HEK-Blue™ IFN-α/β Cells

Human Type I IFN Sensor Cells

HEK-Blue™ IFN-α/β cells allow the detection of bioactive human type I IFNs 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 in sufficient amounts.

The cells were further transfected with a SEAP 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 in the supernatant can be easily determined with QUANTI-Blue™.

Stimulation of HEK-Blue™ IFN-α/β cells with human IFN-α can be blocked by anti-hIFN-α-IgA, a neutralizing monoclonal antibody.

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)

Description

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.

Details