Human TLR4/NF-kB NF-kB/SEAP - (SEAPorter™) Stable Reporter Cell Line

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Human TLR4/NF-kB NF-kB/SEAP - (SEAPorter™) Stable Reporter Cell Line Summary

• Description: The TLR4 reporter cell line is a stably co-transfected cell line which expresses full-length human Toll-like receptor 4 (TLR4) and the secreted alkaline phosphatase (SEAP) reporter gene under the transcriptional control of an NF-kB response element.
• Target Species: Human
• Specificity: TLR4/NF-kB/SEAP
• Selection Agent: Blasticidin and G418.
• Growth Properties: Adherent Morphology: Epithelial
• RCL Type: NF-kB/SEAP - (SEAPorter™)
• Reporter Gene: Secreted alkaline phosphatase (SEAP)
• Host: HEK293

Applications/Dilutions

• Dilutions:
  • Flow Cytometry
  • Ligand Activation
• Application Notes: The TLR4 reporter line can be used for TLR4-dependent functional assays as well as screening of TLR4 agonists or antagonists. The TLR4 reporter cell line has been validated by flow cytometry (Fig. 1). Functional activity of this cell line has been validated by transfecting the MD2 plasmid and analyzing SEAP activity with SEAPorter™ Assay Kit (NBP2-25285) (Fig. 2).
  
  Contents: 3~4 x 10^6 cells
  Biosafety Level: 2
• Agonist:
  • LPS from E. Coli, TLR4 ligand NBP2-25295
• Readout System:
  • Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric) NBP2-25285
  • Cell Surface Staining Flow Cytometry Kit NBP2-26247

Packaging, Storage & Formulations

• Storage: Store in gas phase of liquid nitrogen.
• Reconstitution Instructions: Complete Growth Medium: DMEM with 4.5 g/L glucose + 10% FBS + 4 mM L-glutamine + 1 mM sodium pyruvate + 100 units/ml penicillin + 0.1 mg/ml streptomycin + 10 ug/ml blasticidin + 0.5 mg/ml G418 (Geneticin).

Details for Array

• Type: Cell

Notes

Assume all cultures are hazardous since they may harbor latent viruses or other organisms that are uncharacterized. The following safety precautions should be observed.
- Use pipette aids to prevent ingestion and keep aerosols down to a minimum.
- No eating, drinking or smoking while handling the TLR4 reporter line.
- Wash hands after handling the TLR4 reporter line and before leaving the lab.
- Decontaminate work surface with disinfectant or 70% ethanol before and after working with cells.
- All waste should be considered hazardous.
- Dispose of all liquid waste after each experiment and treat with bleach.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Reporter Cell Lines are guaranteed for 1 year from date of receipt.

Publications for TLR4/NF-kB Reporter Cell Line (NBP2-26276)(7)

Publications using NBP2-26276

• Lee Y, Lee IY, Yun HJ et al. BAT3 negatively regulates lipopolysaccharide-induced NF-kB signaling through TRAF6 Biochem. Biophys. Res. Commun. 2016-09-16 [PMID: 27501752] (WB, Rat)
• O'Neill S, Humphries D, Tse G et al. Heat shock protein 90 inhibition abrogates TLR4-mediated NF-kB activity and reduces renal ischemia-reperfusion injury. Sci Rep. 2015-08-07 [PMID: 26248657] (Func)
• Chan M, Hayashi T, Mathewson RD et al. Identification of substituted pyrimido[5,4-b]indoles as selective Toll-like receptor 4 ligands. J Med Chem. 2013-06-13 [PMID: 23656327]
• Ahmed I, Roy B, Chandrakesan P et al. Evidence of functional cross talk between the Notch and NF-kB pathways in nonneoplastic hyperproliferating colonic epithelium. Am J Physiol Gastrointest Liver Physiol. 2013-02-15 [PMID: 23203159]
  
  Details:
  1. TLR4/NF-kB/SEAPorter HEk 293 cell line (IML-104).
• Feng D, Sangster-Guity N, Stone R et al. Differential requirement of histone acetylase and deacetylase activities for IRF5-mediated proinflammatory cytokine expression. J Immunol. 2010-11-15 [PMID: 20935208]
• Chandrakesan P, Ahmed I, Anwar T et al. Novel changes in NF-{kappa}B activity during progression and regression phases of hyperplasia: role of MEK, ERK, and p38. J Biol Chem. 2010-10-22 [PMID: 20710027]
• Greene CM, Carroll TP, Smith SG et al. TLR-induced inflammation in cystic fibrosis and non-cystic fibrosis airway epithelial cells. J Immunol. 2005-02-01 [PMID: 15661927] (Func)
  
  Details:
  Functional Assays: 1.TLR2/NF-kB/SEAP Stably Transfected HEK 293 Cells (IML-102). 2. TLR4/MD-2/NF-kB/SEAP Stably Transfected HEK 293 Cells (IML-104MD2). 3. TLR4/NF-kB/SEAP Stably Transfected HEK 293 Cells (IML-104).4.TLR7/NF-kB/SEAP Stably Transfected HEK 293 Cells (IML-107). 5. LPS from E. Coli, TLR4 Ligand (IMG-2204). 6. SEAPorter Assay Kit (10055K).

FAQs for TLR4/NF-kB Reporter Cell Line (NBP2-26276). (Showing 1 - 3 of 3 FAQs).

1. We are interested in your product, TLR4/NF-kB/SEAP Stable Reporter Cell Line (#NBP2-26276. We want to know sequence of NF-kB response element that was used for this product. If you could disclose such information, could you please let us know?
   • Thank you for your inquiry about the TLR4/NF-kB SEAP reporter cell line, Cat no NBP2-26276. I am sorry but the sequence of the NF-kB response element is proprietary and we are not able to disclose that information.
2. The protocol describes that cells settle down in 1-3 days; however, the cells took longer to grow. Why is this?
   • The time to settle down can vary and can take longer than 1-3 days depending on a number of variables, including cell culture conditions, cell recovery from shipment/storage, etc. One - 3 days is an approximate time. It would be impossible to predict exactly how long it would take the cells to settle down in a given investigators hands and conditions. Remember the cells have travelled around the world and outside of their usual liquid nitrogen storage, and then are brought out of storage into culture. They are exposed to many variables throughout that process, they make take up to 5-7 days to settle down.
3. Why are these cells in biosafety level 2?
   • These cell lines were developed from the HEK293 cell line. The HEK293 line (ATCC CRL-1573) itself is designated as biosafety level two, please see the designation on this link. Hence, since the cell lines were derived from the cell line they are also designated as safety level two.