LPS-EB Biotin

Biotinylated LPS from Escherichia coli 0111:B4

LPS-EB Biotin is biotinylated ultrapure smooth lipopolysaccharide (sLPS) from the Gram-negative bacteria E. coli  0111:B4.

LPS is recognized by toll-like receptor 4 (TLR4) which interacts with three different extracellular proteins: LPS binding protein (LBP), CD14, and, myeloid differentiation protein 2 (MD-2), to induce a signaling cascade leading to the activation of NF-κB and the production of proinflammatory cytokines [1, 2]. 

Most LPS preparations on the market are contaminated by other bacterial components, such as lipoproteins, thus activating TLR2 signaling as well as TLR4 signaling. The ultrapure preparation that InvivoGen provides only activates the TLR4 pathway.

In this product, biotin has been bound to LPS’s oxidized carbohydrates using biotin-LC-hydrazide as previously described [3]. The labeled ligand retains the biological activity of LPS. LPS‑EB Biotin can be used for the detection of LPS binding sites by cytometry or fluorescence microscopy.  

Key features of LPS-EB Biotin:

• Highly specific and potent TLR4 agonist
• Ultrapure grade of LPS that only activates TLR4
• Each lot is functionally tested

   

References:

1. Kuzmich, N.N. et al., 2017. TLR4 signaling pathway modulators as potential therapeutics in inflammation and sepsis. Vaccines (Basel) 5(4):34.
2. Hirschfeld M. et al., 2000. Cutting edge: repurification of lipopolysaccharide eliminates signaling through both human and murine toll-like receptor 2. J Immunol. 165(2):618-22.
3. Luk J.M. et al., 1995. Biotinylated lipopolysaccharide binds to endotoxin receptor in endothelial and monocytic cells. Anal Biochem 232:217.

Learn more about TLR4 and its co-receptors

Figures

Intracellular detection of LPS-EB Biotin. RAW-Blue™ cells were incubated with LPS-EB Biotin for 16h - 24h. After, the cells were fixed and permeabilized, following the incubation with an Anti-Biotin-PE secondary antibody. The intracellular staining was analyzed by flow cytometry.

LPS-EB Biotin is a potent activator of human (h)TLR4. HEK-Blue™ hTLR2 and HEK-Blue™ hTLR4 cells were incubated with increasing concentrations of LPS-EB Biotin. After overnight incubation in HEK-Blue™ detection medium, a SEAP detection growth medium, the response of hTLR2 and hTLR4 was assessed by determining the presence of SEAP in the supernatant. Data are expressed as optical density at 630 nm (±SEM).

Specifications

Specificity: TLR4 agonist

Working Concentration: 10 ng - 10 μg/ml

Endotoxin Level:  ≥1 x 10⁵ EU/mg (measurement with HEK-Blue™  LPS Detection Kit 2)

Quality Control

• Activation of TLR4 by LPS-EB Biotin has been confirmed using a HEK-Blue™ TLR4 cellular assay.
• Absence of bacterial contamination (e.g. lipoproteins) has been confirmed using a HEK-Blue™ TLR2 cellular assay.
• Biotin coupling has been validated by flow cytometry.

Contents

• 500 μg biotinylated ultrapure E. coli O111:B4 LPS (LPS-EB Biotin)
• 1.5 ml endotoxin-free water

LPS-EB Biotin is shipped at room temperature

Stored at -20°C.