RabMAb

Anti-IKB alpha抗体[E130] (ab32518)

概述

性能

应用

Our Abpromise guarantee covers the use of ab32518 in the following tested applications.

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

应用 Ab评论 说明
ICC/IF 1/50.
WB 1/1000 - 1/100000. Detects a band of approximately 35 kDa (predicted molecular weight: 36 kDa).
IHC-P 1/100.
IP 1/20.

靶标

  • 功能Inhibits the activity of dimeric NF-kappa-B/REL complexes by trapping REL dimers in the cytoplasm through masking of their nuclear localization signals. On cellular stimulation by immune and proinflammatory responses, becomes phosphorylated promoting ubiquitination and degradation, enabling the dimeric RELA to translocate to the nucleus and activate transcription.
  • 疾病相关Ectodermal dysplasia, anhidrotic, with T-cell immunodeficiency autosomal dominant
  • 序列相似性Belongs to the NF-kappa-B inhibitor family.
    Contains 5 ANK repeats.
  • 翻译后修饰Phosphorylated; disables inhibition of NF-kappa-B DNA-binding activity. Phosphorylation at positions 32 and 36 is prerequisite to recognition by UBE2D3 leading to polyubiquitination and subsequent degradation.
    Sumoylated; sumoylation requires the presence of the nuclear import signal. Sumoylation blocks ubiquitination and proteasome-mediated degradation of the protein thereby increasing the protein stability.
    Monoubiquitinated at Lys-21 and/or Lys-22 by UBE2D3. Ubiquitin chain elongation is then performed by CDC34 in cooperation with the SCF(FBXW11) E3 ligase complex, building ubiquitin chains from the UBE2D3-primed NFKBIA-linked ubiquitin. The resulting polyubiquitination leads to protein degradation. Also ubiquitinated by SCF(BTRC) following stimulus-dependent phosphorylation at Ser-32 and Ser-36.
    Deubiquitinated by porcine reproductive and respiratory syndrome virus Nsp2 protein, which thereby interferes with NFKBIA degradation and impairs subsequent NF-kappa-B activation.
  • 细胞定位Cytoplasm. Nucleus. Shuttles between the nucleus and the cytoplasm by a nuclear localization signal (NLS) and a CRM1-dependent nuclear export.
  • Information by UniProt
  • 数据库链接
  • 别名
    • I kappa B alpha antibody
    • I-kappa-B-alpha antibody
    • IkappaBalpha antibody
    • IkB-alpha antibody
    • IKBA antibody
    • IKBA_HUMAN antibody
    • IKBalpha antibody
    • MAD 3 antibody
    • MAD3 antibody
    • Major histocompatibility complex enhancer-binding protein MAD3 antibody
    • NF kappa B inhibitor alpha antibody
    • NF-kappa-B inhibitor alpha antibody
    • NFKBI antibody
    • NFKBIA antibody
    • Nuclear factor of kappa light chain gene enhancer in B cells antibody
    • Nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor alpha antibody
    see all

Anti-IKB alpha antibody [E130] 图像

  • Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using unpurified ab32518 at 1/50 dilution.

  • ICC/IF image of unpurified ab32518 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32518, 1/1000 dilution) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • All lanes : Anti-IKB alpha antibody [E130] (ab32518) at 1/10000 dilution (purified)

    Lane 1 : PC12 cell lysate
    Lane 2 : NIH3T3 cell lysate
    Lane 3 : RAW264.7 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    HRP goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size : 36 kDa
    Observed band size : 35 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • All lanes : Anti-IKB alpha antibody [E130] (ab32518) at 1/10000 dilution (purified)

    Lane 1 : HeLa cell lysate
    Lane 2 : K562 cell lysate
    Lane 3 : HepG2 cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    HRP goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size : 36 kDa
    Observed band size : 35 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • Anti-IKB alpha antibody [E130] (ab32518) at 1/1000 dilution (purified) + Human fetal liver at 10 µg

    Secondary
    HRP anti-rabbit IgG, specific to the non reduced form of IgG at 1/1000 dilution

    Predicted band size : 36 kDa
    Observed band size : 35 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • Anti-IKB alpha antibody [E130] (ab32518) at 1/1000 dilution (purified) + SH-SY5Y cell lysate at 10 µg

    Secondary
    HRP goat anti-rabbit (H+L) at 1/1000 dilution

    Predicted band size : 36 kDa
    Observed band size : 35 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • Immunohistochemical staining of paraffin embedded human stomach with purified ab32518 at a working dilution of 1 in 100. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

  • Immunohistochemical staining of paraffin embedded rat kidney with purified ab32518 at a working dilution of 1 in 100. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

  • ab32518 (purified) at 1/20 immunoprecipitating IKB alpha in HeLa cell lysate (Lane 1). For western blotting a HRP-conjugated goat anti-rabbit IgG was used as the secondary antibody (1/1000).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunofluorescence staining of HeLa cells with purified ab32518 at a working dilution of 1 in 50, counter-stained with DAPI. The secondary antibody was ab150077, Alexa Fluor® 488 goat anti rabbit, used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab32518 was used at a dilution of 1/50 followed by ab150120, Alexa Fluor® 594 goat anti-mouse antibody at a dilution of 1/500.

  • Anti-IKB alpha antibody [E130] (ab32518) at 1/10000 dilution (unpurified) + HeLa cell lysate

    Predicted band size : 36 kDa
    Observed band size : 35 kDa (why is the actual band size different from the predicted?)
  • Unpurified ab32518 used to immunoprecipitate IKB alpha from human HeLa whole cell lysate. The antibody was further used to Western blot the protein.

    Lane 1 IKB alpha IP

    Lane 2 Control immunoprecipitate

    Lane 3 Input (20%)

    See Abreview

  • Unpurified ab32518 used to immunoprecipitate IKB alpha from rat PC12 whole cell lysate. The antibody was further used to Western blot the protein.

    Lane 1 IKB alpha IP

    Lane 2 Control immunoprecipitate

    Lane 3 Input (20%)

    See Abreview

  • Unpurified ab32518 at 1/100 staining mouse kidney tissue sections by IHC-P. The tissue was paraformaldehyde fixed and a heat mediated antigen retrieval step in citrate buffer was performed before the tissue was blocked and incubated with the antibody for 1 hour. An HRP conjugated goat anti-rabbit antibody was used as the secondary.

    See Abreview

  • Unpurified ab32518 staining IkBα/&beta in RAW 264.7 cells treated with FK506 (ab120223), by ICC/IF. Decrease in IkBα/&beta expression correlates with increased concentration of FK506, as described in literature.
    The cells were incubated at 37°C for 3h in media containing different concentrations of ab120223 (FK506) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab32518 (1/100 dilution) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight® 488 goat anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

Anti-IKB alpha antibody [E130] (ab32518)参考文献

This product has been referenced in:
  • Sinha K  et al. Morin protects gastric mucosa from nonsteroidal anti-inflammatory drug, indomethacin induced inflammatory damage and apoptosis by modulating NF-?B pathway. Biochim Biophys Acta 1850:769-783 (2015). Read more (PubMed: 25603542) »
  • Tang YL  et al. TLR4/NF-?B signaling contributes to chronic unpredictable mild stress-induced atherosclerosis in ApoE-/- mice. PLoS One 10:e0123685 (2015). WB ; Mouse . Read more (PubMed: 25860573) »

See all 28 Publications for this product

Product Wall

Application Western blot
Sample Rat Tissue lysate - whole (Skeletal muscle)
Gel Running Conditions Reduced Denaturing (12%)
Loading amount 20 µg
Specification Skeletal muscle
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
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Abcam user community

Verified customer

提交于 Feb 18 2016

Application Western blot
Sample Mouse Tissue lysate - whole (Kidney)
Gel Running Conditions Reduced Denaturing
Loading amount 20 µg
Treatment 28 min IRI
Specification Kidney
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Username

Dr. Wesley Konsavage

Verified customer

提交于 Sep 15 2015

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
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Verified customer

提交于 Aug 31 2015

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12% gel)
Sample Mouse Tissue lysate - whole (Skeletal muscle (AT))
Specification Skeletal muscle (AT)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Mar 27 2014

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12%)
Sample Rat Tissue lysate - whole (Liver)
Specification Liver
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

提交于 Jun 17 2013

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12%)
Sample Mouse Tissue lysate - whole (Liver)
Specification Liver
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

提交于 Jun 17 2013

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12%)
Sample Human Tissue lysate - whole (Liver)
Specification Liver
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

提交于 Jun 17 2013



Selon Uniprot: http://www.uniprot.org/uniprot/P25963 la protéine peut être phosphoryle sur 8 résidus.

Pour ab5682 qui détecte la protéine phosphoryle sur deux résidus, le poids molécul...

Read More

ab32518 recognises both the phosphorylated and non-phosphorylated forms of the serine 32 region of IKB alpha. This means that the band at 37 kDa represents the unphosphorylated protein and the band at ˜47 kDa is likely to be due to phosphorylations of ...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (smooth muscle cells)
Loading amount 25 µg
Specification smooth muscle cells
Treatment infection with interested gene for 48 hours
Gel Running Conditions Reduced Denaturing (10%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Jan 16 2013

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