Recombinant
RabMAb

Anti-NFkB p100 antibody [EPR18756] (ab191594)

Overview

  • Product name
    Anti-NFkB p100 antibody [EPR18756]
    See all NFkB p100 primary antibodies
  • Description
    Rabbit monoclonal [EPR18756] to NFkB p100
  • Tested applications
    Suitable for: Flow Cyt, ICC/IF, IP, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Recombinant fragment within Human NFkB p100 aa 700 to the C-terminus. The exact sequence is proprietary.
    Database link: Q00653

  • Positive control
    • WB: Human fetal brain and fetal kidney lysates; HeLa, Jurkat, MCF7, Daudi, C6 and NIH/3T3 whole cell lysates; Mouse heart and kidney lysates; Rat brain, heart, kidney and spleen lysates. ICC/IF: HeLa and NIH/3T3 cell lines. Flow Cyt: HeLa cell line. IP: HeLa whole cell lysate.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

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

Properties

Applications

Our Abpromise guarantee covers the use of ab191594 in the following tested applications.

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

Application Abreviews Notes
Flow Cyt 1/60.
ICC/IF 1/250.
IP 1/20.
WB 1/1000. Detects a band of approximately 110 kDa (predicted molecular weight: 97 kDa).

Target

  • Relevance
    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 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. In a non-canonical activation pathway, the MAP3K14-activated CHUK/IKKA homodimer phosphorylates NFKB2/p100 associated with RelB, inducing its proteolytic processing to NFKB2/p52 and the formation of NF-kappa-B RelB-p52 complexes. The NF-kappa-B heterodimeric RelB-p52 complex is a transcriptional activator. The NF-kappa-B p52-p52 homodimer is a transcriptional repressor. NFKB2 appears to have dual functions such as cytoplasmic retention of attached NF-kappa-B proteins by p100 and generation of p52 by a cotranslational processing. The proteasome-mediated process ensures the production of both p52 and p100 and preserves their independent function. p52 binds to the kappa-B consensus sequence 5'-GGRNNYYCC-3', located in the enhancer region of genes involved in immune response and acute phase reactions. p52 and p100 are respectively the minor and major form; the processing of p100 being relatively poor. Isoform p49 is a subunit of the NF-kappa-B protein complex, which stimulates the HIV enhancer in synergy with p65. In concert with RELB, regulates the circadian clock by repressing the transcriptional activator activity of the CLOCK-ARNTL/BMAL1 heterodimer.
  • Cellular localization
    Cytoplasmic and Nuclear
  • Database links
  • Alternative names
    • CVID10 antibody
    • DNA binding factor KBF2 antibody
    • H2TF1 antibody
    • Lymphocyte translocation chromosome 10 protein antibody
    • LYT 10 antibody
    • NF kB2 antibody
    • NFKB p52/p100 subunit antibody
    • Nuclear factor Kappa B subunit 2 antibody
    • Nuclear factor of kappa light polypeptide gene enhancer in B cells 2 (p49/p100) antibody
    • Nuclear factor of kappa light polypeptide gene enhancer in B cells 2 antibody
    • Oncogene Lyt 10 antibody
    • p100 antibody
    • Transcription factor NFKB2 antibody
    see all

Images

  • All lanes : Anti-NFkB p100 antibody [EPR18756] (ab191594) at 1/1000 dilution

    Lane 1 : Human fetal brain lysate
    Lane 2 : Human fetal kidney lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

    Predicted band size : 97 kDa
    Observed band size : 110 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    Blocking/Dilution buffer: 5% NFDM/TBST.

    The molecular weight observed is consistent with what have been described in the literature (PMID: 8606850, 17015635).

  • All lanes : Anti-NFkB p100 antibody [EPR18756] (ab191594) at 1/5000 dilution

    Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 2 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate
    Lane 3 : MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
    Lane 4 : Daudi (Human Burkitt's lymphoma cell line) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 97 kDa
    Observed band size : 110 kDa (why is the actual band size different from the predicted?)

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure time:

    Lane 1, 2 & 3: 30 seconds; Lane 4: 5 seconds.

    The molecular weight observed is consistent with what have been described in the literature (PMID: 18606850, 17015635).

  • All lanes : Anti-NFkB p100 antibody [EPR18756] (ab191594) at 1/1000 dilution

    Lane 1 : Mouse heart lysate
    Lane 2 : Mouse kidney lysate
    Lane 3 : Rat brain lysate
    Lane 4 : Rat heart lysate
    Lane 5 : Rat kidney lysate
    Lane 6 : Rat spleen lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 97 kDa
    Observed band size : 110 kDa (why is the actual band size different from the predicted?)

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure time:

    Lane 1 & 2: 3 minutes; Lane 3 & 4: 30 seconds; Lane 5 & 6: 5 seconds.

    The molecular weight observed is consistent with what have been described in the literature (PMID: 18606850, 17015635).

  • All lanes : Anti-NFkB p100 antibody [EPR18756] (ab191594) at 1/1000 dilution

    Lane 1 : C6 (Rat glial tumor cell line) whole cell lysate
    Lane 2 : NIH/3T3 (Mouse embyro fibroblast cell line) whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size : 97 kDa
    Observed band size : 110 kDa (why is the actual band size different from the predicted?)


    Exposure time : 1 second

    Blocking/Dilution buffer: 5% NFDM/TBST.

    The molecular weight observed is consistent with what have been described in the literature (PMID: 18606850, 17015635).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cell line labeling NFkB p100 with ab191594 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing cytoplasmic staining on HeLa cell line.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab191594 at 1/250 dilution followed by ab150120 at 1/1000 dilution.

    -ve control 2: ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embyro fibroblast cell line) cell line labeling NFkB p100 with ab191594 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing cytoplasmic staining on NIH/3T3 cell line.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab191594 at 1/250 dilution followed by ab150120 at 1/1000 dilution.

    -ve control 2: ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cell line labeling NFkB p100 with ab191594 at 1/60 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti Rabbit IgG (Alexa Fluor® 488) at 1/500 dilution was used as the secondary antibody.

  • NFkB p100 was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab191594 at 1/20 dilution.

    Western blot was performed from the immunoprecipitate using ab191594 at 1/1000 dilution.

    VeriBlot for IP secondary antibody (HRP) (ab131366) was used as secondary antibody at 1/10000 dilution.

    Lane 1: HeLa whole cell lysate, 10ug (Input).

    Lane 2: ab191594 IP in HeLa whole cell lysate.

    Lane 3: Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab191594 in HeLa whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 3 seconds.

References

ab191594 has not yet been referenced specifically in any publications.

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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