell line is demonstrated by the lack of an effect of the HeLa-Mx2 cell culture's age on the performance of the reporter gene assay. The reporter gene assay demonstrated reproducible dose-response curves for IFN-α2a in the range of 1-10,000 IU/ml. The 95% confidential limit and total coefficient of variation estimates ranged between 96 and 116 and 10.51% in the reducible range mentioned above, respectively. In conclusion, we established a stable IFN-responsible HeLa-Mx2 cell line, which has advantages with regard to simplicity, selectivity, and reliability over conventional cytopathic effect reduction assays used to quantify IFN-α activity. © 2009 S. Karger AG, Basel.
DOI
10.1159/000235158
Appears in Collections:
약학대학 > 약학과 > Journal papers
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