HEK-Blue™ IFN-α/β Cells

HEK-Blue™ IFN-α/β Cells

Human Type I IFN Reporter Cells

HEK-Blue™ IFN-α/β cells allow the detection of bioactive human type I interferons (i.e. IFN-α and IFN-β) by monitoring the activation of the ISGF3 pathway.

These cells were generated by stable transfection of HEK293 cells with the human STAT2 and IRF9 genes to obtain a fully active type I IFN signaling pathway. The other genes of the pathway (IFNAR1, IFNAR2, JAK1, TyK2, and STAT1) are naturally expressed by these cells. The cells feature an inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene under the control of the IFN-α/β inducible ISG54 promoter. 

Stimulation of HEK-Blue™ IFN-α/β cells with human IFN-α or IFN-β activates the JAK/STAT/ISGF3 pathway and subsequently induces the production of SEAP. Levels of SEAP are readily assessable in the supernatant using QUANTI-Blue™.

Additionally, the activation of HEK-Blue™ IFN-α/β cells with human IFN-α can be blocked with a neutralizing monoclonal antibody, such as anti-hIFN-α-IgA.

Figures

Stimulation of HEK-Blue™ IFN-α/β cells by recombinant human IFN-α, IFN-β and IFN-γ. After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the optical density (O.D.) at 655 nm.

Ligand	EC50	Response Ratio
hIFN-α	75 +/- 10 IU/ml	20
hIFN-β	10 +/- 2 IU/ml	20

Note: The response ratio was calculated by dividing the OD at 655 nm for the treated cells by the OD at 655 nm for the untreated cells.

HEK-Blue™ IFN-α/β cells were stimulated with various human recombinant cytokines; IFNα (1000 IU/ml), IFNβ (1000 IU/ml), IFNγ (1000 IU/ml), IL-1β (100 ng/ml), IL-4 (100 ng/ml), IL-6 (100 ng/ml), IL-13 (100 ng/ml), IL-18 (100 ng/ml), TGF-β (10 ng/ml), and TNF-α (100 ng/ml). After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the O.D. at 655 nm.

Anti-hIFN-α-IgA was incubated with 500 IU/ml of human IFN-α for 30 minutes prior to the addition of HEK-Blue™ IFN-α/β cells, which were incubated with the antibody and cytokine for a further 24 h. Levels of SEAP in the supernatant were measured using QUANTI-Blue™ and reading the O.D. at 655 nm.

Specifications

Antibiotic resistance: blasticidin, Zeocin™

Growth medium: DMEM, 4.5 g/l glucose, 2-4 mM L-glutamine, 10% (v/v) fetal bovine serum, 50 U/ml penicillin, 50 μg/ml streptomycin, 100 μg/ml Normocin™

Guaranteed mycoplasma-free

Detects human type I interferons:

• Detection range for human IFN-α: 1 - 10⁴ IU/ml
• Detection range for human IFN-β: 10 - 10⁴ IU/ml

Contents

• 1 vial containing 3-7 x 10⁶ cells
• 100 μl Blasticidin (10 mg/ml)
• 100 μl Zeocin™ (100 mg/ml)
• 1 ml Normocin™ (50 mg/ml)
• 1 pouch of QUANTI-Blue™ (SEAP detection medium)

Shipped on dry ice (Europe, USA & Canada)

Details

Type I interferons, in particular interferon alpha (IFN-α) and interferon beta (IFN-β), play a vital role in host resistance to viral infections.

They signal mainly through the JAK-STAT pathway. Following their production, IFN-α and IFN-β bind to a common receptor (IFNAR) and recruit the Janus kinases (JAK1 and TyK2).

JAKs phosphorylate STAT1 and STAT2, which then dimerize and interact with IFN regulatory factor 9 (IRF9), forming a complex named ISGF3.

ISGF3 binds to IFN-stimulated response elements (ISRE) in the promoters of IFN-stimulated genes (ISG) to regulate their expression.

Stimulation of HEK-Blue™ IFN-α/β cells with human IFN-α or IFN-β activates the JAK/STAT/ISGF3 pathway and subsequently induces the production of SEAP.