IFN-α/β Reporter HEK 293 Cells

Notification:  A new clone is provided with an improved Type I IFN response. The cat code has been changed accordingly (hkb-ifnabv2).
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HEK-Blue™ IFN-α/β Cells signaling pathway

Human Type I IFNs Reporter Cells

HEK-Blue™ IFN-α/β cells were engineered from the human embryonic kidney HEK 293 cell line to detect bioactive human type I interferons (e.g. IFN-α, IFN-β) by monitoring the activation of the ISGF3 pathway.
IFN-α and IFN-β are important anti-viral cytokines that also have anti-proliferative and immunomodulatory functions [1, 2]. They bind a cell-surface receptor, composed of two subunits, IFNAR1 and IFNAR2, which are associated with TyK2 and JAK1, respectively [1].

 More details

Description:

HEK-Blue™ IFN-α/β cells were generated by stable expression of the genes encoding the human STAT2, IRF9, and a STAT1/2-inducible secreted embryonic alkaline phosphatase (SEAP) reporter 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. STAT1/2-dependent SEAP activity is readily assessable in the supernatant using QUANTI-Blue™ Solution, a detection reagent. The activation of HEK-Blue™ IFN-α/β cells can be blocked with a neutralizing monoclonal antibody, such as anti-hIFN-α-IgG. Of note, HEK-Blue™ IFN-α/β cells do not respond to human type II and type III IFNs (IFN-γ/λ; see figures).
 

Key Features:

• Fully functional IFN-α/β signaling pathway
• Strong response to human IFN-α and IFN-β 
• Poor response to murine (m) IFN-α
• Unresponsive to  IFN-γ (type II IFN) and IFN-λ (type III IFN)
• Readily assessable SEAP reporter activity

Applications:

• Detection of human IFN-α and IFN-β 
• Screening of anti-IFN-α/β and anti-IFNAR antibodies

References:

1. Schreiber G. 2017. The molecular basis for differential type I interferon signaling. J. Biol. Chem. 292:7285-94.
2. McNab F. et al., 2015. Type I interferons in infectious disease. Nat Rev Immunol. 15(2):87-103.

Figures

Dose-response of HEK-Blue™ IFN-α/β cells to recombinant type I IFNs. Cells were stimulated with increasing concentrations of recombinant human (h)IFN-α and hIFN-b . After overnight incubation, the ISGF3 response was determined using QUANTI‑Blue™ Solution, a SEAP detection reagent. The optical density (OD) at 630 nm is shown as mean ± SEM.

Response of HEK-Blue™ IFN-α/β cells to a panel of cytokines. Cells were stimulated with various human recombinant cytokines: 100 U/ml hIFN-α2b or hIFN-b-1a, 10000 U/ml mIFN-α or mIFN-b, 100 ng/ml IFN-γ, 100 ng/ml IL-28a, IL-28b, or IL-29. After overnight incubation, SEAP activity was assessed using QUANTI‑Blue™ Solution. The optical density (OD) at 630 nm is shown as mean ± SEM.

Dose-dependent inhibition of HEK-Blue™ IFN-α/β cell response using Anti-IFN-α-IgG. A serial dilution of Anti-IFN-α-IgG monoclonal antibody (mAb) was incubated with 500 U/ml of recombinant human IFN-α2b for 30 minutes prior to the addition of the HEK-Blue™ IFN-α/β cells. After overnight incubation, the ISGF3 response was determined using QUANTI‑Blue™ Solution, a SEAP detection reagent. Data are presented as percentage of neutralization (mean ± SEM).

Specifications

Antibiotic resistance: Blasticidin, Zeocin®

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

Guaranteed mycoplasma-free

Detection ranges:

• 1 - 10³ U/ml for human IFN-α
• 1 - 10³ U/ml for human IFN-β

This product is covered by a Limited Use License (See Terms and Conditions).

Contents

• 1 vial containing 3-7 x 10⁶ cells
• 1 ml of Blasticidin (10 mg/ml)
• 1 ml of Zeocin® (100 mg/ml)
• 1 ml Normocin™ (50 mg/ml)
• 1 ml of QB reagent and 1 ml of QB buffer (sufficient to prepare 100 ml of QUANTI-Blue™ Solution, a SEAP detection reagent)

Shipped on dry ice (Europe, USA, Canada and some areas in Asia)

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 [1, 2]. The type I IFN family is a multi-gene cytokine family that encodes 13 partially homologous IFN-α subtypes in humans (14 in mice), a single IFN-β, and several poorly defined single gene products (IFN-ɛ, IFN-τ, IFN-κ, IFN-ω, IFN-δ, and IFN-ζ) [1, 2].  IFN-α and IFN-β are the best-defined and most broadly expressed type I IFNs [2].

IFN-β and all of the IFN-α subtypes bind to a heterodimeric transmembrane receptor composed of the subunits IFNAR1 and IFNAR2 which are associated with the tyrosine kinases Tyk2 and Jak1 (Janus kinase 1) respectively. These kinases phosphorylate STAT1 and STAT2 which then dimerize and interact with IFN regulatory factor 9 (IRF9), leading to the formation of the ISGF3 complex. ISGF3 binds to IFN-stimulated response elements (ISRE) in the promoters of IFN-stimulated genes (ISG) to regulate their expression. 

Despite their protective effects, studies have shown that aberrantly expression of the type I IFN system can elicit autoimmune disorders, such as interferonopathies and SLE (systemic lupus erythematosus). Recent evidence also implicates type I IFN-dependent signaling as a key inflammatory driver in non-autoimmune diseases [3].

References:

1. Schreiber G. 2017. The molecular basis for differential type I interferon signaling. J. Biol. Chem. 292:7285-94.
2. McNab F. et al., 2015. Type I interferons in infectious disease. Nat Rev Immunol. 15(2):87-103.
3. Crow MK, Ronnblom L. Type I interferons in host defence and inflammatory diseases. Lupus Sci Med. 2019 May 28;6(1):e000336.

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