Jurkat-Dual™ Cells

NF-κB & IRF signaling in Jurkat-Dual™ cells

IRF-SEAP & NF-κB-Luc Reporter T Lymphocytes

Jurkat-Dual™ cells are engineered reporter T lymphocytes designed for the study of transcription factor activation in a physiologically-relevant cell line. These cells stably express two inducible reporter constructs; interferon regulatory factor-inducible secreted embryonic alkaline phosphatase (IRF-SEAP) and NF-κB-Lucia luciferase (NF-κB-Luc) reporter genes. They were derived from the human T lymphocyte-based Jurkat cell line.

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Cell line description:

Jurkat-Dual™ cells feature the Lucia luciferase gene, a secreted luciferase reporter gene, driven by an interferon-β (IFN-β) minimal promoter fused to five copies of the NF-κB consensus transcriptional response element and three copies of the c-Rel binding site. These cells also express a secreted embryonic alkaline phosphatase (SEAP) reporter gene under the control of an IFN-stimulated gene 54 (ISG54) minimal promoter in conjunction with five IFN-stimulated response elements. As a result, Jurkat-Dual™ cells allow the simultaneous study of the NF-κB pathway, by monitoring the activity of Lucia luciferase, and the IRF pathway, by assessing the activity of SEAP. Both reporter proteins are readily measurable in the cell culture supernatant when using QUANTI-Luc™ 4 Lucia/Gaussia, a Lucia and Gaussia luciferase detection reagent, and QUANTI-Blue™ Solution, a SEAP detection reagent.

Jurkat-Dual™ cells induce the activation of NF-κB in response to TNF-α and T-Lymphocyte mitogens, such as phytohemagglutinin and concanavalin A. They trigger the IRF pathway upon stimulation with type I IFNs and poly(I:C).

Features of Jurkat-Dual™ cells:

• Readily assessable SEAP and Lucia luciferase reporter activity
• The stability for 20 passages has been verified
• Functionally tested and guaranteed mycoplasma-free

Application of Jurkat-Dual™ cells:

• Monitoring IRF and NF-κB activation in T lymphocytes

Figures

NF-κB/IRF dual response of Jurkat-Dual™ cells to various stimuli. Cells were incubated with 20 μg/ml of the mitogens phytohemagglutinin-P (PHA-P, the protein form of PHA) or concanavalin A (ConA), 300 IU/ml IFN-α, 300 IU/ml IFN-β, 100 ng/ml TNF-α or 10 μg/ml poly(I:C). After 24h incubation, NF-кB and IRF activation was assessed by measuring the levels of SEAP and Lucia luciferase in the supernatant using QUANTI-Luc™ or QUANTI‑Blue™ Solution, respectively.

Specifications

Antibiotic resistance: Zeocin®, blasticidin

Growth medium: IMDM, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum (FBS; 30 min at 56 °C), 100 μg/ml  Pen-Strep (100 U/ml-100 μg/ml), Normocin™

Quality control:

• Reporter activity has been validated using functional assays.
• The stability for 20 passages following thawing has been verified.
• These cells are guaranteed mycoplasma-free.

Contents

• 1 vial of Jurkat-Dual™ cells (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 tube of QUANTI-Luc™ 4 Reagent, a Lucia luciferase detection reagent (sufficient to prepare 25 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

The human Jurkat cell line was established from acute T cell leukemia. Jurkat cells have been extensively used in vitro to delineate the signaling pathways induced by the engagement of T-cell receptors and to study the expression of various chemokine receptors susceptible to viral entry, particularly HIV [1]. T cell activation results in the nuclear translocation of transcription factors, such as IRFs (interferon regulatory factors) and NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) to induce the expression of target genes. Knowledge of their signaling cascades, ease of culture, and transfection make Jurkat cells a convenient tool to screen for T-cell activation, anti-viral or anti-cancer drugs.

References:

1. Montano M. 2014. Model systems. Translational Biology in Medicine. 9-33.

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