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HEK-Blue™ hTLR4

HEK-Blue™ hTLR4 cells Unit size Cat. code Docs Qty Price
human TLR4-expressing HEK 293 cells
3-7 x 10e6 cells
hkb-htlr4
+-
$1,142.00

HEK-Blue™-hTLR4 cells were obtained by co-transfection of the human TLR4, MD-2 and CD14 co-receptor genes, and an inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene into HEK293 cells.

The SEAP reporter gene is placed under the control of an IL-12 p40 minimal promoter fused to five NF-κB and AP-1-binding sites.

Stimulation with a TLR4 ligand activates NF-κB and AP-1 which induce the production of SEAP.

Levels of SEAP can be easily determined with HEK-Blue™ Detection, a cell culture medium that allows for real-time detection of SEAP.

The parental cell line of HEK-Blue™ hTLR4 cells is HEK-Blue™ Null2 cells.

TLR/NOD Induction Response of HEK-Blue™ hTLR4 cells to TLR and NOD agonists
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™and by reading the OD at 655 nm.

Response of HEK-Blue™ hTLR4 cells to TLR4 agonists
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
TLR4 agonists dose response

HEK-Blue™ hTLR4 cells were incubated in HEK-Blue™ Detection medium and stimulated with increasing concentrations of TLR4 agonists. After 24h incubation, the levels of NF-kB-induced SEAP were determined by reading the OD at 655 nm. The response ratio was calculated by dividing the OD at 655 nm for the treated cells by the OD at 655 nm for the untreated cells.

Ligand EC50 Response Ratio
 LPS-EB up 2 +/- 1 ng/ml 13
LPS-EK 0.07 +/- 0.01 ng/ml  16
MPLAs 10 +/- 3 ng/ml 12
Neutralization of TLR4 dose response
Neutralization of TLR4 dose response

HEK-Blue™ TLR4 cells were pre-incubated for 1h with increasing concentrations of anti-hTLR4 IgA2, then stimulated with 5 ng/ml of LPS-EB Ultrapure for 18h. NF-kB-induced SEAP activity was assessed using QUANTI-Blue™ by reading the OD at 655 nm.

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Specifications

Antibiotic resistance: blasticidin, hygromycin, Zeocin™

Growth medium: DMEM, 4.5 g/l glucose, 2-4 mM L-glutamine, 10% (v/v) fetal bovine serum, 50 U/ml penicillin, 50 μg/ml streptomycin, 100 μg/ml Normocin™

Guaranteed mycoplasma-free.

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Contents

1 vial containing 3-7 x 106 cells

Shipped on dry ice.

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Citations

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