IFN-γ Reporter HEK 293 Cells

Human Interferon-γ Reporter Cells

HEK-Blue™ IFN-γ Cells signaling pathway

HEK-Blue™ IFN-γ cells allow the detection of bioactive human interferon-γ (IFN-γ) by monitoring the activation of the JAK/STAT-1 pathway. 
IFN-γ, a Type II IFN, is a pleiotropic cytokine with anti-viral, anti-tumor, and immunomodulatory functions [1].  IFN-γ binds a heterodimeric receptor consisting of two subunits, IFNGR1 and IFNGR2, associated with JAK1 and JAK2, respectively. Upon binding to this receptor, IFN-γ triggers JAK/STAT signaling.  Activated STAT1 molecules form homodimers and are translocated to the nucleus where they bind interferon-gamma-activated sites (GAS) in the promoter of IFN-γ inducible genes.

 More details on interferon-γ

Cell line description:

HEK-Blue™ IFN-γ cells were generated by stable transfection of the human embryonic kidney HEK293 cell line with the human STAT1 gene to obtain a fully active STAT1 pathway.

The other genes of the pathway are naturally expressed in sufficient amounts.

These cells were further transfected with a SEAP (secreted embryonic alkaline phosphatase) reporter gene under the control of an ISG54 promoter fused to four interferon-gamma-activated sites (GAS).

HEK-Blue™ IFN-γ cells produce SEAP in response to IFN-γ stimulation only. They are unresponsive to type I IFNs.

Levels of SEAP in the supernatant can be easily determined with QUANTI-Blue™ Solution.

Features of HEK-Blue™ IFN-γ cells:

• Fully functional IFN-γ signaling pathway
• Do not respond to IFN-α/β (Type I IFNs)
• Readily assessable SEAP reporter activity
• Functionally tested and guaranteed mycoplasma-free

Applications of HEK-Blue™ IFN-γ cells:

• Detection of human IFN-γ
• Screening of anti-hIFN-γ antibodies

Reference:

1. Ivashkiv L.B., 2018. IFNγ: signalling, epigenetics and roles in immunity, metabolism, disease and cancer immunotherapy. Nat Rev Immunol. 18(9):545-558.

Figures

Cells were stimulated with increasing concentrations of hIFN-α (IU/ml), hIFN-β (IU/ml), or hIFN-γ (ng/ml). After a 24h incubation, the levels of IFN-induced SEAP were determined using QUANTI-Blue™, a SEAP detection reagent, and by reading the optical density (OD) at 655 nm.

Ligand	EC50	Response Ratio
Human IFN-γ	0.7 ng/ml	25

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.

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 range for human IFN-γ: 0.1 - 10 ng/ml

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 of 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

Interferon-gamma (IFN-γ), a Type II interferon, is secreted from CD4+ T-helper 1 (Th1) cells and activated natural killer (NK) cells. It plays a role in activating lymphocytes to enhance anti-microbial and anti-tumor effects [1-3]. In addition, IFN-γ plays a role in regulating the proliferation, differentiation, and response of lymphocyte subsets. 

IFN-γ exerts its action by first binding to a heterodimeric receptor consisting of two chains, IFNGR1 and IFNGR2, causing its dimerization and the activation of specific Janus family kinases (JAK1 and JAK2) [4, 5]. Two STAT1 molecules then associate with this ligand-activated receptor complex and are activated by phosphorylation. Activated STAT1 forms homodimers and are translocated to the nucleus where they bind interferon-gamma-activated sites (GAS) in the promoter of IFN-γ inducible genes.

1. Ivashkiv L.B., 2018. IFNγ: signalling, epigenetics and roles in immunity, metabolism, disease and cancer immunotherapy. Nat Rev Immunol. 18(9):545-558.
2. Shtrichman R. & Samuel CE., 2001. The role of gamma interferon in antimicrobial immunity. Curr Opin Microbiol. 4(3):251-9.
3. Sato A. et al., 2006. Antitumor activity of IFN-lambda in murine tumor models. J Immunol. 176(12):7686-94.
4. Platanias L.C., 2005. Mechanisms of type-I- and type-II-interferon-mediated signalling. Nat Rev Immunol. 5(5):375-86.
5. Schroder K. et al., 2004. Interferon-gamma: an overview of signals, mechanisms, and functions. J Leukoc Biol. 75(2):163-89.

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