TransAM® NFκB Activation Assays | Colorimetric Kits

DNA-binding ELISA kits for studying activity of NFkB family members p50, p52, p65, c-Rel, and RelB

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NFκB Introduction & Background

Nuclear Factor κB (NFκB) is an important transcription factor complex that plays critical roles in regulating the expression of many genes in cells, including pathways that control the immune response, inflammation, development, cellular growth, and apoptosis. When NFκB is misregulated it can lead to many different disease states, including multiple types of cancer, neurodegenerative diseases, heart diseases, and inflammation. These disorders are usually due to overactive NFκB complexes, which highlights the importance of being able quickly, accurately, and reliably measure its activity in samples.
 

Using TransAM DNA-Binding ELISAs to Measure NFκB Activity

TransAM NFκB kits are DNA-binding ELISAs that provide everything needed to study NFκB in biological samples, including a positive control extract. These assays are available with antibodies specific for the activated form of p50, p52, and p65. A family kit is also available with the ability to screen the members of the NFκB family (p50, p52, p65, c-Rel, and RelB).
 
The kit manual and other important documents can be downloaded from the Documents tab below.
 
Recombinant p50 and Recombinant p65 proteins are available separately to generate an optional protein standard curve in the TransAM NFκB Transcription Factor ELISA kits.
 
Flow chart of the TransAM DNA-binding transcription factor ELISA

 

Figure 1. Flow chart of the TransAM process. Activated transcription factor in the cell extract binds to the consensus-binding site on the oligo immobilized in the well. Incubation with the supplied primary and secondary antibodies specifically quantifies the amount of activated transcription factor.

Read more about the TransAM transcription factor activity assay system >>

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TransAM NF-kB Family Kit Data

TransAM NFκB Assay is More Sensitive than a Gel Shift

TransAM NFkB data vs. gel shift
 
Figure 2. TransAM NFκB is more sensitive than gelshift. Human fibroblast WI-38 cells are stimulated with IL-1α for 30 minutes. Increasing amounts of whole-cell extract are assayed using (A) the TransAM NFκB p50 Kit or (B) a gel shift assay.

  

NFκB Family Profiling in Multiple Cell Lines

NFkB family profiling of DNA binding activation in various cell lines
 
Figure 3. Nuclear extracts prepared from HeLa, HeLa treated with TNF-α, Jurkat, Jurkat treated with PMA and calcium ionophore, and Raji cells were assayed at 10 µg/well for p65, p50, c-Rel, p52 and RelB activity using the TransAM NFκB Family Kit.
 

TransAM NF-kB Family Kit Contents & Storage

Kit components: One, two or five NFκB 96-well assay plate(s) with plate sealer(s), NFκB p50, p52 or p65 or p50, p52, p65, c-Rel and RelB primary antibody(ies), HRP-conjugated secondary antibody, NFκB wild-type and mutated oligonucleotides, positive control cell extract, DTT, Protease Inhibitor Cocktail, Herring Sperm DNA, Lysis, Binding, 10X Washing and 10X Antibody Binding Buffers, and Developing & Stop Solutions.

Kit storage: Reagent storage conditions vary from room temperature to -80°C, see manual for details. All reagents are guaranteed stable for 6 months when stored properly.

NOTE: the p50 and p52 antibodies can be used with human and mouse extracts, while the p65 and RelB antibodies can be used with human, mouse, and rat extracts. The c-Rel antibody can only be used with human extracts.

Technology covered under EAT-filed patents and licensed to Active Motif. Use of TransAM in NFκB-related drug discovery may be covered under U.S. Patent No. 6,150,090 and require a license from Ariad Pharmaceuticals (Cambridge, MA, USA).

TransAM promotion
 
Name Format Cat No. Price  
TransAM® NFκB Family 2 x 96 rxns 43296 $1,220 Buy
TransAM® NFκB p50 1 x 96 rxns 41096 $725 Buy
5 x 96 rxns 41596 $2,980 Buy
TransAM® NFκB p52 1 x 96 rxns 48196 $725 Buy
5 x 96 rxns 48696 $2,980 Buy
TransAM® NFκB p65 1 x 96 rxns 40096 $725 Buy
5 x 96 rxns 40596 $2,980 Buy
TransAM NFκB Family: Catalog Number 43296
TransAM NFκB p50: Catalog Number 41096
TransAM NFκB p50: Catalog Number 41596
TransAM NFκB p52: Catalog Number 48196
TransAM NFκB p52: Catalog Number 48696
TransAM NFκB p65: Catalog Number 40096
TransAM NFκB p65: Catalog Number 40596