Key Product Details

Validated by

Knockout/Knockdown, Biological Validation

Species Reactivity

Validated:

Human, Mouse, Rat

Cited:

Human, Mouse

Applications

Validated:

Knockout Validated, Immunohistochemistry, Western Blot, Immunocytochemistry, Simple Western, Chromatin Immunoprecipitation (ChIP)

Cited:

Western Blot, Immunoprecipitation, Chromatin Immunoprecipitation (ChIP)

Label

Unconjugated

Antibody Source

Polyclonal Sheep IgG
View all RelA/NFkB p65 Primary Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human RelA/NF kappa B p65 isoform 1
Asn456-Ser551
Accession # Q04206

Specificity

Detects human, mouse, and rat RelA/NF kappa B p65 in Western blots.

Clonality

Polyclonal

Host

Sheep

Isotype

IgG

Scientific Data Images for Human/Mouse/Rat RelA/NF kappa B p65 Antibody

Detection of Human RelA/NF?B p65 antibody by Western Blot.

Detection of Human RelA/NF kappa B p65 by Western Blot.

Western blot shows lysates of K562 human chronic myelogenous leukemia cell line, Daudi human Burkitt's lymphoma cell line, and LNCaP human prostate cancer cell line. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for RelA/NF kappa B p65 at approximately 70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of Human and Mouse RelA/NF?B p65 antibody by Western Blot.

Detection of Human and Mouse RelA/NF kappa B p65 by Western Blot.

Western blot shows lysates of K562 human chronic myelogenous leukemia cell line, HeLa human cervical epithelial carcinoma cell line, and Neuro-2A mouse neuroblastoma cell line. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for RelA/NF kappa B p65 at approximately 70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of Mouse and Rat RelA/NF?B p65 antibody by Western Blot.

Detection of Mouse and Rat RelA/NF kappa B p65 by Western Blot.

Western blot shows lysates of NIH-3T3 mouse embryonic fibroblast cell line and C6 rat glioma cell line. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human/Mouse RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for RelA/NF kappa B p65 at approximately 65 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of RelA/NF kappa B p65-regulated Genes by Chromatin Immuno-precipitation.

Detection of RelA/NF kappa B p65-regulated Genes by Chromatin Immuno-precipitation.

Jurkat human acute T cell leukemia cell line treated with 50 ng/mL PMA and 200 ng/mL calcium ionomycin for overnight was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. RelA/NF kappa B p65/DNA complexes were immunoprecipitated using 5 µg Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) or control antibody (Catalog # 5-001-A) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Sheep IgG Secondary Antibody (Catalog # BAF016). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. Thep21promoter was detected by standard PCR.

RelA/NF?B p65 antibody in HeLa Human Cell Line by Immunocytochemistry (ICC).

RelA/NF kappa B p65 in HeLa Human Cell Line.

RelA/NF kappa B p65 was detected in immersion fixed HeLa human cervical epithelial carcinoma cells untreated (upper panel) or treated (lower panel) with 20 ng/mL Recombinant Human TNF-alpha (Catalog # 210-TA) for 10 minutes using Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm in untreated cells and nuclei in treated cells. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

RelA/NF?B p65 antibody in Human Squamous Cell Carcinoma by Immunohistochemistry (IHC-P).

RelA/NF kappa B p65 in Human Squamous Cell Carcinoma.

RelA/NF kappa B p65 was detected in immersion fixed paraffin-embedded sections of human squamous cell carcinoma using Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) at 3 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in cancer cells. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of Human, Mouse, and Rat RelA/NF?B p65 antibody by Simple WesternTM.

Detection of Human, Mouse, and Rat RelA/NF kappa B p65 by Simple WesternTM.

Simple Western lane view shows lysates of HeLa human cervical epithelial carcinoma cell line, Neuro-2A mouse neuroblastoma cell line, and C6 rat glioma cell line, loaded at 0.5 mg/mL. A specific band was detected for RelA/NF kappa B p65 at approximately 65 kDa (as indicated) using 20 µg/mL of Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) followed by 1:50 dilution of HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western Blot Shows Human RelA/NF?B p65 Antibody Specificity by Using Knockout Cell Line.

Western Blot Shows Human RelA/NF kappa B p65 Specificity by Using Knockout Cell Line.

Western blot shows lysates of HeLa human cervical epithelial carcinoma parental cell line and RelA/NF kappa B p65 knockout HeLa cell line (KO). PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human/Mouse/Rat RelA/NF kappa B p65 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5078) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for RelA/NF kappa B p65 at approximately 65 kDa (as indicated) in the parental HeLa cell line, but is not detectable in knockout HeLa cell line. GAPDH (Catalog # AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Applications for Human/Mouse/Rat RelA/NF kappa B p65 Antibody

Application
Recommended Usage

Chromatin Immunoprecipitation (ChIP)

5 µg/5 x 106 cells
Sample: PMA and calcium ionomycin treated Jurkat human acute T cell leukemia cell line chromatin, p21 promoter detected by standard PCR

Immunocytochemistry

5-15 µg/mL
Sample: Immersion fixed HeLa human cervical epithelial carcinoma cells untreated or treated with 20 ng/mL Recombinant Human TNF-alpha (Catalog # 210-TA)

Immunohistochemistry

5-15 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human squamous cell carcinoma

Knockout Validated

RelA/NF kappa B p65 is specifically detected in HeLa human cervical epithelial carcinoma parental cell line but is not detectable in RelA/NF kappa B p65 knockout HeLa cell line.

Simple Western

20 µg/mL
Sample: HeLa human cervical epithelial carcinoma cell line, Neuro‑2A mouse neuroblastoma cell line, and C6 rat glioma cell line

Western Blot

1 µg/mL
Sample: K562 human chronic myelogenous leukemia cell line, Daudi human Burkitt's lymphoma cell line, LNCaP human prostate cancer cell line, HeLa human cervical epithelial carcinoma cell line, Neuro‑2A mouse neuroblastoma cell line, NIH‑3T3 mouse embryonic fibroblast cell line, and C6 rat glioma cell line

Reviewed Applications

Read 2 reviews rated 5 using AF5078 in the following applications:

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: RelA/NFkB p65

RelA belongs to a family of transcription factors (NF kappa B (nuclear factor kappa from B cells) complex) that play a fundamental role in inflammatory and immune responses. The NF kappa B complex is composed of a heterodimer of a Rel family member (RelA, c-Rel, RelB) and either NF kappa B1 or NF kappa B2 subunits. RelA and NF kappa B1 are the most common heterodimeric pair. The NF kappa B complex is sequestered in the cytoplasm by inhibitory I kappa B proteins. Upon cellular activation, the ubiquitin-proteosome pathway degrades the I kappa B proteins allowing the NF kappa B complex to translocate to the nucleus and activate gene transcription.

Long Name

v-rel Reticuloendotheliosis Viral Oncogene Homolog A

Alternate Names

NFkB p65, NFKB3, p65RelA

Entrez Gene IDs

5970 (Human)

Gene Symbol

RELA

UniProt

Q04206

Additional RelA/NFkB p65 Products

Product Documents for Human/Mouse/Rat RelA/NF kappa B p65 Antibody

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Human/Mouse/Rat RelA/NF kappa B p65 Antibody

For research use only

Citations for Human/Mouse/Rat RelA/NF kappa B p65 Antibody

Customer Reviews for Human/Mouse/Rat RelA/NF kappa B p65 Antibody (2)

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  • Human/Mouse/Rat RelA/NF kappa B p65 Antibody
    Name: Anonymous
    Application: ELISA
    Sample Tested: xenograft PD sample
    Species: Human
    Verified Customer | Posted 10/24/2019
    Human/Mouse/Rat RelA/NF kappa B p65 Antibody AF5078
  • Human/Mouse RelA/NF kappa B p65 Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Human fibroblast
    Species: Human
    Verified Customer | Posted 08/23/2018
    Human/Mouse/Rat RelA/NF kappa B p65 Antibody AF5078

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