ODN 1826 FITC - TLR9 Agonist

FITC-labeled CpG ODN, Class B, Mouse-preferred

ABOUT

FITC labeled CpG oligonucleotide - Murine TLR9-preferred ligand

ODN 1826 FITC is an FITC (fluorescein isothiocyanate)-labeled version of ODN 1826, a Class B CpG oligonucleotide (ODN) with a preference for mouse Toll-like receptor 9 (mTLR9). This fluorescent label allows to monitor the internalization of the CpG ODN into the cells without affecting its TLR9-activating properties. ODN 1826 is a short synthetic single-stranded DNA molecule containing unmethylated CpG dinucleotides (CpG motifs). It contains a full phosphorothioate backbone and is nuclease-resistant. These synthetic unmethylated CpG motifs mimic microbial DNA and act as immunostimulants via TLR9.

More details More details

Mode of action

Stimulatory CpG ODNs are internalized and activate the endosomal receptor TLR9. Activation of TLR9 triggers NF-κB- and interferon regulatory factor (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]. Class B (also called Type K) CpG ODNs, such as ODN 1826, contain a full phosphorothioate backbone with one or more CpG dinucleotides. They strongly activate B cells but weakly stimulate IFN-α secretion in plasmacytoid dendritic cells [5].

ODN 1826 FITC can be used to evaluate CpG ODN cellular uptake and intracellular localization by confocal laser-scanning microscopy (excitation 495 nm, emission 520 nm) (data not shown) or flow cytometry (see figure). The labeled ODN retains its TLR9 biological activity as verified using HEK-Blue™ mTLR9 cells.

Key features of ODN 1826 FITC

  • Potent activator of mouse TLR9
  • FITC labeled
  • Synthetic ODN with unmethylated CpG motifs
  • Each lot is functionally tested

more info Get more information about CpG ODNs Classes.

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. Krieg A.M. et al., 1995. CpG motifs in bacterial DNA trigger direct B-cell activation. Nature. 374(6522):546-9.

All products are for research use only, and not for human or veterinary use.

SPECIFICATIONS

Specifications

Source
Synthetic
Target

TLR9

Sequence

5’-tccatgacgttcctgacgtt-3’ (20 mer) 

Note: Regular letters represent phosphorothioate bonds, bold letters represent CpG dinucleotides.

Molecular weight
6901 g/mol
Tag
3' FITC
Working concentration

1-5 μM for cellular assays

Solubility

5 mg/ml in water

Appearance (form)
Lyophilized
Reconstitution buffer
Endotoxin-free water (provided)
Endotoxin

Negative (tested using EndotoxDetect™ assay)

Applications

Flow cytometry (tested), TLR9 activation in cellular assays (tested)

Quality control

Each lot is functionally tested and validated using cellular assays.

CONTENTS

Contents

  • Product: 
    ODN 1826 FITC
  • Cat code: 
    tlrl-1826f
  • Quantity: 
    50 µg
Includes:

1 ml of sterile endotoxin-free water

Notes:

ODN 1826 labeled with FITC at the 3’ terminus

Shipping & Storage

  • Shipping method:  Room temperature
  • Storage:

    • -20°C

    Caution:

    • Avoid repeated freeze-thaw cycles
    • Protect from light

Details

CpG ODNs

Synthetic oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODNs), such as ODN 1018, have been extensively studied as adjuvants [1]. These CpG motifs are present at a 20-fold greater frequency in bacterial DNA compared to mammalian DNA [2]. CpG ODNs are recognized by the Toll-like receptor 9 (TLR9), which is expressed on human B cells and plasmacytoid dendritic cells (pDCs), thereby inducing Th1-dominated immune responses [3]. Pre-clinical studies, conducted in rodents and non-human primates, as well as human clinical trials, have demonstrated that CpG ODNs can significantly improve vaccine-specific antibody responses [1]. Three types of stimulatory CpG ODNs have been identified, types A, B, and C, which differ in their immune-stimulatory activities [4-5]. 

Toll-like receptor 9

The Toll-like Receptor 9 (TLR9) is an endosomal receptor that triggers NF-κB- and interferon regulatory factor (IRF)-mediated pro-inflammatory responses upon the recognition of unmethylated cytosine-phosphorothioate-guanosine (CpG) forms of DNA [6-8]. Unmethylated CpG dinucleotides are a hallmark of microbial (bacterial, viral, fungal, and parasite) DNA, as well as mitochondrial self-DNA [8,9]. 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 [6,8].

TLR9 is mainly expressed in subsets of Dendritic Cells and B cells of all mammals. In rodents, but not in humans, TLR9 is also expressed in monocytes and macrophages [8]. The structure of the receptor varies by 24% between human TLR9 (hTLR9) and mouse TLR9 (mTLR9) [8]. They recognize different CpG motifs, the optimal sequences being GTCGTT and GACGTT for hTLR9 and mTLR9, respectively [10].

References:

1. Steinhagen F. et al., 2011. TLR-based immune adjuvants. Vaccine 29(17):3341-55.
2. Hemmi H. et al., 2000. A Toll-like receptor recognizes bacterial DNA. Nature 408:740-5.
3. Coffman RL. et al., 2010. Vaccine adjuvants: Putting innate immunity to work. Immunity 33(4):492-503.
4. Krug A. et al., 2001. Identification of CpG oligonucleotide sequences with high induction of IFN-alpha/beta in plasmacytoid dendritic cells. Eur J Immunol, 31(7): 2154-63.
5. Marshall JD. et al., 2005. Superior activity of the type C class of ISS in vitro and in vivo across multiple species. DNA Cell Biol. 24(2):63-72.
6. Kumagai Y. et al., 2008. TLR9 as a key receptor of the recognition of DNA. Adv. Drug. Deliv. Rev. 60(7):795-804.
7. Heinz L.X. et al., 2021. TASL is the SLC15A4-associated adaptor for IRF5 activation by TLR7-9. Nature. 581(7808):316-322.
8. Kayraklioglu N. et al., 2021. CpG oligonucleotides as vaccine adjuvants. DNA Vaccines: Methods and Protocols. Methods in Molecular Biology. Vol. 2197. p51-77.
9. Kumar V., 2021. The trinity of cGAS, TLR9, and ALRs: guardians of the cellular galaxy against host-derived self-DNA. Front. Immunol. 11:624597.
10. 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

ODN 1826 FITC

Technical Data Sheet

Safety Data Sheet

Validation Data Sheet

Certificate of analysis

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