Description:
NFAT Responsive Luciferase Reporter NIH/3T3 Stable Cell Line is derived from mouse fibroblast, and stably express firefly luciferase reporter gene under the control of NFAT response element. This cell line is an ideal cellular model for monitoring the activation of Calcium Signaling Pathway triggered by stimuli treatment, enforced gene expression and gene knockdown.Principle:
NFATs are a family of transcriptional factors that play an important role in immune response as well as in the development of cardiac, skeletal muscle, and nervous systems. NFATs are regulated by calcium signaling. Through calcium activation of the phosphatase calcineurin, NFATc proteins translocate from the cytoplasm into the nucleus, where they cooperate with other proteins to mediate gene expression. Nuclear import of NFAT is blocked by kinases such as PKA and GSK3. NFATs are also implicated in breast cancer. Signosis has established a NFAT luciferase reporter cell line that has been stably transfected with a NFAT-luciferase reporter construct. Via the analysis of luciferase, the cell line can be employed to monitor the cellular changes of NFAT activities that are triggered by stimulation, compound treatment, enforced gene expression or gene knockdown.
The cell line is established by transfection using apTA-NFAT-luciferase reporter vector, which contains NFAT binding sites, a minimal promoter upstream of the firefly luciferase coding region,along with hygromycin expression vector followed by hygromycin selection. The hygromycin resistant clones were subsequently screened for PMA+ionomycin-induced luciferase activity.
Data:
Analysis of NFAT Luciferase Reporter NIH/3T3 Stable Cell Line.The cells were seeded on a 96-well plate for overnight with DMEM including 10% FBS. The cells then were treated with or without 10ng/ml PMA and 1uM ionomycin in DMEM and 0.1% FBS for 16 hours. More than 5 fold increase in luciferase activity was detected when compared to untreated cells.
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