NFYA DNA-Binding ELISA Kit

Root Catalog/Products/ELISA Kits, Root Catalog/Products/ELISA Kits/Transcription Factor DNA Binding

NFYA

12 x 8-Well Microstrips

The protein encoded by this gene is one subunit of a trimeric complex, forming a highly conserved transcription factor that binds to CCAAT motifs in the promoter regions in a variety of genes. Subunit A associates with a tight dimer composed of the B and C subunits, resulting in a trimer that binds to DNA with high specificity and affinity. The sequence specific interactions of the complex are made by the A subunit, suggesting a role as the regulatory subunit. In addition, there is evidence of post-transcriptional regulation in this gene product, either by protein degradation or control of translation. Further regulation is represented by alternative splicing in the glutamine-rich activation domain, with clear tissue-specific preferences for the two isoforms.

The NFYA DNA-Binding ELISA Kit detects endogenous levels of total NFYA protein.

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.