ssRNA40/LyoVec™

Activation of TLR7/8 by ssRNA40/LyoVec™

TLR7/8 Agonist - HIV-1-derived ssRNA - complexed with LyoVec™

ssRNA40 is a 20-mer phosphorothioate-protected single-stranded RNA oligonucleotide containing a GU-rich sequence. ssRNA40, also known as R-1075, is a U-rich single-stranded RNA derived from the long terminal repeat of HIV-1.

ssRNA derived from HIV-1 or the influenza virus were shown to induce the production of proinflammatory cytokines by activation of the Toll-like receptors TLR7 and/or TLR8.

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Description

InvivoGen's ssRNA40/LyoVec™ is complexed with the cationic lipid LyoVec™ to protect it from degradation and facilitate its uptake. Moreover, phosphorothioate linkages were incorporated in order to extend the effective molecular lifetime by minimizing extra and intracellular nuclease degradation. ssRNA40/LyoVec™ is a weak TLR7 and strong TLR8 agonist, when tested using our HEK-Blue™ reporter cell lines expressing human or mouse TLR7 or TLR8 (see figure). 

Key features of ssRNA40/LyoVec™

• Activator of TLR7 and TLR8
• Negative control ssRNA41/LyoVec™ is available
• Complexed with LyoVec™ 
• Each lot of ssRNA40/LyoVec™ is functionally tested

Read our review about TLR7 and TLR8.

Figures

NF-κB response of HEK-Blue™-derived cells to ssRNA40/LyoVec™. HEK-Blue™ cells expressing hTLR7, mTLR7, or hTLR8 cultured in HEK-Blue™ Detection reagent and stimulated with increasing concentrations of ssRNA40/LyoVec™. After 24h incubation, the NF-κB-induced SEAP activity was assessed by measuring the SEAP level in the supernatant. Data are shown as optical density (OD) at 650 nm (mean ± SEM). Of note, HEK-Blue™ Null* comprises data from parental cell lines HEK-Blue Null1, HEK-Blue Null1-v, HEK-Blue Null2-k.

Specifications

Specificity: human TLR8/mouse TLR7 agonist

Working Concentration: 0.25-5 μg/ml

Sequence: ssRNA40 5’-G*C*C*C*G*U*C*U*G*U*U*G*U*G*U*G*A*C*U*C-3’
(“*” depicts the phosphorothioate linkage)

Solubility: 0.05 mg/ml in water

Quality control:

• The biological activity of ssRNA40/LyoVec™ has been validated using cellular assays.
• The absence of bacterial contamination (e.g. lipoproteins and endotoxins) has been confirmed using HEK-Blue™ hTLR2 and HEK-Blue™ hTLR4 cells.

Contents

• 4x 25 μg lyophilized ssRNA40/LyoVec™ 1:2 ratio (w/w)

Note: Each vial contains 25 μg of ssRNA40 complexed with 50 μg LyoVec™.

• 10 ml endotoxin-free water

ssRNA40/LyoVec™ is provided lyophilized and shipped at room temperature.

Store at -20 ̊C.

Lyophilized product is stable 1 year at -20 ̊C.

Upon resuspension, store product at 4°C. Resuspended product is stable 1 week at 4°C.

Details

ssRNA40 is a 20-mer phosphothioate-protected single-stranded RNA oligonucleotide containing a GU-rich sequence [1].

ssRNA40 is complexed with the cationic lipid LyoVec™, to protect it from degradation and facilitate its uptake, and lyophilized to generate ssRNA40/LyoVec™. When complexed to cationic lipids, ssRNA can substitute for viral RNAs in inducing TNF-α and IFN-α production in peripheral blood mononuclear cells [1, 2].

Murine dendritic cells deficient for TLR7 failed to produce IFN-α in response to ssRNA40. In contrast, the response to CpG-ODNs was unaffected, suggesting that TLR7 plays a critical role in viral ssRNA recognition [1].

In human cells, TLR8 was shown to be the key receptor for viral ssRNA, implying a species specificity difference in ssRNA recognition.

During infection, some viral particles are degraded by the endosomal proteases, exposing the viral genome and allowing TLR7 and/or TLR8 signaling, known to occur in endosomes [3].

TLR7 and TLR8 can recognize both self and viral RNA but can distinguish the presence of viral RNA by detecting their abnormal localization in the endosome rather than a particular RNA motif.

1. Heil F. et al., 2004. Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8. Science. 5;303(5663):1526-9.
2. Diebold SS. et al., 2004. Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA. Science. 5;303(5663):1529-31
3. Heil F. et al., 2003 The Toll-like receptor 7 (TLR7)-specific stimulus loxoribine uncovers a strong relationship within the TLR7, 8 and 9 subfamily. Eur J Immunol. 33(11):2987-97.