Our Products
- Same product category
- Order HEK-Blue™ hTLR4 cells
HEK-Blue™ TLR4 cells
InvivoGen introduces HEK-Blue™ human TLR4 cells, part of a collection of engineered HEK 293 cell lines designed to provide a rapid, sensitive and reliable method to screen and validate TLR agonists or antagonists.
HEK-Blue™-hTLR4 cells are designed for studying the stimulation of human TLR4 (hTLR4) by monitoring the activation of NF-kB.
HEK-Blue™-hTLR4 cells were obtained by co-transfection of the hTLR4 and MD-2/CD14 co-receptor genes and an NF-κB-inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene.
These HEK 293 cells were thoroughly tested and validated by InvivoGen. The following data were obtained using the QUANTI-Blue™ or HEK-Blue™ Detection assays. These assays allow the detection of SEAP production following TLR/NOD activation by reading the OD at 655 nm. Performance of these assays was validated under optimized conditions.
The parental cell line of HEK-Blue™ hTLR4 cells is HEK-Blue™ Null2 that express the SEAP reporter gene under the control of the IL-12 p40 minimal promoter fused to five NF-κB binding sites.
Validation sheet for HEK-Blue™ hTLR4 Cells
TLR/NOD Induction
Response of HEK-Blue™ hTLR4 cells to TLR and NOD agonists
HEK-Blue™ hTLR4 cells were stimulated with various TLR and NOD agonists: Pam3CSK4 (100 ng/ml), Poly(I:C) (50 ng/ml), LPS-EB ultrapure (100 ng/ml), recombinant flagellin from S. typhimurium (10 ng/ml), CL264 (1 μg/ml), CL097 (3 μg/ml), ssRNA40/LyoVec™ (5 μg/ml), ODN 2006 (10 μg/ml), C12-iE-DAP (100 ng/ml ), L18-MDP (100 ng/ml ), and TNF-α (100 ng/ml ). After 18h incubation (24h incubation for CL264, C12-iE-DAP and L18-MDP ligands), NF-κB-induced SEAP activity was assessed using QUANTI-Blue™.
Response of HEK-Blue™ hTLR4 cells to TLR4 agonists
HEK-Blue™ hTLR4 and HEK-Blue™ Null2 (control) cells were incubated in HEK-Blue™ Detection medium and stimulated with 10 ng/ml LPS-EB ultrapure, 10 ng/ml LPS-EK ultrapure, 100 ng/ml MPLA or 100 ng/ml MPLAs. After 24h incubation, the levels of NF-κB-induced SEAP were determined by reading the OD at 655 nm.
TLR4 agonists dose response
LPS-EK Ultrapure (from E.coli K12) EC50 = 580 +/- 40 pg/ml
LPS-EB Ultrapure (from E.coli O111:B4) EC50 = 2.8 +/- 1 ng/ml
MPLAs (Synthetic MPLA) EC50 = 15.3 +/- 0.6 ng/ml
Response Ratio = 7.3
Optimum cell number = 25,000 cells/well in a 96-well plate.
Dose response of HEK-Blue™ hTLR4 to TLR4 agonists.
After 18 h incubation with TLR4 agonists, NF-κB-induced SEAP activity was assessed using QUANTI-Blue™, a SEAP-detection medium.
TLR4 inhibition
TLR4 antagonists dose response
LPS-RS (LPS from R. sphaeroides) IC50 7.9 +/- 1 ng/ml
Inhibition Ratio = 4
Optimum cell number = 25,000 cells/well in a 96-well plate
Agonist LPS-EB Ultrapure conc = 10 ng/ml
HEK-Blue™ TLR4 cells were pre-incubated for 6h with increasing concentrations of LPS-RS, then stimulated with 10 ng/ml of LPS-EB Ultrapure for 18h. NF-κB-induced SEAP activity was assessed using QUANTI-Blue™.
Neutralization of TLR4 response
Anti-hTLR4 IgA (neutralizing antibody) IC50 136 +/- 15 ng/ml
Inhibition Ratio = 3.5
Optimum cell number = 25,000 cells/well in a 96-well plate
Agonist LPS-EB Ultrapure conc = 10 ng/ml
HEK-Blue™ TLR4 cells were pre-incubated for 6h with anti-hTLR4 IgA, then stimulated with 10 ng/ml of LPS-EB Ultrapure for 18h. NF-κB-induced SEAP activity was assessed using QUANTI-Blue™.
Content and storage
Cells are provided frozen in a cryotube containing 5-7 x 106 cells and supplied with the corresponding antibiotics, 1 ml Normocin™ (50 mg/ml)and 1 pouch of QUANTI-Blue™.
Cells are shipped on dry ice.
| Product | Description | TDS | MSDS | Qty | Cat. Code | Price | Order |
|---|---|---|---|---|---|---|---|
| HEK-Blue™ hTLR4 cells | human TLR4-expressing HEK 293 cells | 5-7 x 10e6 cells | hkb-htlr4 | check |  |