Human TLR9 Dual Reporter THP-1 Cells

NF-κB-SEAP and IRF-Lucia reporter monocytes

ABOUT

Dual reporter monocytes for TLR9 pathway studies

InvivoGen offers a human monocyte-derived cell line, specifically designed for the study of human TLR9 (Toll-Like Receptor 9) signaling pathways:

• THP1-Dual™ hTLR9 cells

THP1-Dual™ hTLR9 cells overexpress the human TLR9 (hTLR9) gene. They also feature two inducible reporter genes, allowing the concomitant study of the IRF and NF-κB pathways, by monitoring the Lucia luciferase and SEAP (secreted embryonic alkaline phosphatase) activities, respectively.

Unlike their parental cells, THP1-Dual™, which weakly express TLR9, THP1-Dual™ hTLR9 cells are responsive to stimulation with TLR9 agonists, such as oligonucleotides containing CpG motifs (CpG ODNs). As expected, hTLR9 overexpression in THP1-Dual™ cells allows potent NF-κB and IRF responses upon incubation with CpG-ODNs of class A (ODN 2216), class B (ODN 2006, ODN 1826), and class C (ODN 2395). Of note, THP1-Dual™ hTLR9 cells are more responsive to the class B, human-preferred, ODN 2006 (see Figures).

 

Background:

Toll-Like Receptor 9 (TLR9) is one of the most studied pattern recognition receptors (PRRs) for nucleic acids. It is an endosomal receptor that triggers NF-κB- and IRF-mediated pro-inflammatory responses upon the recognition of unmethylated cytosine-phosphorothioate-guanosine (CpG) forms of DNA [1-3]. TLR9 agonists can be mimicked by synthetic oligonucleotides containing CpG motifs (CpG ODNs) [1,3].

More details More details

Key features:

  • Verified overexpression of TLR9 gene by qRT-PCR
  • Functionally validated with a selection of TLR9 agonists
  • Readily assessable Lucia luciferase and SEAP reporter activities

Applications:

  • Study of IRF and NF-κB-dependent TLR9 signaling pathways
  • Screening of novel specific activators or inhibitors of the TLR9 signaling pathways


References

1. Kumagai Y. et al., 2008. TLR9 as a key receptor of the recognition of DNA. Adv. Drug. Deliv. Rev. 60(7):795-804.
2. Heinz L.X. et al., 2021. TASL is the SLC15A4-associated adaptor for IRF5 activation by TLR7-9. Nature. 581(7808):316-322.
3. Kayraklioglu N. et al., 2021. CpG oligonucleotides as vaccine adjuvants. DNA Vaccines: Methods and Protocols. Methods in Molecular Biology. Vol. 2197. p51-77.

Disclaimer:  These cells are for internal research use only and are covered by a Limited Use License (See Terms and Conditions). Additional rights may be available.

SPECIFICATIONS

Specifications

Species
Human
Tested applications

Dual reporter assays for TLR9 pathway, NF-κB and IRF signaling pathways

Cell type
Monocytic
Growth properties
Suspension
Tissue origin
Human monocytes
Reporter gene
Lucia®
SEAP
Antibiotic resistance
Blasticidin
Zeocin®
Puromycin
Growth medium

Complete RPMI 1640 (see TDS)

Mycoplasma-free

Verified using Plasmotest

Quality control

Each lot is functionally tested and validated.

CONTENTS

Contents

  • Product: 
    THP1-Dual™ hTLR9 Cells
  • Cat code: 
    thpd-htlr9
  • Quantity: 
    3-7 x 10^6 cells
Includes:

1 ml of Blasticidin (10 mg/ml), 1 ml of Zeocin® (100 mg/ml), 1 ml of Normocin® (50 mg/ml), 1 tube of QUANTI-Luc™ 4 Reagent (Lucia luciferase detection reagent), 1 ml of QB reagent and 1 ml of QB buffer (SEAP detection reagent)

Shipping & Storage

  • Shipping method:  Dry ice
  • Storage:

    • Liquid nitrogen
    Stability: 20 passages

Details

Toll-Like Receptor 9 (TLR9) is an endosomal receptor that triggers NF-κB- and IRF-mediated pro-inflammatory responses upon the recognition of unmethylated cytosine-phosphorothioate-guanosine (CpG) forms of DNA [1-3]. Unmethylated CpG dinucleotides are a hallmark of microbial (bacterial, viral, fungal, and parasite) DNA, as well as mitochondrial self-DNA [3,4]. These TLR9 agonists can be mimicked by synthetic oligonucleotides containing CpG motifs (CpG ODNs), which have been extensively studied to improve adaptive immune responses in the context of vaccination [1,3].

TLR9 is mainly expressed in subsets of Dendritic Cells and in B cells of all mammals. In rodents, but not in humans, TLR9 is also expressed in monocytes and macrophages [3]. The structure of the receptor varies by 24% between human TLR9 (hTLR9) and mouse TLR9 (mTLR9) [3]. They recognize different CpG motifs, the optimal sequences being GTCGTT and GACGTT for hTLR9 and mTLR9, respectively [5].
 

More Information Get more information about CpG-ODNs Classes.

 

References

1. Kumagai Y. et al., 2008. TLR9 as a key receptor of the recognition of DNA. Adv. Drug. Deliv. Rev. 60(7):795-804.
2. Heinz L.X. et al., 2021. TASL is the SLC15A4-associated adaptor for IRF5 activation by TLR7-9. Nature. 581(7808):316-322.
3. Kayraklioglu N. et al., 2021. CpG oligonucleotides as vaccine adjuvants. DNA Vaccines: Methods and Protocols. Methods in Molecular Biology. Vol. 2197. p51-77.
4. Kumar V., 2021. The trinity of cGAS, TLR9, and ALRs: guardians of the cellular galaxy against host-derived self-DNA. Front. Immunol. 11:624597.
5. Bauer S. et al., 2001. Human TLR9 confers responsiveness to bacterial DNA via species-specific CpG motif recognition. Proc Natl Acad Sci USA, 98(16):9237-42.

DOCUMENTS

Documents

THP1-Dual™ hTLR9 Cells

Technical Data Sheet

Safety Data Sheet

Validation Data Sheet

Certificate of analysis

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