Specifications

Antibiotic resistance: hygromycin B, Zeocin™

Growth medium: DMEM, 4.5 g/l glucose, 2 mM L-Glutamine, 10% (v/v) heat-inactivated fetal bovine serum (30 min at 56°C), 50 U/ml penicillin, 50 µg/ml streptomycin, 100 µg/ml Normocin™

Guaranteed mycoplasma-free

Shipped on dry ice

Detects human and murine IL-1β (α)
• Detection range for human IL-1β: 100 pg - 100 ng/ml
• Detection range for murine IL-1β: 10 ng - 1 µg/ml

Documents

• TDS HEK-Blue™ IL-1β cells : 3-7 x 10e6 (hkb-il1b)
• TDS HEK-Blue™ IL-1β kit : 1 kit (hkb-il1b-kit)
• MSDS HEK-Blue™ IL-1β cells : 3-7 x 10e6 (hkb-il1b)

Contents

• 1 vial containing 3-5 x 10⁶ cells

• 100 μl HygroGold™ (100 mg/ml)

• 100 μl Zeocin™ (100 mg/ml)

• 1 ml Normocin™ (50 mg/ml)

• 1 pouch of QUANTI-Blue™

Additionally provided with the HEK-Blue™  IL-1β kit:

• 100 μg anti-hIL1-β IgA antibody

Description

Interleukin-1β (IL-1β) is an important mediator of the inflammatory response to infection and injury. IL-1β binds to the type 1 IL-1 receptor (IL-1R) which requires the IL-1 receptor accessory protein (IL-1RAcP) to transduce a signal. IL-1β signaling is mediated by MyD88, IRAK-1/2 and TRAF-6 leading to the activation of NF-κB and AP-1.

Binding of IL-1β to its receptor IL-1R on the surface of HEK-Blue™ IL-1β cells triggers a signaling cascade leading to the activation NF-κB and the subsequent production of SEAP.

THP-1/HEK-Blue™ IL-1β Assay

1- Production of IL-1β by THP-1 cells: Typically, THP-1 cells are pretreated with phorbol 12-myristate acetate (PMA) to become more susceptible to inflammasome activators, then are primed with lipopolysaccharide (LPS). These treatments induce the production of pro-IL-1β, the immature form of IL-1β.
Subsequent stimulation with inflammasome inducers, such as crystals or ATP, leads to NRLP3 and caspase-1 activation resulting in IL-1β maturation and secretion.

2- Detection of IL-1β by HEK-Blue™ IL-1β cells: IL-1β-containing THP-1 supernatants are added to HEK-Blue™ IL-1β cells leading to NF-κB activation and the subsequent production of SEAP. The presence of SEAP in HEK-Blue™ IL-1β supernatants is assessed using QUANTI-Blue™, a SEAP detection medium.

Details

THP1 cells pretreated with PMA and primed with LPS (1 μg/ml) were stimulated with ATP (5 mM), nigericin (1 μM), alum (200 μg/ml), MSU (200 μg/ml) or CPPD (200 μg/ml). After 24h incubation, THP-1 supernatants or recombinant IL-1β (0.1 ng/ml) were added to HEK-Blue™ IL-1β cells. IL-1β - induced NF-kB activation was assessed by measuring the levels of SEAP in the supernatant of HEK-Blue™ IL-1β cells using the QUANTI-Blue™ assay.

Validation sheet for HEK-Blue™ IL-1β Cells (PDF)

Response of HEK-Blue™ IL-1β cells to IL-1β and TNF-α

Stimulation of HEK-Blue™ IL-1β cells by recombinant human and murine IL-1β and human TNF-α. After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the optical density (O.D.) at 655 nm.

Note: 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.

Cytokine response profile of HEK-Blue™ IL-1β cells

HEK-Blue™ IL-1β cells were stimulated with various human recombinant cytokines; IFNα (1000 IU/ml), IFNβ (1000 IU/ml), IFNγ (1000 IU/ml), IL-1β (100 ng/ml), IL-4 (100 ng/ml), IL-6 (100 ng/ml), IL-13 (100 ng/ml), IL-18 (100 ng/ml), TGF-β (10 ng/ml), and TNF-a (100 ng/ml). After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the O.D. at 655 nm.

Neutralization of IL-1β response

Anti-hIL-1β-IgA          IC50 = 50 +/- 20 ng/ml

Anti-hIL-1β-IgA was incubated with 1 ng/ml of hIL-1β for 30 minutes prior to the addition of HEK-Blue™ IL-1β cells, which were incubated with the antibody and cytokine for a further 24 h. Levels of SEAP in the supernatant were measured using QUANTI-Blue™ and reading the O.D. at 655 nm.