Description
Key facts
Detection method:Colorimetric,
Sample types:Nuclear Extracts,
Reacts with:Mouse, Rat, Human,
Assay type:Semi-quantitative,
Sensitivity:> 600 ng/well,
Assay time:3h 30m,
Assay Platform:Microplate reader
Product details:
Nrf2 Transcription Factor Assay Kit (Colorimetric) (ab207223) is a high throughput assay to quantify Nrf2 activation in nuclear extracts. This assay combines a quick ELISA format with a sensitive and specific non-radioactive assay for transcription factor activation.
A specific double stranded DNA sequence containing the Nrf2 consensus binding site (5'-GTCACAGTGACTCAGCAGAATCTG-3') has been immobilized onto a 96-well plate. Active Nrf2 present in the nuclear extract specifically binds to the oligonucleotide. Nrf2 is detected by a primary antibody that recognizes an epitope of Nrf2 accessible only when the protein is activated and bound to its target DNA. An HRP-conjugated secondary antibody provides sensitive colorimetric readout at OD 450 nm. This product detects only human, mouse and rat Nrf2.
Key performance and benefits:
Assay time: 3.5 hours (cell extracts preparation not included).
Detection limit: < 0.6 ug nuclear extract/well.
Detection range: 0.6 - 10 ug nuclear extract/well.
Other Notes
Nrf2 (NF-E2 related factor, NFE2L2, from nuclear factor erythroid-derived 2-like 2) is a basic leucine zipper (bZIP) transcription factor. Nrf2 binds to the antioxidant response element (ARE) and positively regulates the expression of detoxifying enzyme genes (such as NAD(P)H:quinone oxidoreductase1, NQO1) in response to antioxidants and xenobiotics. Higher levels of NQO1 gene expression has been shown in liver, lung, colon, and breast tumors.
A cytosolic inhibitor of Nrf2, Keap1/INrf2, retains Nrf2 in the cytoplasm under normal conditions where the interaction of Nrf2 with INrf2 targets Nrf2 for ubiquitination and proteasomal degradation. However, after oxidative stress, Nrf2 is released from INrf2, translocates to the nucleus, and results in the activation of ARE-mediated gene expression. Nrf2 is also synthesized
de novo
after exposure to stress. In addition, Nrf2 controls its own degradation by regulating expression and induction of INrf2.
It has been shown that nuclear export and degradation pathways are activated by around two hours after treatment with tert-butylhydroquinone (t-BHQ).
Nrf2 activation and degradation are important sensing mechanisms in the cellular response for oxidative and electrophilic stressors.
Properties and Storage Information:
Shipped at conditions-Dry Ice, Appropriate short-term storage conditions-Multi, Appropriate long-term storage conditions-Multi, Storage information-Please refer to protocols
Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
Nrf2 also known as nuclear factor erythroid 2-related factor 2 is a transcription factor with a molecular weight of approximately 66 kDa. It plays a mechanical role in regulating the expression of antioxidant proteins that protect against oxidative damage triggered by injury and inflammation. Nrf2 is ubiquitously expressed in various tissues including the liver lungs and skin. Nrf2 activation occurs when it translocates from the cytoplasm to the nucleus to bind the antioxidant response element (ARE) in the DNA starting transcription of target genes.
Biological function summary
Nrf2 acts as an important regulator of the cellular antioxidant response. It works in conjunction with its partner protein Keap1 forming a complex that controls its stability and degradation. Under normal conditions Keap1 keeps Nrf2 in the cytoplasm where it is targeted for ubiquitination and degradation. Once activated by oxidative stress or electrophiles Nrf2 dissociates from Keap1 thereby avoiding degradation and relocates to the nucleus to activate the transcription of ARE-dependent genes. This activity boosts the cellular response to oxidative stress by inducing genes involved in detoxification and cellular defense.
Pathways
Nrf2 plays a significant role in the oxidative stress response and detoxification pathways. Nrf2 activation is linked closely to the PI3K/Akt signaling pathway which influences cell survival growth and metabolism. This pathway also interacts with other important proteins like GSK-3β which can modulate Nrf2 activity and stability. Through these pathways Nrf2 orchestrates a defense mechanism against reactive oxygen species (ROS) by boosting the expression of antioxidant enzymes and detoxifying proteins.
Nrf2 has been associated with conditions like cancer and neurodegenerative diseases. In cancer aberrant Nrf2 activation may lead to enhanced tumor survival by increasing expression of cytoprotective genes which makes cancer cells resistant to chemotherapy. Nrf2 also interacts with proteins such as p53 which play roles in tumor suppression and cellular stress responses. In neurodegenerative disorders reduced Nrf2 activity can contribute to oxidative stress leading to neuron damage and disease progression with proteins like amyloid-beta also being linked with oxidative processes affected by Nrf2 functionality.
