TLR-expressing NF-κB–SEAP and IL-8–Lucia reporter cells
HEK-Blue-Lucia™ TLR& cells (formerly named HEK-Dual™ TLRs (NF/IL8) cell lines) are a collection of the human embryonic kidney 293 (HEK293)-derived cell lines designed to provide a rapid, sensitive, and reliable method to screen and validate TLR agonists or antagonists.
These cells express two inducible reporter genes for SEAP (secreted embryonic alkaline phosphatase) and Lucia luciferase. Stimulation with TLR agonists 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. IL-8 (interleukin 8) is a chemokine produced in response to TLR agonists in an NF-κB/AP-1-dependent manner [1-2].
This 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.
Key features
- Stable expression of a human or mouse TLR gene
- Stable expression of an NF-κB/AP1-inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene
- Stable expression of the Lucia luciferase reporter gene under the control of the endogenous IL-8 promoter (responsive to NF-κB)
- No activity of TLR3, TLR5, and TNFR (Tumor Necrosis Factor receptor) due to verified knockout of these three receptors
- Readily assessable NF-κB activation by assessing the SEAP and/or Lucia luciferase activities
Applications
- Defining the role of TLR-dependent NF-κB signaling pathways
- Screening for novel TLR agonists and inhibitors
- Comparing the TLR-dependent signaling between human and mouse TLR
- Choice of readout depending on the laboratory equipment (spectrophotometer for SEAP or luminometer for Lucia luciferase detection).
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.
