NF-Kappa B p65 DNA-Binding ELISA Kit (OKAG00423)

Catalog No: OKAG00423

Size:12x8WELLSTRIPS

NF-κB p65 DNA-Binding ELISA detects active NF-κB p65 in HepG2 Nuclear Extract. The HepG2 cells were grown 3 days in DMEM with 10% FBS and harvested for nuclear extract.

Product Info

• Predicted Species Reactivity: Human, Mouse, Rat
• Product Format: 12 x 8-Well Microstrips
• Application: Enzyme-linked immunosorbent assay
• ELISA Kit Detection Method: Colorimetric 450 nm
• ELISA Kit Principle: The Aviva DNA-Binding ELISA Kit contains components necessary for detection of active transcription factors in eukaryotic nuclear or cell lysates. This particular immunoassay utilizes the qualitative technique of an indirect ELISA. Streptavidin is bound to the immunoassay plate and specific biotinylated double-stranded (dsDNA) oligonucleotides are then added to bind to the streptavidin via a high affinity biotin-streptavidin interaction. After subsequent blocking of extraneous binding sites in each well, the sample containing the target of interest can be added. Primary antibody is added to bind activated transcription factors bound to the dsDNA oligonucleotide, which has been immobilized via the plate-coated streptavidin. A HRP-conjugated secondary antibody specific for rabbit IgGs is added, which allows for specific binding to the Primary Antibody, and consequently colorimetric detection upon addition of the TMB substrate.
  For color development, TMB (3, 3’, 5, 5’-Tetramethylbenzidine) is added to each well. After addition of the substrate, a peroxidase catalyzed reaction will produce a blue TMB Diimine product that is proportional to the target concentration in the sample. Color development is quenched by addition of Stop Solution, or 2 N Sulfuric Acid, which turns the solution yellow. The absorbance can then be read by a spectrophotometer at 450 nm and subsequently allowing for determination of the target concentration in the sample.
  
• ELISA Kit Component:
  • dsDNA Oligonucleotide Coated Microplate: 96 Wells (12 x 8 Strips)Microstrips
  • 100X Primary Antibody: 100 uL
  • 100X Primary Phospho-Antibody: 100 uL
  • HRP-Conjugated Anti-Rabbit IgG Antibody: 2 x 6 mL
  • Nuclear Lysate Positive Control: Lyophilized
  • Wild-Type Consensus dsDNA Oligonucleotide: 20 uL
  • Mutant Consensus dsDNA Oligonucleotide: 20 uL
  • 10X Wash Buffer: 50 mL
  • 2X Binding Buffer: 12 mL
  • Primary Antibody Diluent: 12 mL
  • 100X Protease and Phosphatase Inhibitors: 100 uL
  • Nuclear Wash Buffer: 12 mL
  • Cytoplasmic Extraction Buffer: 6 mL
  • Nuclear Extraction Buffer: 6 mL
  • Ready-to-Use Substrate: 12 mL
  • Stop Solution: 12 mL
  
  OMIM: 164014
  UniGene: Hs.502875
  Note: The product is for research use only, not for use in diagnostic or therapeutic procedures.
• Reconstitution and Storage: Store as indicated in product manual.
• Specificity: The NF-kappaB p65 DNA-Binding ELISA Kit detects endogenous levels of total NF-kappaB p65 protein.
• Assay Info: Assay Type: Qualitative DNA Binding ELISA

Target Info

• Gene Symbol: RELA
• Gene Full Name: RELA proto-oncogene, NF-kB subunit
• Alias Symbols: p65, CMCU, NFKB3
• NCBI Gene Id: 5970
• Protein Name: transcription factor p65
• Description of Target: NF-kappa-B is a ubiquitous transcription factor involved in several biological processes. It is held in the cytoplasm in an inactive state by specific inhibitors. Upon degradation of the inhibitor, NF-kappa-B moves to the nucleus and activates transcription of specific genes. NF-kappa-B is composed of NFKB1 or NFKB2 bound to either REL, RELA, or RELB. The most abundant form of NF-kappa-B is NFKB1 complexed with the product of this gene, RELA. Four transcript variants encoding different isoforms have been found for this gene.
• Uniprot ID: Q04206