For the best experience on the Abcam website please upgrade to a modern browser such as Google Chrome

We use cookies to make our site as useful as possible.

Our Cookie Policy explains how you can opt-out of the cookies we use.

If you continue without changing your cookie settings, we'll assume you’re happy with this.

Continue Continue

United States
Your country/region is currently set to:

If incorrect, please enter your country/region into the box below, to view site information related to your country/region.

Call (888) 77-ABCAM (22226) or contact us
Need help? Contact us

  • My account
  • Sign out
Sign in or Register with us

Welcome

Sign in or

Don't have an account?

Register with us
My basket
Quick order
Abcam homepage

  • Research Products
    By product type
    Primary antibodies
    Secondary antibodies
    ELISA and Matched Antibody Pair Kits
    Cell and tissue imaging tools
    Cellular and biochemical assays
    Proteins and Peptides
    By product type
    Proteomics tools
    Agonists, activators, antagonists and inhibitors
    Cell lines and Lysates
    Multiplex miRNA assays
    Multiplex Assays
    By research area
    Cancer
    Cardiovascular
    Cell Biology
    Epigenetics
    Metabolism
    Developmental Biology
    By research area
    Immunology
    Microbiology
    Neuroscience
    Signal Transduction
    Stem Cells
  • Diagnostic & Therapeutic Solutions
    Custom solutions & partnerships

    Custom antibody development and commercial partnerships to advance your diagnostic and therapeutic discovery.

    Create custom solutions with us

    Partner with us

  • Support
    Support hub

    Access advice and support for any research roadblock

    View support hub

    Protocols

    Your experiments laid out step by step

    View protocols

  • Events
    • Conference calendar
    • Cancer
    • Cardiovascular
    • Epigenetics & Nuclear signaling
    • Immunology
    • Neuroscience
    • Stem cells
    • Tradeshows
    • Scientific webinars
    Keep up to date with the latest events

    Full event breakdown with abstracts, speakers, registration and more

    View global event calendar

  • Pathways
    Cell signalling pathways

    View all pathways

    View all interactive pathways

Supporting our customers and employees during the COVID-19 pandemic. Read more

  1. Link

    nf-kb-p65-antibody-e379-ab32536.pdf

  1. Send me a copy of this email
    I agree to the terms and conditions.
Cell Biology Apoptosis Intracellular NFkB RelA (p65)
Share by email
Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-NF-kB p65 antibody [E379] (ab32536)

  • Datasheet
  • SDS
Reviews (14)Q&A (2)References (146)

Product price, shipping and contact information

Currently unavailable

Sorry, we can't display this right now.

Please contact us to place your order, or try again later.

 

Loading size & price…

 

Shipping and order information

Abpromise

Guaranteed product quality, expert customer support.

Find out more.

Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunoprecipitation - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
  • Anti-NF-kB p65 antibody [E379] (ab32536)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [E379] to NF-kB p65
  • Suitable for: WB, IHC-P, ICC/IF, IP
  • Knockout validated
  • Reacts with: Mouse, Human

You may also be interested in

Protein
Product image
Native Clostridium difficile Toxin B protein (ab124001)
Assay
Product image
NFkB p65 Transcription Factor Assay Kit (ab133112)
Primary
Product image
Alexa Fluor® 488 Anti-NF-kB p65 antibody [E379] (ab190205)

View more associated products

Overview

  • Product name

    Anti-NF-kB p65 antibody [E379]
    See all NF-kB p65 primary antibodies
  • Description

    Rabbit monoclonal [E379] to NF-kB p65
  • Host species

    Rabbit
  • Specificity

    ab32536 recognises NF-kB p65. The mouse recommendation is based on the WB results. We do not guarantee IHC-P for mouse. This antibody is unsuitable for detecting NF-KB p65 in mouse tissue lysates. The expression of NF-KB p65 is increased by lipopolysaccharides treatment reported by PMID: 18036230. Mouse brain and spleen are positive reported by PMID: 21479220 and 20008488
  • Tested Applications & Species

    Application Species
    ICC/IF
    Human
    IHC-P
    Human
    IP
    Human
    WB
    Mouse
    Human
    See all applications and species data
  • Immunogen

    Synthetic peptide within Human NF-kB p65 aa 500 to the C-terminus (C terminal). The exact sequence is proprietary.
    Database link: Q04206

  • Positive control

    • WB: Wild-type HAP1 cell lysate. HeLa, MEF, RAW 264.7 and A431 cell lysate; Human fetal brain, kidney and lung tissue lysates. IHC-P: Human breast carcinoma and colon carcinoma tissue. Mouse colon tissue. ICC/IF: HeLa cells. IP: NF-kB p65 IP in HeLa whole cell lysate (ab150035).
  • General notes

    Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

    Abcam recommended secondaries - Goat Anti-Rabbit HRP (ab205718) and Goat Anti-Rabbit Alexa Fluor® 488 (ab150077).

    See other anti-rabbit secondary antibodies that can be used with this antibody.

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing the problem with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation.

    One factor contributing to the crisis is the use of antibodies that are not suitable. This can lead to misleading results and the use of incorrect data informing project assumptions and direction. To help address this challenge, we have introduced an application and species grid on our primary antibody datasheets to make it easy to simplify identification of the right antibody for your needs.

    Learn more here.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.21% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    E379
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Apoptosis
    • Intracellular
    • NFkB
    • RelA (p65)
    • Neuroscience
    • Neurology process
    • Neurodegenerative disease
    • Alzheimer's disease
    • Other
    • Microbiology
    • Interspecies Interaction
    • Host Virus Interaction
    • Signal Transduction
    • Signaling Pathway
    • Nuclear Signaling
    • NFkB Pathway
    • Epigenetics and Nuclear Signaling
    • Nuclear Signaling Pathways
    • NFkB pathway
    • Cancer
    • Signal transduction
    • Nuclear signaling
    • NFkB pathway
    • Cardiovascular
    • Atherosclerosis
    • Vascular Inflammation
    • Inflammatory mediators
    • Microbiology
    • Interspecies Interaction
    • Host Immune Response
    • Metabolism
    • Types of disease
    • Obesity
    • Cancer
    • Cell Death
    • Apoptosis
    • Apoptosis Markers
    • NFkB
    • RelA (p65)

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-NF-kB p65 antibody [E379] (ab190205)
    • Alexa Fluor® 647 Anti-NF-kB p65 antibody [E379] (ab190589)
    • Biotin Anti-NF-kB p65 antibody [E379] (ab205823)
    • Anti-NF-kB p65 antibody [E379] - BSA and Azide free (ab207297)
    • PE Anti-NF-kB p65 antibody [E379] (ab208750)
    • Alexa Fluor® 555 Anti-NF-kB p65 antibody [E379] (ab214634)
    • Alexa Fluor® 594 Anti-NF-kB p65 antibody [E379] (ab214846)
  • Assay kits

    • NFkB p65 Transcription Factor Assay Kit (ab133112)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773)
    • Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed (ab96899)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • HeLa whole cell lysate (ab29545)
  • Recombinant Protein

    • Recombinant Human NF-kB p65 protein (Tagged) (ab114150)
  • Related Products

    • Prestained Protein Ladder - Broad molecular weight (10-245 kDa) (ab116028)
    • Human NF-kB p65 peptide (ab16636)
    • NF kappaB p65 ELISA Kit (ab176648)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab32536 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Guaranteed

Tested applications are guaranteed to work and covered by our Abpromise guarantee.

Predicted

Predicted to work for this combination of applications and species but not guaranteed.

Incompatible

Does not work for this combination of applications and species.

Application Species
ICC/IF
Human
IHC-P
Human
IP
Human
WB
Mouse
Human
Application Abreviews Notes
WB (11)
1/1000 - 1/100000. Detects a band of approximately 65 kDa (predicted molecular weight: 65 kDa).

This antibody might not detect target band in mouse tissues.

IHC-P
1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

The mouse recommendation is based on the WB results. We do not guarantee IHC-P for mouse.

ICC/IF (2)
1/100.

For unpurified use at 1/250 -1/500.

IP
1/30.

For unpurified use at 1/200. 

Notes
WB
1/1000 - 1/100000. Detects a band of approximately 65 kDa (predicted molecular weight: 65 kDa).

This antibody might not detect target band in mouse tissues.

IHC-P
1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

The mouse recommendation is based on the WB results. We do not guarantee IHC-P for mouse.

ICC/IF
1/100.

For unpurified use at 1/250 -1/500.

IP
1/30.

For unpurified use at 1/200. 

  • Application notes
    Is unsuitable for Flow Cyt.
  • Target

    • Function

      NF-kappa-B is a pleiotropic transcription factor which is present in almost all cell types and is involved in many biological processed 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 and the heterodimeric p65-p50 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. 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. NF-kappa-B heterodimeric p65-p50 and p65-c-Rel complexes are transcriptional activators. The NF-kappa-B p65-p65 complex appears to be involved in invasin-mediated activation of IL-8 expression. The inhibitory effect of I-kappa-B upon NF-kappa-B the cytoplasm is exerted primarily through the interaction with p65. p65 shows a weak DNA-binding site which could contribute directly to DNA binding in the NF-kappa-B complex. Associates with chromatin at the NF-kappa-B promoter region via association with DDX1.
    • Sequence similarities

      Contains 1 RHD (Rel-like) domain.
    • Domain

      the 9aaTAD motif is a transactivation domain present in a large number of yeast and animal transcription factors.
    • Post-translational
      modifications

      Ubiquitinated, leading to its proteasomal degradation. Degradation is required for termination of NF-kappa-B response.
      Monomethylated at Lys-310 by SETD6. Monomethylation at Lys-310 is recognized by the ANK repeats of EHMT1 and promotes the formation of repressed chromatin at target genes, leading to down-regulation of NF-kappa-B transcription factor activity. Phosphorylation at Ser-311 disrupts the interaction with EHMT1 without preventing monomethylation at Lys-310 and relieves the repression of target genes.
      Phosphorylation at Ser-311 disrupts the interaction with EHMT1 and promotes transcription factor activity (By similarity). Phosphorylation on Ser-536 stimulates acetylation on Lys-310 and interaction with CBP; the phosphorylated and acetylated forms show enhanced transcriptional activity.
      Reversibly acetylated; the acetylation seems to be mediated by CBP, the deacetylation by HDAC3. Acetylation at Lys-122 enhances DNA binding and impairs association with NFKBIA. Acetylation at Lys-310 is required for full transcriptional activity in the absence of effects on DNA binding and NFKBIA association. Acetylation can also lower DNA-binding and results in nuclear export. Interaction with BRMS1 promotes deacetylation of 'Lys-310'.
    • Cellular localization

      Nucleus. Cytoplasm. Nuclear, but also found in the cytoplasm in an inactive form complexed to an inhibitor (I-kappa-B). Colocalized with RELA in the nucleus upon TNF-alpha induction.
    • Target information above from: UniProt accession Q04206 The UniProt Consortium
      The Universal Protein Resource (UniProt) in 2010
      Nucleic Acids Res. 38:D142-D148 (2010) .

      Information by UniProt
    • Database links

      • Entrez Gene: 5970 Human
      • Entrez Gene: 19697 Mouse
      • Omim: 164014 Human
      • SwissProt: Q04206 Human
      • SwissProt: Q04207 Mouse
      • Unigene: 502875 Human
      • Unigene: 249966 Mouse
      • Alternative names

        • Avian reticuloendotheliosis viral (v rel) oncogene homolog A antibody
        • MGC131774 antibody
        • NF kappa B p65delta3 antibody
        • nfkappabp65 antibody
        • NFkB p65 antibody
        • NFKB3 antibody
        • Nuclear factor kappaB antibody
        • Nuclear Factor NF Kappa B p65 Subunit antibody
        • Nuclear factor NF-kappa-B p65 subunit antibody
        • Nuclear factor of kappa light polypeptide gene enhancer in B cells 3 antibody
        • Nuclear factor of kappa light polypeptide gene enhancer in B-cells 3 antibody
        • OTTHUMP00000233473 antibody
        • OTTHUMP00000233474 antibody
        • OTTHUMP00000233475 antibody
        • OTTHUMP00000233476 antibody
        • OTTHUMP00000233900 antibody
        • p65 antibody
        • p65 NF kappaB antibody
        • p65 NFkB antibody
        • relA antibody
        • TF65_HUMAN antibody
        • Transcription factor NFKB3 antibody
        • Transcription factor p65 antibody
        • v rel avian reticuloendotheliosis viral oncogene homolog A (nuclear factor of kappa light polypeptide gene enhancer in B cells 3 (p65)) antibody
        • V rel avian reticuloendotheliosis viral oncogene homolog A antibody
        • v rel reticuloendotheliosis viral oncogene homolog A (avian) antibody
        • V rel reticuloendotheliosis viral oncogene homolog A, nuclear factor of kappa light polypeptide gene enhancer in B cells 3, p65 antibody
        see all

      Images

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)

        Lane 1: Wild-type HAP1 cell lysate (20 µg)
        Lane 2: NF-kB p65 knockout HAP1 cell lysate (20 µg)
        Lane 3: HeLa cell lysate (20 µg)
        Lane 4: A431 cell lysate (20 µg)

        Lanes 1 - 4: Merged signal (red and green). Green - ab32536 observed at 70 kDa. Red - ab8245 loading control, observed at 37 kDa.


        Unpurified ab32536 was shown to specifically react with NF-kB p65 in wild-type HAP1 cells. No band was observed when NF-kB p65 knockout samples were used. Wild-type and NF-kB p65 knockout samples were subjected to SDS-PAGE. ab32536 (NF-kB p65) and ab8245 (loading control to GAPDH) were diluted to 1/50 000 and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

      • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)

        Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling NF-kB p65 with purified ab32536 at 1:100 dilution.

        Cells were fixed in 100% methanol. ab150077 Goat anti rabbit IgG(Alexa Fluor® 488) was used as the secondary antibody at 1:1000 dilution. DAPI nuclear counterstain.

        PBS instead of the primary antibody was used as the secondary antibody only control.

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        All lanes : Anti-NF-kB p65 antibody [E379] (ab32536) at 1/1000 dilution

        Lane 1 : Human fetal brain lysates
        Lane 2 : Human fetal kidney lysates
        Lane 3 : Human fetal lung lysates

        Lysates/proteins at 20 µg per lane.

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

        Predicted band size: 65 kDa
        Observed band size: 65 kDa



        Blocking/Diluting buffer and concentration: 5% NFDM/TBST

        Exposure time: 37 second for Lane1 and 3 seconds for Lanes 2 and 3

        Normal brain might express low level of p65 (PMID: 21479220)

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)Souslova et al PLoS One. 2010 Mar 25;5(3):e9902. doi: 10.1371/journal.pone.0009902. Fig 2. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

        MMP3 and NFκB are co-localized in human atherosclerotic plaques.

        Sections (4 µm) of paraffin embedded tissue blocks of atherosclerotic plaques were subjected to double immunostaining of MMP3 and p50 (A, B, and C) or MMP3 and p65 (D, E and F, shown), using rabbit polyclonal NFκB p50 and MMP3 antibodies (Abcam) and ab32536, respectively, and goat anti-rabbit secondary antibodies.

        The double immunostaining was visualized with Liquid Permanent Red chromogen and Vector Blue chromogen. Pink colour indicates expression of p50 (A, B and C) or p65 (D, E and F). Blue colour indicates expression of MMP3 (A, B, C, D, E and F.

        Arrow indicates cell co-expressing MMP3 with p50 (B and C) or with p65 (E and F).

        200× magnification in A and D; 400× magnification in B, C, E and F.

      • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)Fazalul Rahiman et al PLoS One. 2016 Apr 7;11(4):e0153005. doi: 10.1371/journal.pone.0153005. eCollection 2016. Fig 1. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

        NF-κB/p65 nuclear localisation in differentiated THP-1 (Human monocytic leukemia cell line) cells.

        Cells were fixed, permeabilized, stained with ab32536 and visualised with Alexa Fluor® 555 goat anti-rabbit IgG (green). The nuclei were counterstained with DAPI (blue). Microscopy images A. Inactive NF-κB/p65 proteins localization in the cytoplasm of the non-stimulated THP-1 cells (top row) and B. Translocated NF-κB/p65 proteins into the nuclei of THP-1 cells following LPS stimulation (bottom row).

        Images were acquired for each fluorescence channel, using suitable TRITC and DAPI filters and a 63× objective.

        Scale bar: 10μm

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)Image from Bel S et al., J Biol Chem. 2012 Jul 20;287(30):25631-9. Epub 2012 May 2. Fig 8.; doi: 10.1074/jbc.M112.364786; July 20, 2012, The Journal of Biological Chemistry, 287, 25631-25639.

        Immunohistochemical analysis of colon sections from mice, staining NF-kB p65 with unpurified ab32536.

        Antigen retrieval was performed by microwave heating in citrate buffer, pH 6. Sections were incubated overnight with primary antibody (1/250) and staining was detected using ab80437 EXPOSE Rabbit specific HRP/DAB detection IHC kit.

      • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)Image from Ali, Ahmed Atef Ahmed et al. PLoS ONE 11.4 (2016): e0154278. Fig #8. doi: 10.1371/journal.pone.0154278. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

        Immunocytochemistry/ Immunofluorescence analysis of human cancer cells labeling NF-kB p65 with unpurified ab32536 (Middle panel).

        Briefly, the tested cells were seeded on coverslips treated with HCl and ethanol, and autoclaved prior to use. Immunostaining of the p65 subunit of NF-κB was done by permeabilizing the cells with Triton X-10, then by treating the cells with anti-NF-κB p65 rabbit monoclonal primary antibody [E379] (ab32536), followed by Alexa Fluor® 488 Donkey anti-rabbit IgG secondary antibody. Nuclei of cells were stained with DAPI (Left panel).

        Merge (Right panel).

        Images were acquired using fluorescence microscope. 

      • Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunocytochemistry/ Immunofluorescence - Anti-NF-kB p65 antibody [E379] (ab32536)

        Immunofluorescent staining of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells using unpurified ab32536.

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        All lanes : Anti-NF-kB p65 antibody [E379] (ab32536) at 1000 µg

        Lane 1 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
        Lane 2 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) treated with 1ug/ml lipopolysaccharides for 6 hours whole cell lysates
        Lane 3 : Mouse brain lysates
        Lane 4 : Mouse spleen lysates

        Lysates/proteins at 15 µg per lane.

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

        Predicted band size: 65 kDa


        Exposure time: 3 seconds


        This antibody is unsuitable for detecting mouse tissue lysates.

        The expression of NF-KB p65 is increased by lipopolysaccharides treatment reported by PMID: 18036230.

        Mouse brain and spleen are positive reported by PMID: 21479220 and 20008488

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)Image courtesy of an anonymous Abreview

        Anti-NF-kB p65 [E379] antibody (unpurified ab32536) reactivity with reduced wild type (WT) and p65 knockout (KO) mouse embryonic fibroblast (MEF) lysate.

        After SDS-PAGE, membranes were blocked in 5% milk in TBS + 0.1% Tween for 1 hour at 25°C before incubation with unpurified ab32536 (1:1,000 dilution in 5% milk TBS + 0.1% Tween) for 16 hours at 4ºC. Blots was then incubated with an anti-Rabbit HRP-conjugated secondary antibody before developing with ECL.

        See Abreview

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)

        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon carcinoma tissue sections labeling NF-kB p65 with Purified ab32536 at 1:2000 dilution (0.2 μg/ml).

        Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used.

        PBS instead of the primary antibody was used as the negative control.

      • Immunoprecipitation - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunoprecipitation - Anti-NF-kB p65 antibody [E379] (ab32536)

        ab32536 (purified) at 1:30 dilution (2 μg) immunoprecipitating NF-kB p65 in HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate.

        Lane 1 (input): HeLa whole cell lysate 10μg
        Lane 2 (+): ab32536 & HeLa whole cell lysate
        Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32536 in HeLa whole cell lysate

        For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
        Blocking and diluting buffer: 5% NFDM/TBST.

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Anti-NF-kB p65 antibody [E379] (ab32536) at 1/5000 dilution (purified) + HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 15 µg

        Secondary
        Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

        Predicted band size: 65 kDa



        Blocking and diluting buffer: 5% NFDM/TBST.

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Anti-NF-kB p65 antibody [E379] (ab32536) at 1/5000 dilution (purified) + MEF (Mouse embryonic fibroblast (immortalized)) whole cell lysates at 15 µg

        Secondary
        Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

        Predicted band size: 65 kDa



        Blocking and diluting buffer: 5% NFDM/TBST.

      • Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Western blot - Anti-NF-kB p65 antibody [E379] (ab32536)
        Anti-NF-kB p65 antibody [E379] (ab32536) at 1/100000 dilution (unpurified) + HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysate

        Predicted band size: 65 kDa
        Observed band size: 65 kDa

      • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)
        Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NF-kB p65 antibody [E379] (ab32536)

        Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using unpurified ab32536.

      • Anti-NF-kB p65 antibody [E379] (ab32536)
        Anti-NF-kB p65 antibody [E379] (ab32536)

      Protocols

      To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

      Click here to view the general protocols

      Datasheets and documents

      • SDS download

      • Datasheet download

        Download

      References (146)

      Publishing research using ab32536? Please let us know so that we can cite the reference in this datasheet.

      ab32536 has been referenced in 146 publications.

      • Zhu H  et al. CircGCN1L1 promotes synoviocyte proliferation and chondrocyte apoptosis by targeting miR-330-3p and TNF-a in TMJ osteoarthritis. Cell Death Dis 11:284 (2020). PubMed: 32332704
      • Li Q  et al. Epigenetic modifier trichostatin A enhanced osteogenic differentiation of mesenchymal stem cells by inhibiting NF-?B (p65) DNA binding and promoted periodontal repair in rats. J Cell Physiol N/A:N/A (2020). PubMed: 32399963
      • Li L  et al. MicroRNA-584 Impairs Cellular Proliferation and Sensitizes Osteosarcoma Cells to Cisplatin and Taxanes by Targeting CCN2. Cancer Manag Res 12:2577-2587 (2020). PubMed: 32346311
      • Kong Q  et al. Autophagy inhibits TLR4-mediated invasiveness of oral cancer cells via the NF-?B pathway. Oral Dis N/A:N/A (2020). PubMed: 32291890
      • Maisto R  et al. Resolvin D1 Modulates the Intracellular VEGF-Related miRNAs of Retinal Photoreceptors Challenged With High Glucose. Front Pharmacol 11:235 (2020). PubMed: 32210819
      View all Publications for this product

      Customer reviews and Q&As

      Show All Reviews Q&A
      Submit a review

      Filter by Application

      Filter by Species

      Filter by Ratings

      1-10 of 14 Abreviews

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Human Cell lysate - whole cell (HMC3 cells)
      Gel Running Conditions
      Reduced Denaturing (4-20% tris-glycin gel)
      Loading amount
      40 µg
      Specification
      HMC3 cells
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%
      Read More

      Abcam user community

      Verified customer

      Submitted Nov 03 2020

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Good
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Deer mouse Tissue lysate - whole (Liver)
      Gel Running Conditions
      Non-reduced Denaturing
      Loading amount
      20 µg
      Specification
      Liver
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
      Read More

      Abcam user community

      Verified customer

      Submitted Oct 28 2019

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Good
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Mouse Cell lysate - whole cell (mouse neural stem cell)
      Gel Running Conditions
      Reduced Denaturing (4-12%)
      Loading amount
      40 µg
      Specification
      mouse neural stem cell
      Blocking step
      BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
      Read More

      Abcam user community

      Verified customer

      Submitted Jun 12 2018

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Human Cell lysate - whole cell (mesenchymal stem cells)
      Gel Running Conditions
      Reduced Denaturing
      Loading amount
      40 µg
      Specification
      mesenchymal stem cells
      Blocking step
      Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 22°C
      Read More

      Dr. Kate Hawkins

      Verified customer

      Submitted Feb 22 2017

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Mouse Cell lysate - whole cell (Colon)
      Gel Running Conditions
      Reduced Denaturing (12.5%)
      Loading amount
      1e+006 cells
      Specification
      Colon
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C
      Read More

      Abcam user community

      Verified customer

      Submitted Dec 23 2015

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Human Cell lysate - whole cell (HeLa)
      Gel Running Conditions
      Reduced Denaturing (12.5%)
      Loading amount
      1e+006 cells
      Specification
      HeLa
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C
      Read More

      Abcam user community

      Verified customer

      Submitted Dec 23 2015

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Average
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Human Cell lysate - whole cell (human head and neck cells)
      Gel Running Conditions
      Reduced Denaturing
      Loading amount
      20 µg
      Treatment
      10 nM PMA
      Specification
      human head and neck cells
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
      Read More

      Abcam user community

      Verified customer

      Submitted Aug 31 2015

      Western blot abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Human Cell lysate - whole cell (Human THP-1 macrophages)
      Gel Running Conditions
      Reduced Denaturing (10% SDS-PAGE)
      Loading amount
      70 µg
      Treatment
      100 µg/ml oxidized LDL (oxLDL) +/- 10 µg/ml apoA-I, 24h
      Specification
      Human THP-1 macrophages
      Blocking step
      Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
      Read More

      Dr. Anca Sima

      Verified customer

      Submitted Jun 04 2015

      Immunocytochemistry/ Immunofluorescence abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      abreview image
      Application
      Immunocytochemistry/ Immunofluorescence
      Blocking step
      Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 22°C
      Sample
      Human Cell (MCF-7)
      Specification
      MCF-7
      Permeabilization
      Yes - Triton X-100
      Fixative
      Paraformaldehyde
      Read More

      Abcam user community

      Verified customer

      Submitted Sep 23 2014

      Flow Cytometry abreview for Anti-NF-kB p65 antibody [E379]

      Excellent
      Abreviews
      Abreviews
      This product is known to not work in this application or species.
      abreview image
      Application
      Flow Cytometry
      Fixation
      Paraformaldehyde
      Permeabilization
      Yes - 70% methanol
      Sample
      Human Cell (MCF-7)
      Specification
      MCF-7
      Gating Strategy
      Live cells gated
      Preparation
      Cell harvesting/tissue preparation method: Cell dissociation buffer
      Sample buffer: Enzyme free
      Read More

      Abcam user community

      Verified customer

      Submitted Aug 15 2014

      1-10 of 14 Abreviews

      •  Previous
      • 1
      • 2
      • Next 

      Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
      For licensing inquiries, please contact partnerships@abcam.com

      Get resources and offers direct to your inbox Sign up
      A-Z by research area
      • Cancer
      • Cardiovascular
      • Cell biology
      • Developmental biology
      • Epigenetics & Nuclear signaling
      • Immunology
      • Metabolism
      • Microbiology
      • Neuroscience
      • Signal transduction
      • Stem cells
      A-Z by product type
      • Primary antibodies
      • Secondary antibodies
      • Biochemicals
      • Isotype controls
      • Flow cytometry multi-color selector
      • Kits
      • Loading controls
      • Lysates
      • Peptides
      • Proteins
      • Slides
      • Tags and cell markers
      • Tools & Reagents
      Help & support
      • Support
      • Make an Inquiry
      • Protocols & troubleshooting
      • Placing an order
      • RabMAb products
      • Biochemical product FAQs
      • Training
      • Browse by Target
      Company
      • Corporate site
      • Investor relations
      • Company news
      • Careers
      • About us
      • Blog
      Events
      • Tradeshows
      • Conferences
      International websites
      • abcam.cn
      • abcam.co.jp

      Join with us

      • LinkedIn
      • facebook
      • Twitter
      • YouTube
      • Terms of sale
      • Website terms of use
      • Cookie policy
      • Privacy policy
      • Legal
      • Modern slavery statement
      © 1998-2021 Abcam plc. All rights reserved.