HEK-Blue™ ISG Cells
|HEK-Blue™ ISG Cells||Unit size||Cat. code||Docs||Qty||Price|
Interferon Regulatory Factor (IRF)-Inducible SEAP Reporter HEK293 Cells
3-7 x 10e6 cells
Interferon regulatory factor (IRF)-inducible SEAP reporter HEK293 cells
HEK-Blue™ ISG cells were specifically designed to study the activation of the STING/TBK1/IRF3 signaling pathway by CDNs.
HEK-Blue™ ISG cells were derived from the PEAKrapid cell line (similar to ATCC® CRL-2828™) which itself was derived from the HEK293 cell line.
HEK-Blue™ ISG cells express a secreted embryonic alkaline phosphatase (SEAP) under the control of the IRF-inducible promoter comprised of five IFN-stimulated response elements (ISRE) fused to an ISG54 minimal promoter.
CDNs in the cytosol bind directly to STING leading to TBK1-mediated IRF3 activation and type I IFN production. IFNs activate the JAK-STAT pathway with the subsequent activation of IFN-stimulated response elements (ISRE) in the promoters of IFN-stimulated genes (ISG). Hence, the presence of CDNs in the cytosol of HEK-Blue™ ISG cells will induce the production of the IRF-inducible SEAP reporter directly by activating the STING/TBK1/IRF3 pathway and indirectly through the activation of the JAK/STAT/IRF9 pathway with type I IFN.
Levels of SEAP in the supernatant can be easily determined with QUANTI- Blue™, a reagent that turns purple/blue in the presence of SEAP and by reading the OD at 620-655 nm.
HEK-Blue™ ISG cells respond strongly to non-canonical CDNs, namely 2’3’-cGAMP and 2’2’-cGAMP but poorly to cytosolic DNA, DMXAA and canonical cyclic dinucleotides (CDNs).
Stimulation of HEK-Blue™ ISG-KO-STING and HEK-Blue™ ISG cells (wild-type cell line) with poly(dA:dT)/LyoVec™ (1 µg/ml), 2’3’-cGAMP (1 µg/ml), 3’3’-cGAMP (10 µg/ml) and 2’3’-c-di-AMP (10 µg/ml). Human IFN-α (1x103 U/ml) and IFN-β (1x103 U/ml) serve as positive controls. After 24h incubation, IRF activation was determined using QUANTI-Blue™, a SEAP detection reagent, and by reading the optical density (OD) at 655 nm. The IRF induction of each ligand is expressed relative to that of mIFN-β at 1x103 U/ml (taken as 100%).
Growth Medium: DMEM, 4.5 g/l glucose, 10% (v/v) fetal bovine serum, 50 U/ml penicillin, 50 µg/ml streptomycin, 100 µg/ml Normocin™, 2 mM L-glutamine supplemented with Zeocin™ selective antibiotic only
- Reporter activity is validated upon stimulation with IFN-α or IFN-β and IRF3 activators such as 2’2’-cGAMP.
- These cells are guaranteed mycoplasma-free.
- 1 vial of HEK-Blue™ ISG Cells (3-7 x 106 cells)
- 100 μl Zeocin™ (100 mg/ml).
- 1 ml Normocin™ (50 mg/ml).
- 1 pouch of QUANTI-Blue™ (SEAP detection medium).
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