HEK-Blue™ CD40L Cells
|HEK-Blue™ CD40L cells||Unit size||Cat. code||Docs||Qty||Price|
Human CD40L SEAP Reporter Cells
3-7 x 10e6 cells
CD40L Reporter Cells
HEK-Blue™ CD40L cells can serve to measure the bioactivity of CD40L through the secretion of embryonic alkaline phosphatase (SEAP) upon NF-κB activation following CD40 stimulation.
These cells were generated by stable transfection of HEK293 cells with the human CD40 gene and an NF-κB-inducible SEAP construct.
Binding of CD40L to its receptor CD40 triggers a signaling cascade leading to the activation of NF-κB and the subsequent production of SEAP.
CD40L-CD40 interaction can be monitored by assessing the levels of SEAP using QUANTI-Blue™.
HEK293 cells express endogenously the receptors for the cytokines IL-1β and TNF-α which share a common signaling pathway with CD40L.
Consequently, HEK-Blue™ CD40L cells also respond to IL-1β and TNF-α. IL-1β- and TNF-α-mediated SEAP production can be blocked using neutralizing antibodies, such as anti-hIL-1β-IgG and anti-hTNF-α-IgA respectively.
Stimulation of HEK-Blue™ CD40L cells by recombinant human CD40L. After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the optical density (O.D.) at 655 nm.
|CD40||20 +/- 10 ng/ml||25|
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.
HEK-Blue™ CD40L 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), TNF-α (100 ng/ml) and CD40L (100 ng/ml). After a 24 h incubation, SEAP activity was assessed using QUANTI-Blue™ and reading the O.D. at 655 nm.
Anti-hCD40L-IgA was incubated with 5 ng/ml of hCD40L for 30 minutes prior to the addition of HEK-Blue™ CD40L 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.
Growth medium: DMEM, 4.5 g/l glucose, 2 mM L-glutamine, 10% (v/v) heat-inactivated fetal bovine serum, 100 U/ml penicillin, 100 µg/ml streptomycin, 100 µg/ml Normocin™
Screen for molecules that interfere with CD40L-CD40 cross-talk
Detection range for human CD40L: 5 ng - 1 μg/mlBack to the top
- 1 vial containing 3-7 x 106 cells
- 1 ml of Blasticidin (10 mg/ml)
- 1 ml of Zeocin™ (100 mg/ml)
- 1 ml Normocin™ (50 mg/ml)
- 1 ml of QB reagent and 1 ml of QB buffer (sufficient to prepare 100 ml of QUANTI-Blue™ Solution, a SEAP detection reagent)
Shipped on dry ice (Europe, USA & Canada)Back to the top
CD40 ligand (CD40L) is a member of the tumor necrosis factor (TNF) family of cell surface interaction molecules.
It is mainly expressed in CD4+ T cells and interacts with CD40 on antigen-presenting cells to regulate both humoral and cellular immune responses.
The CD40 cytoplasmic domain binds directly to several TNF receptor-associated factors (TRAFs), and this interaction is thought to initiate CD40 signaling.
CD40-mediated signaling results in NF-κB, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) activation.
In HEK-Blue™ CD40L cells, binding of CD40L to its receptor CD40 triggers a signaling cascade leading to the activation of NF-κB and the subsequent production of SEAP.Back to the top