Pam3CSK4 (TLR1/2) and LPS (TLR4) were obtained from InvivoGen (Carlsbad, CA, USA). Mouse recombinant IFN-β was purchased from PBL InterferonSource (now PBL Assay Science, Piscataway, NJ, USA) and mouse IFN-γ was from R&D Systems (Minneapolis, MN, USA). Antibodies used in this study were obtained as follows. Anti-H-PGDS and Anti-COX-2 antibodies were produced by Cayman Chemical (Ann Arbor, MI, USA). Anti-STAT1 and anti-pY701-STAT1 antibodies were from Cell Signaling Technology (Danvers, MA, USA). Anti-actin antibody was from BD Biosciences (Franklin Lakes, NJ, USA). PGs were purchased from Cayman Chemical (Ann Arbor, MI, USA). All other chemicals were from Sigma (St. Louis, MO, USA), unless stated otherwise.
Lps tlr4
Manufactured by InvivoGen
Sourced in United States
LPS (TLR4) is a laboratory product that functions as a ligand for the Toll-like receptor 4 (TLR4). TLR4 is a pattern recognition receptor that binds to lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria. The interaction between LPS and TLR4 is a key step in the innate immune response to bacterial infection.
Automatically generated - may contain errors
Lab products found in correlation
Anti stat1, by Cell Signaling Technology (1 mentions)
Anti py701 stat1, by Cell Signaling Technology (1 mentions)
Pam3csk4 tlr1 2, by InvivoGen (1 mentions)
Anti h pgds, by Cayman Chemical (1 mentions)
Anti cox 2 antibody, by Cayman Chemical (1 mentions)
Mouse ifn γ, by R&D Systems (1 mentions)
Anti actin antibody, by BD (1 mentions)
Brefeldin a, by Merck Group (1 mentions)
Facs lysing solution, by BD (1 mentions)
2 protocols using lps tlr4
1
Toll-like Receptor Signaling in Inflammation
2
Standardized Cytokine Profiling Assay
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To minimize technical artifacts, we used a highly standardized, stringently controlled protocol as described previously (50) . Briefly, premade 96-well plates contained the following specific PRR ligands were prepared: PAM3CSK4 (PAM; TLR2/1; InvivoGen) at 1 g/ml; LPS (TLR4; InvivoGen) at 10 ng/ml; R848 (TLR7/8; InvivoGen) at 10 M and media alone. Whole blood, diluted 1:1 with sterile prewarmed RPMI 1640, was added to each well containing the specific TLR ligands.
For the intracellular cytokine staining (ICS), 10 µg/ml of brefeldin A (final concentration -Sigma-Aldrich) was added to each well, and samples were incubated for 6h at 37°C in 5% CO2, then treated with 2 mM EDTA (final concentration) for 10 min at 37°C. The cells were collected and resuspended in BD FACS Lysing Solution, placed into fresh tubes, and stored at -80°C.
For multiplex analysis, samples were incubated with whole blood for 24h at 37°C in 5% CO2.
The supernatant was collected and stored at -80°C.
For the intracellular cytokine staining (ICS), 10 µg/ml of brefeldin A (final concentration -Sigma-Aldrich) was added to each well, and samples were incubated for 6h at 37°C in 5% CO2, then treated with 2 mM EDTA (final concentration) for 10 min at 37°C. The cells were collected and resuspended in BD FACS Lysing Solution, placed into fresh tubes, and stored at -80°C.
For multiplex analysis, samples were incubated with whole blood for 24h at 37°C in 5% CO2.
The supernatant was collected and stored at -80°C.
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