Western blot analysis of extracts from THP-1 cells, differentiated with TPA #4174 (80 nM for 24 hr) and treated with 1 μg/ml LPS for the indicated times, using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb #3033 (upper) and NF-κB p65 (C22B4) Rabbit mAb #4764 (lower).
Western blot analysis of extracts from THP-1 cells, untreated (-) or LPS-treated (100 ng/ml, 3 hr; +), using IL-1β (3A6) Mouse mAb #12242 (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
For a 1 mg/ml stock, reconstitute the 10 mg in 10 ml sterile PBS. Working concentrations and length of treatment can vary depending on the desired effect, but it is typically used at 10-1000 ng/ml for 15 min-24 hours. Soluble in PBS and cell culture medium at 5 mg/ml and 1 mg/ml, respectively.
Store lyophilized at 4ºC. In lyophilized form, the product is stable for 24 months. Once in solution store at -80ºC and use within 3 months to prevent loss of potency. Aliquot to avoid multiple freeze/thaw cycles.
LPS is supplied as a lyophilized powder and is from E. coli serotype O111:B4. It is purified via phenol extraction.
Lipopolysaccharide (LPS), also known as endotoxin, is a major glycolipid constituent of the outer cell wall of gram-negative bacteria. LPS molecules typically consist of a strain-specific distal polysaccharide side chain known as the O-antigen, a hydrophilic core oligosaccharide, and a hydrophobic domain referred to as lipid A. Lipid A is covalently bound to the outer bacterial membrane and is responsible for the toxicity of LPS (1-3). LPS is a potent activator of the proinflammatory response in many mammalian cell types, including macrophages, monocytes, and endothelial cells. Investigators have demonstrated that LPS binds to the CD14/TLR4/MD2 receptor complex, which in turn induces inflammatory cytokines including TNF-α, Interleukin-1, and IFN-α, as well as numerous inflammatory proteins such as iNOS, NF-κB, RIG-1, and IRF-3 (4-6).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.