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AB133112

NFkB p65 Transcription Factor Assay试剂盒

NFkB p65 Transcription Factor Assay Kit

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(1 Review)

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(113 Publications)

NFkB p65 Transcription Factor Assay Kit ab133112 is a non-radioactive, sensitive ELISA-based method for detecting specific transcription factor NF-kB (p65) DNA binding activity in nuclear extracts.

- No cross-reactivity with NF-kB (p50)
- Colorimetric assay - 450nm readout; works on any plate reader
- Cited in over 95 publications

查看别名

NFKB3, RELA, Transcription factor p65, Nuclear factor NF-kappa-B p65 subunit, Nuclear factor of kappa light polypeptide gene enhancer in B-cells 3

4 Images
Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)
  • FuncS

PubMed

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)

Determining p65-NF-κB by ELISA using ab133112.

After the treatment with LPS (10 μg/ml for 6 hrs), cells were lysed with hypotonic HEPES lysis buffer (pH 7.4) and centrifuged at 1000 g for 10 min at 4°C, supernatants were collected and used for the determination of intracellular p65- NF-κB by ELISA. The absorbance was read at 450 nm using spectrophotometer.

Saha D et al., PLoS One, 12(2). Fig 9a. doi: 10.1371/journal.pone.0171084 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)
  • FuncS

Lab

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)

Titration of positive control with or without inhibitor, background signal subtracted (duplicates; +/- SD).

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)
  • FuncS

Supplier Data

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)

Assay of cell lysates isolated from stimulated (20 ng/ml TNF alpha for 30 minutes) and nonstimulated HeLa cells demonstrating NFkB (p65) activity.

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)
  • FuncS

Lab

Functional Studies - NFkB p65 Transcription Factor Assay Kit (AB133112)

Jurkat cells were treated with PMA and ionomycin (+). Nuclear lysates were extracted (ab113474) and 40 uL, corresponding to 4e6 cells, were tested in duplicates (+/- SD).

关键信息

检测方法

Colorimetric

样品类型

Nuclear Extracts

反应种属

Mouse, Rat, Human

检测类型

Semi-quantitative

检测时间

3h 30m

检测平台

Microplate reader

产品详情

NFkB p65 Transcription Factor Assay Kit ab133112 is a non-radioactive, sensitive ELISA-based method for detecting specific transcription factor DNA binding activity in nuclear extracts.

In the NFkB p65 assay, a double stranded DNA sequence containing the NFkB response element is immobilized onto the bottom of the wells of a 96-well plate. NFkB contained in a nuclear extract, binds to the NFkB response element, and is detected using an anti NFkB p65 antibody. A secondary antibody conjugated to HRP is added to provide a colorimetric readout at 450 nm.

NFkB p65 transcription factor assay protocol summary:
- prepare nuclear extracts from cells
- add samples to wells
- incubate for 1 hr or o/n
- wash with wash buffer
- add NFkB antibody and incubate for 1 hr , then wash
- add HRP-conjugated secondary antibody and incubate for 1 hr, then wash
- add developing solution and incubate for 15-45 min
- add stop solution
- analyze with microplate reader

REACH authorisation
Abcam has not and does not intend to apply for the REACH Authorisation of customers' uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.

规格

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性能和储存信息

运输条件
Dry Ice
推荐的短期储存条件
Multi
推荐的长期储存条件
Multi
储存信息
Please refer to protocols

补充信息

This supplementary information is collated from multiple sources and compiled automatically.

NF-kB p65 also known as RelA is a significant component of the NF-kB protein complex. This complex usually involves a molecular weight for p65 of approximately 65 kDa. NF-kB p65 is mechanistically a transcription factor that regulates genes involved in inflammation cell survival and immune response. Expression of p65 is widely seen in various cell types including immune cells and epithelial cells suggesting its role in numerous physiological processes.
Biological function summary

NF-kB p65 acts as part of the larger NF-kB complex usually forming a heterodimer with other family members like p50. This complex translocates to the nucleus upon activation where it binds specific DNA sequences to regulate gene expression. The p65 subunit is essential for transactivating target genes involved in immune and inflammatory responses. Its regulation is important for proper cellular functioning especially in the maintenance of immune homeostasis and inflammatory response.

Pathways

NF-kB p65 participates in critical pathways like the NF-kB signaling and Toll-like receptor pathways. These pathways are fundamental for initiating immune responses and contribute to the regulation of apoptosis and cellular stress responses. In the context of these pathways the IKK complex plays a pivotal role in NF-kB activation leading to the phosphorylation and subsequent degradation of inhibitors that retain NF-kB p65 in the cytoplasm therefore allowing its movement to the nucleus.

NF-kB p65 is implicated in conditions such as cancer and autoimmune diseases. Its dysregulation can lead to persistent activation of inflammatory pathways contributing to tumorigenesis and chronic inflammation. In the context of these diseases proteins such as the p50 subunit and the IKK complex are often involved reflecting the importance of tight regulation of NF-kB signaling in preventing pathological conditions.

产品实验方案

靶点信息

NF-kappa-B is a pleiotropic transcription factor present in almost all cell types and is the endpoint of a series of signal transduction events that are initiated by a vast array of stimuli related to many biological processes such as inflammation, immunity, differentiation, cell growth, tumorigenesis and apoptosis. NF-kappa-B is a homo- or heterodimeric complex formed by the Rel-like domain-containing proteins RELA/p65, RELB, NFKB1/p105, NFKB1/p50, REL and NFKB2/p52. The heterodimeric RELA-NFKB1 complex appears to be most abundant one. The dimers bind at kappa-B sites in the DNA of their target genes and the individual dimers have distinct preferences for different kappa-B sites that they can bind with distinguishable affinity and specificity. Different dimer combinations act as transcriptional activators or repressors, respectively. The NF-kappa-B heterodimeric RELA-NFKB1 and RELA-REL complexes, for instance, function as transcriptional activators. NF-kappa-B is controlled by various mechanisms of post-translational modification and subcellular compartmentalization as well as by interactions with other cofactors or corepressors. NF-kappa-B complexes are held in the cytoplasm in an inactive state complexed with members of the NF-kappa-B inhibitor (I-kappa-B) family. In a conventional activation pathway, I-kappa-B is phosphorylated by I-kappa-B kinases (IKKs) in response to different activators, subsequently degraded thus liberating the active NF-kappa-B complex which translocates to the nucleus. The inhibitory effect of I-kappa-B on NF-kappa-B through retention in the cytoplasm is exerted primarily through the interaction with RELA. RELA shows a weak DNA-binding site which could contribute directly to DNA binding in the NF-kappa-B complex. Beside its activity as a direct transcriptional activator, it is also able to modulate promoters accessibility to transcription factors and thereby indirectly regulate gene expression. Associates with chromatin at the NF-kappa-B promoter region via association with DDX1. Essential for cytokine gene expression in T-cells (PubMed : 15790681). The NF-kappa-B homodimeric RELA-RELA complex appears to be involved in invasin-mediated activation of IL-8 expression. Key transcription factor regulating the IFN response during SARS-CoV-2 infection (PubMed : 33440148).
See full target information RELA

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