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Human NF-kB Reporter Assay Kit

1 x-96 well format assays
3 x-32 assays in-96 well format
1 x-384 well format assays
1 x-96 well format assays
3 x-32 assays in-96 well format
1 x-384 well format assays

Product Description and Product Data

This is an all-inclusive cell-based luciferase reporter assay kit targeting the Human Nuclear Factor kappa-light-chain enhancer of activated B cells (NF-kB). INDIGO’s NF-kB reporter assay utilizes proprietary mammalian cells that have been engineered to provide constitutive expression of NF-kB. In addition to NF-kB Reporter Cells, this kit provides two optimized media for use during cell culture and in diluting the user’s test samples, a reference agonist, Luciferase Detection Reagent, and a cell culture-ready assay plate. The principal application of this assay is in the screening of test samples to quantify any functional activity, either agonist or antagonist, that they may exert against human NF-kB. This kit provides researchers with clear, reproducible results, exceptional cell viability post-thaw, and consistent results lot to lot. Kits must be stored at -80C. Do not store in liquid nitrogen. Note: reporter cells cannot be refrozen or maintained in extended culture.


  • Clear, Reproducible Results

  • All-Inclusive Assay Systems
  • Exceptional Cell Viability Post-Thaw
  • Consistent Results Lot to Lot

Product Specifications

Target TypeTranscription Factor
Receptor FormNative
Assay ModeAgonist, Antagonist
Kit Components
  • NFkB Reporter Cells
  • Cell Recovery Medium (CRM)
  • Compound Screening Medium (CSM)
  • PMA, (ref. agonist; in DMSO)
  • Detection Substrate
  • Detection Buffer
  • White, sterile, cell-culture ready assay plate
Shelf Life6 months
Orthologs AvailableNo
Shipping RequirementsDry Ice
Storage temperature-80C


PMA and TNFa activation of Human NF-kB. Activation of NF-kB is demonstrated by treating reporter cells with (a.) Phorbol 12-myristate 13-acetate (PMA; provided), and (b.) TNFa (Tocris) for 22 hours, following the protocol for activation assays depicted in Figure 1. Average relative light units (RLU) and corresponding standard deviation (SD) values were determined for each treatment concentration (n ≥ 4). Fold-activation and Z’ values were calculated as described by Zhang, et al. (1999). Non-linear regression and EC50 analyses were performed using GraphPad Prism software. High Z' scores confirm the robust performance of this assay, and its suitability for HTS
Human NF-κB Reporter Cells were treated with ~EC80 of PMA and challenged with the inhibitors Chromomycin A3, Bortezomib, (S)-MG132, PS-1145 and BMS 345541 (all from Cayman Chemical). Reporter cells were treated for 6 hours, following the protocol for inhibition assays depicted in Figure 1.

Target Background

NF-kB is a signal transduction dependent transcription factor. INDIGO’s Human NF-kB Reporter Assay System utilizes proprietary human cells that express NF-kB (nuclear factor kappa-light-chain enhancer of activated B Cells), and contain the luciferase reporter gene functionally linked to upstream NF-kB genetic response elements. Thus, quantifying changes in luciferase expression provides a sensitive surrogate measure of changes in the level of NF-kB activation. This NF-kB reporter cell line is validated to provide a robust dose-dependent activation response when treated with TNFa, or the Protein Kinase C activator Phorbol 12-myristate 13-acetate (PMA). As such, the principle application of this assay system is in the screening of test samples to quantify any functional activities that they may exert to modulate, either induce or suppress, NF-kB activities.


The ongoing rise in antibiotic resistance, and a waning of the introduction of new antibiotics, has resulted in limited treatment options for bacterial infections, including these caused by methicillin-resistant Staphylococcus aureus, leaving the world in a post-antibiotic era. Here, we set out to examine mechanisms by which theaflavin 3,3’-digallate (TF3) might act as an anti-hemolytic compound. In the presented study, we found that TF3 has weak bacteriostatic and bactericidal effects on Staphylococcus aureus, and strong inhibitory effect towards the hemolytic activity of its α-hemolysin (Hla) including its production and secretion. A supportive SPR assay reinforced these results and further revealed binding of TF3 to Hla with KD = 4.57×10−5 M. Interestingly, TF3 was also able to protect human primary keratinocytes from Hla-induced cell death, being at the same time non-toxic for them. Further analysis of TF3 properties revealed that TF3 blocked Hla-prompting immune reaction by inhibiting production and secretion of IL1β, IL6, and TNFα in vitro and in vivo, through affecting NFκB activity. Additionally, we observed that TF3 also markedly attenuated S. aureus-induced barrier disruption, by inhibiting Hla-triggered E-cadherin and ZO-1 impairment. Overall, by blocking activity of Hla, TF3 subsequently subdued the inflammation and protected the epithelial barrier, which is considered as beneficial to relieving skin injury.
SARS-CoV-2 infection still poses health threats especially to older and immunocompromised individuals. New emerging variants of SARS-CoV-2, including Omicron and Arcturus, have been challenging the effectiveness of humoral immunity resulting from repeated vaccination and infection. With recent study implying a wave of new mutants in vaccinated people making them more susceptible to the newest variants and fueling a rapid viral evolution, there is a need for alternative or adjunct approaches against coronavirus infections other than vaccines. Our earlier work indicated that a specific combination of micronutrients and phytochemicals can inhibit key infection mechanisms shared by SARS-CoV-2 and its variants in vitro. Here we demonstrate in vivo that an intake of this micronutrient combination before and during infection of mice with engineered SARS-CoV-2 virions and HCoV-229E virus results in a significant decrease in viral load and level of spike protein in the lungs. This was accompanied by decreased inflammatory response, including TNFα, IL1β, ILα, and IL17. These and our earlier results confirm that by targeting multiple mechanisms simultaneously by a combination treatment we can effectively and safely challenge SARS-CoV-2 and HCoV-229E virus. If clinically confirmed, such an approach could complement already in-use preventive and therapeutic strategies against coronavirus infections.

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Nuclear Factor kappa-light-chain enhancer of activated B cells (NF-kB)

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