THP1-Dual™ Cells

NF-kB-SEAP and IRF-Lucia luciferase Reporter Monocytes

THP1-Dual™ cells were derived from the human THP-1 monocyte cell line by stable integration of two inducible reporter constructs.

As a result, THP1-Dual™ cells allow the simultaneous study of the NF-κB pathway, by monitoring the activity of SEAP, and the IRF pathway, by assessing the activity of a secreted luciferase (Lucia).

Both reporter proteins are readily measurable in the cell culture supernatant when using QUANTI-Blue™, a SEAP detection reagent, and QUANTI-Luc™, a luciferase detection reagent.

THP1-Dual™ cells induce the activation of NF-κB in response to certain TLR agonists, such as Pam3CSK4 and flagellin. They trigger the IRF pathway upon stimulation with type I IFNs and RLR or CDS agonists, such as transfected dsRNA.

THP1-Dual™ cells are resistant to the selectable markers Zeocin™ and blasticidin.

These products are covered by a Limited Use License (See Terms and Conditions).

THP1-Dual™ cells were stimulated with 1 ng/ml Pam3CSK4 (TLR2), 100 ng/ml LPS-EK UP (TLR4), 100 ng/ml FLA-ST UP (TLR5), 10 μg/ml R848 (TLR7/8), 10 μg/ml Tri-DAP (NOD1), 3 μg/ml 2’3’-cGAMP (STING), 1 μg/ml poly(I:C)/LyoVec™ (RLR) or 100 ng/ml poly(dA:dT)/LyoVec™ (CDS). After 24h incubation, NF-κB and IRF activation was assessed by measuring the levels of SEAP and Lucia luciferase using QUANTI‑Blue™ and QUANTI-Luc™, respectively. With QUANTI‑Blue™ the levels of SEAP were determined by reading the optical density (OD) at 655 nm. With QUANTI-Luc™ the levels of Lucia luciferase were determined by measuring the relative light units (RLUs) in a luminometer.

Cells were pretreated or not with PMA and incubated with 1 µg/ml Pam3CSK4, 1 µg/ml poly(I:C)/LyoVec, 1 µg/ml LPS-EK UP, 1 µg/ml FLA-ST UP, 10 µg/ml R848, 10 µg/ml ODN2006, 10 µg/ml Tri-DAP, 10 µg/ml MDP, 1 µg/ml 5’ppp-dsRNA/Lyovec, 1 µg/ml poly(dG:dC)/LyoVec or 10e4 U/ml IFN-α.

After 24h incubation, levels of SEAP and Lucia luciferase were assessed using QUANTI-Blue™ and QUANTI-Luc™, respectively.

 

Specifications

Antibiotic resistance: Zeocin™, blasticidin

Growth medium: RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum, 100 μg/ml Normocin™, Pen-Strep (100 U/ml-100 μg/ml)

Guaranteed mycoplasma-free

Contents

THP1-Dual™ is supplied with:

• 1 vial of THP1-Dual™ cells (3-7 x 106 cells) in Freezing Medium.
• 1 ml Normocin™ (50 mg/ml).
• 100 μl Zeocin™ (100 mg/ml).
• 100 μl Blasticidin (10 mg/ml).
• 1 pouch of QUANTI-Luc™ (Luciferase detection medium).
• 1 pouch of QUANTI-Blue™ (SEAP detection medium).

THP1-Dual™ cells are shipped on dry ice.

See Data Sheet for storage conditions of each component.

Description

THP1-Dual™ cells feature the Lucia gene, a secreted luciferase reporter gene, under the control of an ISG54 minimal promoter in conjunction with five IFN-stimulated response elements.

THP1-Dual™ cells also express a secreted embryonic alkaline phosphatase (SEAP) reporter gene driven by an 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.

As a result, THP1-Dual™ cells allow the simultaneous study of the NF-κB pathway, by monitoring the activity of SEAP, and the IRF pathway, by assessing the activity of Lucia luciferase.