RANKL Reporter HEK 293 Cells

HEK-Blue™ RANKL Cells Unit size Cat. code Docs Qty Price
Human & Mouse RANKL Reporter Cells
3-7 x 10e6 cells

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RANKL Reporter Cells

HEK-Blue RANKL Cells signaling pathway
HEK-Blue™ RANKL Cells signaling pathway

HEK-Blue™ RANKL cells were engineered from the human embryonic kidney HEK293 cell line to detect bioactive human and murine RANKL by monitoring the activation of the NF-κB and AP-1 pathways. In addition, these cells can be used for screening anti-RANKL antibodies, or inhibitors targeting the RANKL/RANK signaling pathway.

The tumor necrosis factor (TNF)-family member Receptor Activator of NF-κB Ligand (RANKL), also known as member 11 of the TNF superfamily (TNFSF11) or TNF-related activation-induced cytokine (TRANCE), exists as a soluble or transmembrane protein produced by osteoblasts and activated T cells [1]. RANKL binding to its receptor RANK and the subsequent signaling events play a pivotal role in bone remodeling and dendritic cell survival, thereby enhancing the induction of T cell responses [1].

RANKL binds to RANK via an obligate trimer configuration [1, 2]. Upon RANKL binding, RANK trimers recruit TNF receptor-associated factor (TRAF) adaptor proteins, such as TRAF6, to TRAF-binding motifs within their cytoplasmic domains [1]. The TRAF6 signaling cascade results in the activation of NF-κB and AP-1 transcription factors. Multiple efforts have focused on the development of anti‑RANKL antibodies or small‑molecule inhibitors for blocking RANKL/RANK signaling to reduce osteoporosis, prevent skeletal-related events (SREs) from bone metastasis in cancer, or improve anti-tumor immunity [1-3]. 

HEK-Blue™ RANKL cells were generated by stable transfection of HEK293 cells with the human gene for RANK (with all three functional TRAF-binding motifs) and an NF-κB/AP1-inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene. Binding of RANKL to its receptor on the surface of HEK-Blue™ RANKL cells triggers a signaling cascade leading to the activation of NF-κB/AP1, and the subsequent production of SEAP.  For your convenience, InvivoGen provides QUANTI‑Blue™ Solution, an easy and rapid means to detect and quantify SEAP activity in the cell culture supernatant.  


Features of HEK-Blue™ RANKL cells:

  • Fully functional RANKL signaling pathway
  • Readily assessable SEAP reporter activity
  • Functionally tested
  • Guaranteed mycoplasma-free

Applications of HEK-Blue™ RANKL cells:

  • Detection of both human and mouse RANKL
  • Screening of anti-RANKL antibodies
  • Screening of inhibitors targeting RANKL/RANK signaling



1. Cheng ML. & Fong L.,  2014. Effects of RANKL-targeted therapy in immunity and cancer. Front. Oncol. 3:329.
2. Ahern al., 2018. Roles of the RANKL-RANK axis in anti-tumour immunity — implications for therapy. Nat. Rev. Clin. Oncol. 15:676-93.
3. Nakai al., 2019. Efficacy of an orally active small-molecule inhibitor of RANKL in bone metastasis. Bone Res. 7:1.


Evaluation of RANKL response in HEK-Blue™ RANKL Cells
Evaluation of RANKL response in HEK-Blue™ RANKL Cells

Dose-response of HEK-Blue™ RANKL cells to recombinant RANKL.

Cells were stimulated with increasing concentrations of recombinant human or murine RANKL. After overnight incubation, the NF-κB response was determined using QUANTI‑Blue™ Solution, a SEAP detection reagent, and reading the optical density (OD) at 630 nm. EC50 values are shown as mean ± SD.

Cytokine and TLR cellular responses
Cytokine and TLR cellular responses

Cytokine response of HEK-Blue™ RANKL cells.

Cells were stimulated with various human and murine recombinant cytokines: 10 ng/ml of hRANKL, mRANKL, hIL‑1β, mIL‑1β, 1x104 IU/ml of hIFN-α, hIFN-β, 100 ng/ml of hIFN-γ, hTNF-α, and 1 µg/ml of the TLR3 ligand Poly(I:C) HMW and the TLR5 ligand FLA-ST (flagellin from S. typhimurium). After overnight incubation, SEAP activity was assessed using QUANTI‑Blue™ Solution. 

Evaluation of RANKL/RANK signaling inhibition
Evaluation of RANKL/RANK signaling inhibition

Dose-dependent inhibition of HEK‑Blue™ RANKL cellular response using an anti-hRANKL antibody.

The antibody was incubated with HEK‑Blue™ RANKL cells  for 30 minutes prior to the addition of 10 ng/ml of hRANKL. After overnight incubation, SEAP activity in the cell culture supernatant was assessed using QUANTI‑Blue™ Solution. Data represent % inhibition of reporter activity without anti-hRANKL antibody.

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Antibiotic resistance: Blasticidin,  Zeocin™

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

Guaranteed mycoplasma-free

Specificity: human and mouse RANKL

Detection range:

  • 3 - 100 ng/ml for human RANKL
  • 1 - 100 ng/ml for murine RANKL


These cells are covered by a Limited Use License (See Terms and Conditions).

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Shipped on dry ice (Europe, USA & Canada)

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