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THP1-Dual™ KO-STING Cells

THP1-Dual KO-STING Cells Unit size Cat. code Docs Qty Price
Human THP-1 Monocytes - STING knockout NF-κB-SEAP and IRF-Lucia Reporter Cells
3-7 x 10e6 cells
thpd-kostg
+-
$1,373.00

June, 18th 2018 - CONTENTS UPDATE NOTIFICATION
Please note that, for your convenience, these cells are now provided with QUANTI-Blue™ Solution.

STING knockout dual reporter monocytes

THP1-Dual™ KO-STING cells were generated from THP1-Dual™ cells by stable knockout of the STING gene. STING (stimulator of interferon genes), alternatively known as MPYS, TMEM173, MITA and ERIS is a direct sensor of cyclic dinucleotides (CDNs) [1-3]. These cell lines were derived from human THP‑1 monocytes, a cell line often used to study DNA sensing pathways as they express all the cytosolic DNA sensors identified so far (with the exception of DAI).

THP1-Dual™ and THP1-Dual™ KO-STING cells stably express two inducible secreted reporter genes: IFN-inducible Lucia luciferase and NF‑κB-inducible SEAP (secreted embryonic alkaline phosphatase). They can be used to study the role of STING by monitoring IRF-induced Lucia luciferase activity, using QUANTI‑Luc™, a Lucia™ luciferase detection reagent.

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

 

1. Burdette DL. et al., 2011. STING is a direct innate immune sensor of cyclic di-GMP. Nature. 478(7370):515-8.
2. Zhang X. et al., 2013. Cyclic GMP-AMP containing mixed phosphodiester linkages is an endogenous high-affinity ligand for STING. Mol Cell. 51(2):226-35.
3. Barber GN. et al., 2015. STING: infection, inflammation and cancer. Nat Rev Immunol. 15(12):760-70.


IRF responses in THP1-Dual™ and THP1-Dual™ KO-STING cells to various stimuli. Cells were incubated with Poly(I:C) LMW/LyoVec™ (100 ng/ml), VACV-70/LyoVec™ (1 µg/ml), 2'3'-cGAMP (3 µg/ml), and c-di-AMP (10 µg/ml). Human IFN-α and IFN-β (1x104 U/ml) serve as positive controls. After a 24h incubation, IRF activation was determined by measuring the relative light units (RLUs) in a luminometer using QUANTI-Luc™, a Lucia luciferase detection reagent. The IRF induction of each ligand is expressed relative to that of hIFN-β at 1x104 U/ml (taken as 100%).

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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)

Quality control:

Biallelic STING knockout is verified by functional assays, PCR and DNA sequencing.

These cells are guaranteed mycoplasma-free.

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

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Contents

  • 1 vial of THP1-Dual™ KO-STING cells (3-7 x 106 cells) in freezing medium
  • 1 ml of Normocin™ (50 mg/ml). Normocin™ is a formulation of three antibiotics active against mycoplasmas, bacteria and fungi.
  • 1 ml of Zeocin™ (100 mg/ml)
  • 1 ml of Blasticidin (10 mg/ml)
  • 1 pouch of QUANTI-Luc™
  • 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)

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Description

THP1-Dual™ and THP1-Dual™ KO-STING cells stably express two inducible secreted reporter genes: Lucia luciferase and SEAP (secreted embryonic alkaline phosphatase). The Lucia luciferase reporter gene is under the control of an ISG54 (interferon-stimulated gene) minimal promoter in conjunction with five IFN-stimulated response elements. The SEAP gene is driven by an IFN-β minimal promoter fused to five copies of the NF‑κB response element. As a result, they allow the simultaneous study of the IFN regulatory factor (IRF) pathway, by assessing the activity of Lucia luciferase and the NF-κB pathway, by monitoring the activity of SEAP. Both reporter proteins are readily measurable in the cell culture supernatant when using QUANTI‑Luc™, a Lucia™ luciferase detection reagent and QUANTI-Blue™, a SEAP detection reagent.

THP1-Dual™ KO-STING and THP1-Dual™ cells can be used to study the role of STING by monitoring IRF-induced Lucia luciferase activity. Unlike the parental cells, THP1-Dual™ KO‑STING cells exhibit no detectable response to cytosolic DNA and CDNs while retaining the ability to respond to type I IFNs. As expected these cells remain responsive to RIG-I ligands such as transfected Poly(I:C).

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