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Human TLR2 HEK293 Reporter Cells (NF-κB and IL-8)

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HEK-Blue-Lucia™ hTLR2

Human TLR2 expressing HEK293 dual reporter cells (NF-κB pathway)

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3-7 x 10e6 cells

hkd-htlr2ni
+-
$1,457

Notification: This cell line has been renamed. It was formerly known as "HEK-Dual™ hTLR2 (NF/IL8)". The cat. code (hkd-htlr2ni) remains unchanged.
This product is for internal research use only. Additional rights may be available. Please visit InvivoGen’s Terms and Conditions.

NF-κB–SEAP and IL-8–Lucia reporter HEK293 cells expressing human TLR2

Signaling pathways in HEK-Blue-Lucia™ hTLR2 cells
Signaling pathways in HEK-Blue-Lucia™ hTLR2 cells

InvivoGen offers a human embryonic kidney 293 (HEK293)-derived cell line, specifically designed to assess the distinct role of the human Toll-like receptor 2 (hTLR2):

— HEK-Blue-Lucia™ hTLR2 cells* 

These cells were generated from the HEK-Blue-Lucia™ Null​ cell line harboring two inducible reporter genes. This feature allows the double readout of the NF-κB/AP-1 pathway, by monitoring the SEAP (secreted embryonic alkaline phosphatase) or Lucia luciferase activities. HEK-Blue-Lucia™ hTLR2 cells also stably express the hTLR2 and CD14 genes. Due to a triple knockout (KO) of TLR3, TLR5, and TNFR, this cell line allows for the independent study of TLR2.

 

Stimulation of HEK-Blue-Lucia™ hTLR2 cells with TLR2 agonists (e.g. Pam3CSK4) triggers the activation of the artificial NF-κB-inducible promoter and the subsequent production of SEAP. It also promotes the expression of Lucia luciferase, which is knocked in (KI) downstream of the endogenous (more physiological) IL-8 promoter (see figures).

IL-8 (interleukin 8) is a chemokine produced in response to TLR agonists in an NF-κB/AP-1-dependent manner [1-2]. This feature enables the double readout study of the NF-κB/AP-1 pathway, by monitoring the activity of SEAP and Lucia luciferase using QUANTI-Blue™ Solution (SEAP detection reagent) or QUANTI-Luc™ 4 Lucia/Gaussia (luciferase detection reagent), respectively. Thus, you may choose the readout depending on your laboratory equipment utilizing a spectrophotometer for SEAP or a luminometer for Lucia luciferase detection.
 

TLR2 is an important pattern recognition receptor (PRR) detecting a large spectrum of microbial pathogen-associated molecular patterns (PAMPs) from Gram-positive and Gram-negative bacteria as well as fungi, parasites, and viruses [3].

More details More details

 

Read our review Read our review on TLR2

 

Key features:

  • Stable overexpression of hTLR2 and CD14
  • Verified KO for the TLR3, TLR5, and TNFR genes 
  • Functionally validated using a selection of PRR ligands and cytokines
  • Readily assessable NF-κB activation by assessing the SEAP and/or Lucia luciferase activities

Applications:

  • Defining the role of TLR2-dependent NF-κB signaling pathway
  • Screening for novel TLR2 agonists and inhibitors
  • Choice of readout depending on the laboratory equipment (spectrophotometer for SEAP or luminometer for Lucia luciferase detection).

 

* formerly named HEK-Dual™ hTLR2 (NF/IL8) cells

 

References:

1. Roebuck KA. 1999. Regulation of interleukin-8 gene expression. J Interferon Cytokine Res:429-38.
2. Ohta K, et al. 2014. Toll-like receptor (TLR) expression and TLR‑mediated interleukin-8 production by human submandibular gland epithelial cells. Mol Med Rep. (5):2377-82.
3. Oliveira-Nascimento, L. et al. 2012. The Role of TLR2 in Infection and Immunity. Front Immunol 3, 79.

Figures

NF-κB - SEAP response
NF-κB - SEAP response

NF-κB–SEAP response. HEK-Blue-Lucia™ hTLR2  and HEK-Blue™ hTLR2 cells were stimulated with various TLR agonists: FSL-1 (TLR2 agonist; 30 pg/ml), Pam3CSK4 (TLR2 agonist; 30 pg/ml), HKLM (TLR2 agonist; 1x107 cells/ml), Poly(I:C) (TLR3 agonist; 3 µg/ml), FLA-ST (flagellin from S. typhimurium, TLR5 agonist; 100 ng/ml), and TNF-α (10 ng/ml). After overnight incubation, the activation of NF-κB was assessed by measuring the activity of SEAP in the supernatant using QUANTI-Blue™ Solution. Data are shown as optical density (OD) at 655 nm (mean ± SEM).

NF-κB (IL-8) - Lucia response
NF-κB (IL-8) - Lucia response

NF-κB (IL-8)–Lucia response using QUANTI-Luc™. HEK-Blue-Lucia™ hTLR2 cells were stimulated with TLR2 agonists: FSL-1 (TLR2 agonist; 30 pg/ml), Pam3CSK4 (TLR2 agonist; 30 pg/ml) and HKLM (1x107 cells/ml). After 24h incubation, activation of the IL-8 was assessed by measuring the activity of Lucia luciferase in the supernatant using QUANTI-Luc™. Data are shown in fold response over non-induced cells (mean ± SEM).

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Specifications

Antibiotic resistance: BlasticidinHygromycinZeocin®

Growth medium: DMEM, 4.5 g/l glucose, 2 mM L-glutamine, 10% (v/v) heat-inactivated fetal bovine serum, 100 U/ml penicillin, 100 μg/ml streptomycin, 100 μg/ml Normocin™

Quality Control:

  • Human TLR2 and CD14 overexpression has been verified by flow cytometry and functional assays.
  • The triple KO of TLR3, TLR5, and TNFR has been verified by DNA sequencing, PCR, and functional assays.
  • The stability for 20 passages, following thawing, has been verified.
  • These cells are guaranteed mycoplasma-free. 

Note: HEK-Blue-Lucia™ hTLR2 cells are resistant to BlasticidinHygromycin, and Zeocin®. They should be maintained in growth medium supplemented with Hygromycin and Zeocin®.

 

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

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Contents

Dry Ice Shipped on dry ice (Europe, USA, Canada, and some areas in Asia)

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Details

Toll-like receptor 2 (TLR2) plays an essential role in detecting a diverse range of microbial pathogen-associated molecular patterns (PAMPs) from Gram-positive and Gram-negative bacteria as well as fungi, parasites, and viruses. These PAMPs include cell-wall components such as lipoproteins, lipoteichoic acid (LTA; Gram-positive bacteria only), lipoarabinomannan (mycobacteria only), and zymosan (yeast) [1]. TLR2 forms a heterodimer on the cell surface, crucial for signaling and ligand specificity, with co-receptors TLR1 or TLR6. For example, TLR2-TLR1 and TLR2-TLR6 heterodimers are known to bind specific lipoproteins depending on whether they are tri- or diacylated, respectively [2, 3]. Moreover, ligand recognition is enhanced by its non-specific delivery to TLR2 by CD14, and sometimes in combination with additional ligand-specific molecules such as CD36 and Dectin-1 [4, 5].

Upon ligand recognition, TLR2-dependent signaling cascades ultimately lead to a MyD88 and MAL/TIRAP-dependent activation of pro-inflammatory transcription factors such as NF-κB and AP-1 [6]. Additionally, the PI3K/Akt pathway may also be activated leading to the production of anti-inflammatory cytokines [7]. Interestingly, microarray data generated by InvivoGen clearly highlights that downstream effects differ depending on whether it’s the TLR2-TLR1 or TLR2-TLR6 heterodimer that is activated upon ligand recognition.

 

References:

1. Oliveira-Nascimento, L. et al. 2012. The Role of TLR2 in Infection and Immunity. Front Immunol 3, 79.
2. Takeuchi, O. et al. 2001. Discrimination of bacterial lipoproteins by Toll-like receptor 6. Int Immunol 13, 933-940.
3. Takeuchi, O. et al. 2002. Cutting edge: role of Toll-like receptor 1 in mediating immune response to microbial lipoproteins. J Immunol 169, 10-14.
4. Jimenez-Dalmaroni, M.J. et al. 2009. Soluble CD36 ectodomain binds negatively charged diacylglycerol ligands and acts as a co-receptor for TLR2. PLoS One 4, e7411.
5. Lotz, S. et al. 2004. Highly purified lipoteichoic acid activates neutrophil granulocytes and delays their spontaneous apoptosis via CD14 and TLR2. J Leukoc Biol 75, 467-477.
6. Piao, W. et al. 2016. Differential adapter recruitment by TLR2 co-receptors. Pathog Dis 74.
7. Santos-Sierra, S. et al. 2009. Mal connects TLR2 to PI3Kinase activation and phagocyte polarization. EMBO J 28, 2018-2027.

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