TLR4 KO Dual Reporter THP1 Cells
THP1-Dual™ KO-TLR4 Cells | Unit size | Cat. code | Docs | Qty | Price |
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Human THP-1 Monocytes - TLR4 knockout NF-κB-SEAP and IRF-Lucia Reporter Cells |
3-7 x 10e6 cells |
thpd-kotlr4 |
TLR4 knockout dual reporter monocytes
NF-κB and IRF signaling pathways in THP1-Dual™ KO-TLR4 cells
THP1-Dual™ KO-TLR4 cells were generated from the THP1-Dual™ cell line, which is derived from the human THP-1 monocytes, through the stable knockout of the TLR4 gene. THP1-Dual™ KO-TLR4 and THP1-Dual™ cells feature two reporter genes allowing the simultaneous study of the IRF pathway, by monitoring the activity of an inducible secreted Lucia luciferase, and the NF-κB pathway by monitoring the activity of an inducible SEAP (secreted embryonic alkaline phosphatase). Lucia luciferase and SEAP activities are readily assessable in the supernatant using QUANTI-Luc™ and QUANTI-Blue™ Solution detection reagents, respectively.
Toll-like receptor 4 (TLR4) was the first TLR identified and is an important pattern recognition receptor (PRR) in innate immunity and inflammation. TLR4 is found both on the cell surface and in endosomes of innate immune cells including monocytes and macrophages, as well as on intestinal epithelium and endothelial cells [1]. TLR4 primarily recognizes and is activated by lipopolysaccharide (LPS) and its toxic moiety Lipid A, a core component of Gram-negative bacteria [2]. TLR4 does not directly interact with LPS and requires essential co-receptors, namely lipid-binding protein (LBP), MD-2, and CD14, to extract and deliver monomeric LPS to TLR4 [3]. There are two distinct signaling cascades triggered by the dimerization of TLR4; the MyD88-dependent (at the cell surface) and TRIF-dependent (in endosomes) pathways. At the cell surface, activation of TLR4 initiates the MyD88-dependent pathway, ultimately leading to the ‘early’ activation of NF-κB and the production of a pro-inflammatory response [3]. Subsequently, the TLR4 complex can be endocytosed into endosomes and result in the ‘late’ activation of NF-κB as well as the stimulation of IRF3 (interferon regulatory factor), which modulates the expression of type I IFNs [4]. TLR4 signaling is crucial in both acute and chronic inflammatory disorders and is thus an attractive target for novel treatments for conditions such as sepsis and cancer [1].
Features of THP1-Dual™ KO-TLR4 cells:
- Verified knockout of the TLR4 gene (PCR, DNA sequencing, and functional assays)
- Functionally validated on a selection of PRR ligands and cytokines
- Readily assessable Lucia luciferase and SEAP reporter activities
- The stability for 20 passages, following thawing, has been verified
- Guaranteed mycoplasma-free
Applications for THP1-Dual™ KO-TLR4 cells:
- Defining the role of TLR4 in PRR-induced signaling, or other cell signaling pathways
- Exclusion of contaminating LPS-dependent signaling
- Highlighting possible overlap between TLR4 and other signaling pathways
References:
1. Ou, T. et al. 2018. The Pathologic Role of Toll-Like Receptor 4 in Prostate Cancer. Front Immunol 9, 1188.
2. Cochet, F. et al. 2017. The Role of Carbohydrates in the Lipopolysaccharide (LPS)/Toll-Like Receptor 4 (TLR4) Signalling. Int J Mol Sci 18
3. Kuzmich, N.N. et al. 2017. TLR4 Signaling Pathway Modulators as Potential Therapeutics in Inflammation and Sepsis. Vaccines (Basel) 5.
4. Marongiu, L. et al. 2019. Below the surface: The inner lives of TLR4 and TLR9. J Leukoc Biol 106, 147-160.
Specifications
Antibiotic resistance: Blasticidin and Zeocin™
Growth medium: RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% (v/v) fetal bovine serum (FBS), 100 U/ml penicillin, 100 µg/ml streptomycin, 100 µg/ml Normocin™
Quality Control:
- TLR4 knockout has been verified by PCR, DNA sequencing, and functional assays.
- The stability for 20 passages, following thawing, has been verified.
- These cells are guaranteed mycoplasma-free.
This product is covered by a Limited Use License (See Terms and Conditions).
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- 3-7 x 106 THP1-Dual™ KO-TLR4 cells in a cryovial or shipping flask
- 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)