NFKB p65 Antibody
Alternative names: RELA, NFKB3, p65
Polyclonal antibody raised in rabbit against NFKB p65 (Rel A), using a KLH-conjugated synthetic peptide.
| Lot | 24468 |
|---|---|
| Concentration | not determined |
| Species reactivity | Human, mouse, rat |
| Type | Polyclonal |
| Purity | Whole antiserum |
| Host | Rabbit |
| Precautions | This product is for research use only. Not for use in diagnostic or therapeutic procedures. |
| Applications | Suggested dilution | References |
|---|---|---|
| ChIP/ChIP-seq * | 1 µl/IP | Fig 1, 2 |
| Western Blotting | 1:2,000 - 1:5,000 | Fig 3 |
| ELISA | 1:5,000 | |
| Immunohistochemistry | 1:500 - 1:2,000 | Fig 4 |
| Gel Shift | 1:500 |
* Please note that the optimal antibody amount per IP should be determined by the end-user. We recommend testing 1-5 µg per IP.
Validation Data
Figure 1. ChIP results obtained with the Diagenode antibody directed against NFkB p65.
ChIP assays were performed using human HeLa cells, treated with TNFalpha, the Diagenode antibody against NFkB p65 (Cat. No. C15310256) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (Cat. No. C01010055), using sheared chromatin from 4 million cells. A titration of the antibody consisting of 1, 2.5 and 5 µl per ChIP experiment was analysed. IgG (1 µg/IP) was used as negative IP control. QPCR was performed with primers for the NFKBIA and CCL20 genes, used as positive controls, and for TSH2B, used as negative control. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against NFkB p65
ChIP was performed on sheared chromatin from 4 million HeLa cells using 1 µg of the Diagenode antibody against NFkB p65 (Cat. No. C15310256) as described above. The IP’d DNA was subsequently analysed on an Illumina HiSeq. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 51 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the enrichment along the complete sequence and a 2 Mb region of human chromosome 2 (fig 2A and B), and in a two genomic regions surrounding the NFKBIA and CCL20 positive control genes.
Figure 3. NFKB p65 antibody western blot results
Whole cell extracts from HeLa cells (35 µg) were analysed by Western blot using the Diagenode antibody against NFkB p65 (Cat. No. C15310256) diluted 1:5,000. The position of the protein of interest is indicated on the right (expected size: 65 kDa); the marker (in kDa) is shown on the left.
Figure 4. NFKB p65 antibody Immunohistochemistry results Formalin fixed paraffin embedded lymphocytes and germinal center cells of the tonsil were stained with the Diagenode antibody against NFkB p65 (Cat. No. C15310256) diluted 1:400 followed by a peroxidase labelled goat anti-rabbit secondary antibody. Figure 4 shows moderate positive nuclear or cytoplasmic staining.
Target Description
This antibody recognizes NFKB p65 which is a component of NFKB. NFKB was originally identified as a factor that binds to the immunoglobulin kappa light chain enhancer in B cells. It was subsequently found in non-B cells in an inactive cytoplasmic form consisting of NFkappaB bound to IkappaB. NFkappaB was originally identified as a heterodimeric DNA binding protein complex consisting of p65 (RelA) and p50 (NFKB1) subunits. Other identified subunits include p52 (NFKB2), c-Rel, and RelB. The p65, cRel, and RelB subunits are responsible for transactivation. The p50 and p52 subunits possess DNA binding activity but limited ability to transactivate. p52 has been reported to form transcriptionally active heterodimers with the NFkappaB subunit p65, similar to p50/p65 heterodimers. The heterodimers of p52/p65 and p50/p65 are regulated by physical inactivation in the cytoplasm by IkB-a. IkB-a binds to the p65 subunit, preventing nuclear localization and DNA binding. Low levels of p52 and p50 homodimers can also exist in cells.
Applications
| ELISA Enzyme-linked immunosorbent assay. Read more |
| IHC Immunohistochemistry Read more |
| WB Western blot : The quality of antibodies used in this technique is crucial for correct and specific protein identification. Diagenode offers huge selection of highly sensitive and specific western blot-validated antibodies. Learn more about: Load... Read more |
| ChIP-seq (ab) Read more |
| ChIP-qPCR (ab) Read more |
Documents
| Datasheet NFKBp65 C15310256 DATASHEET Datasheet description | Download |
| Antibodies you can trust POSTER Epigenetic research tools have evolved over time from endpoint PCR to qPCR to the analyses of lar... | Download |
| Epigenetic Antibodies Brochure BROCHURE More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e... | Download |
Safety sheets
Publications
How to properly cite our product/service in your workWe strongly recommend using this: NFKB p65 Antibody (Hologic Diagenode Cat# C15310256 Lot# 24468). Click here to copy to clipboard. Using our products or services in your publication? Let us know! |
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Constitutive expression of the transcriptional co-activator IκBζ promotes melanoma growth and immunotherapy resistance |
Peroxiredoxin 1-Toll-like receptor 4-p65 axis inhibits receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation |
PBRM1-deficient PBAF complexes target aberrant genomic loci to activatethe NF-κB pathway in clear cell renal cell carcinoma. |
Vitamin C enhances NF-κB-driven epigenomic reprogramming andboosts the immunogenic properties of dendritic cells. |
The CDK4/6-EZH2 pathway is a potential therapeutic target for psoriasis. |
AP-1 activity induced by co-stimulation is required for chromatin opening during T cell activation. |
RRAD, IL4I1, CDKN1A, and SERPINE1 genes are potentially co-regulated by NF-κB and p53 transcription factors in cells exposed to high doses of ionizing radiation. |
Pro-inflammatory cytokine and high doses of ionizing radiation have similar effects on the expression of NF-kappaB-dependent genes. |