B16-Blue™ IFN-γ Cells

Murine IFN-gamma sensor cells

B16-Blue™ IFN-γ cells signaling

B16-Blue™ IFN-γ cells allow the detection of bioactive murine IFN-γ (mIFN-γ) by monitoring the activation of the JAK/STAT/ISRE pathway.

IFN-γ, also known as Type II IFN, is a pleiotropic cytokine with anti-viral, anti-tumor, and immunomodulatory functions [1]. IFN-γ binds a distinct cell-surface receptor, composed of two subunits, IFNGR1 and IFNGR2, which are associated with JAK1 and JAK2, respectively [2]. Upon binding to its receptor, IFN-γ triggers the JAK/STAT/ISRE pathway.

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Cell line description:

B16-Blue™ IFN-γ cells derive from the murine B16 melanoma cell line of C57BL/6 origin after stable transfection with a SEAP (secreted embryonic alkaline phosphatase) reporter gene under the control of the IFN-inducible ISG54 promoter enhanced by a multimeric interferon-sensitive response element (ISRE).
B16-Blue™ IFN-γ cells do not respond to IFN-α/β, due to the inactivation of the type I IFN receptor. B16-Blue™ IFN-γ cells respond specifically to mIFN-γ and do not respond to human IFN-γ.

Stimulation of B16-Blue™ IFN-γ cells with mIFN-γ triggers the production of SEAP. Levels of SEAP in the supernatant can be easily determined with QUANTI-Blue™ Solution, a medium that turns purple/blue in the presence of SEAP and by reading the OD at 655 nm.

Features of B16-Blue™ IFN-γ cells:

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

Applications of B16-Blue™ IFN-γ cells:

• Detection of murine IFN-γ
• Screening of anti-mIFN-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.
2. Platanias LC., 2005. Mechanisms of type-I- and type-II-interferon-mediated signalling. Nat Rev Immunol. 5(5):375-86.

Figures

Dose-response of B16-Blue™ IFN-γ cells to recombinant murine IFN-γ.
Cells were stimulated with increasing concentrations of recombinant murine IFN-γ. After overnight incubation, the ISGF3 response was determined using QUANTI-Blue™ Solution, a SEAP detection reagent, and reading the optical density (OD) at 630 nm. The OD at 630 nm is shown as mean ± SEM.

Response of  B16-Blue™ IFN-γ cells to a panel of cytokines.
Cells were stimulated with various human and murine recombinant cytokines: 10 ng/ml of mIFN-γ, mIFN-λ, hIFN-γ and 1000U/ml of mIFN-αA (also known as mIFN-α3), mIFN-β, hIFN-α2a, hIFN-β. After overnight incubation, SEAP activity was assessed using QUANTI-Blue™ Solution. The OD at 630 nm is shown as mean ± SEM.

Specifications

Detection range for murine IFN-γ: 0.1 ng - 1 µg/ml for mIFN-γ

Antibiotic resistance: Zeocin®

Quality control:

• B16-Blue™ IFN-γ cells were stimulated with murine IFN-α, IFN-β and IFN-γ. The cells produce SEAP only in response to murine IFN-γ
• These cells are guaranteed mycoplasma-free

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

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 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) is the sole member of the type II IFN family. It is secreted from CD4+ Th1 cells and activated NK cells. It plays a role in activating lymphocytes to enhance anti-microbial and anti-tumor effects [1-]. In addition, it 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 molecules form homodimers and are translocated to the nucleus where they bind IFN-stimulated response elements (ISRE) in the promoter of IFN 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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