重组
RabMAb

Anti-IKB alpha抗体[E130] (ab32518)

概述

  • 产品名称
    Anti-IKB alpha抗体[E130]
    参阅全部 IKB alpha 一抗
  • 描述
    兔单克隆抗体[E130] to IKB alpha
  • 特异性
    This antibody detects both the phosphorylated and non-phosphorylated form of the serine 32 region of IKB alpha.
  • 经测试应用
    适用于: ICC/IF, WB, IHC-P, IP, Flow Cytmore details
  • 种属反应性
    与反应: Mouse, Rat, Hamster, Human
    预测可用于: Cow, Pig
  • 免疫原

    Synthetic peptide within Human IKB alpha. The exact sequence is proprietary.
    Database link: P25963

  • 阳性对照
    • Hela cell lysate and human prostate carcinoma tissue.
  • 常规说明

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

    This product is a recombinant rabbit monoclonal antibody.

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.
Flow Cyt 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

  • 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

  • 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.

  • 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.

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

  • Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling IKB alpha with purified ab32518 at 1/20 dilution (10ug/mL) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

  • 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

  • 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.

  • 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.

  • 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

  • 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.

  • 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:
  • Vander Lugt B  et al. Transcriptional determinants of tolerogenic and immunogenic states during dendritic cell maturation. J Cell Biol 216:779-792 (2017). WB ; Mouse . Read more (PubMed: 28130292) »
  • Hu J  et al. BMSC paracrine activity attenuates interleukin-1ß-induced inflammation and apoptosis in rat AF cells via inhibiting relative NF-?B signaling and the mitochondrial pathway. Am J Transl Res 9:79-89 (2017). WB ; Rat . Read more (PubMed: 28123635) »

See all 47 Publications for this product

Product Wall

Application
Western blot
Sample
Mouse Cell lysate - whole cell (Endothelial)
Gel Running Conditions
Reduced Denaturing
Loading amount
10 µg
Specification
Endothelial
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

提交于 Jul 28 2017

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
Username

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
Username

Abcam user community

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

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