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RabMAb

Anti-Caspase-9抗体[E23] (ab32539)

概述

  • 产品名称
    Anti-Caspase-9抗体[E23]
    参阅全部 Caspase-9 一抗
  • 描述
    兔单克隆抗体[E23] to Caspase-9
  • 特异性
    This antibody should recognise both the pro-[40kDa] form and p35 cleaved form of Caspase-9.
  • 经测试应用
    适用于: WB, IHC-P, Flow Cyt, IP, ICC/IFmore details
  • 种属反应性
    与反应: Human
  • 免疫原

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human Caspase-9 aa 250-350.
    Database link: P55211

  • 阳性对照
    • WB: Jurkat and HeLa cell lysates. IHC-P: Human skeletal muscle and cervical carcinoma tissues. ICC/IF: HepG2 cells. Flow Cyt: K562 cells. IP: HeLa whole cell lysate.
  • 常规说明

    This product is a recombinant rabbit monoclonal antibody.

    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.

    Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

    A trial size is available to purchase for this antibody.

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

性能

应用

Our Abpromise guarantee covers the use of ab32539 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
WB 1/1000 - 1/10000. Predicted molecular weight: 46 kDa.

We recommend overnight incubation at 4°C.

IHC-P 1/50 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See protocols (link: http://www.abcam.com/protocols/ihc-antigen-retrieval-protocol).

Flow Cyt 1/100 - 1/250.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IP 1/80 - 1/100.
ICC/IF 1/500.

For unpurified use at 1/50.

靶标

  • 功能
    Involved in the activation cascade of caspases responsible for apoptosis execution. Binding of caspase-9 to Apaf-1 leads to activation of the protease which then cleaves and activates caspase-3. Proteolytically cleaves poly(ADP-ribose) polymerase (PARP).
    Isoform 2 lacks activity is an dominant-negative inhibitor of caspase-9.
  • 组织特异性
    Ubiquitous, with highest expression in the heart, moderate expression in liver, skeletal muscle, and pancreas. Low levels in all other tissues. Within the heart, specifically expressed in myocytes.
  • 序列相似性
    Belongs to the peptidase C14A family.
    Contains 1 CARD domain.
  • 发展阶段
    Expressed at low levels in fetal heart, at moderate levels in neonate heart, and at high levels in adult heart.
  • 翻译后修饰
    Cleavages at Asp-315 by granzyme B and at Asp-330 by caspase-3 generate the two active subunits. Caspase-8 and -10 can also be involved in these processing events.
  • Information by UniProt
  • 数据库链接
  • 别名
    • APAF-3 antibody
    • APAF3 antibody
    • Apoptosis related cysteine peptidase antibody
    • Apoptotic protease Mch-6 antibody
    • Apoptotic protease-activating factor 3 antibody
    • CASP-9 antibody
    • CASP9 antibody
    • CASP9_HUMAN antibody
    • Caspase 9 apoptosis related cysteine peptidase antibody
    • Caspase 9 Dominant Negative antibody
    • Caspase 9c antibody
    • Caspase-9 antibody
    • Caspase-9 subunit p10 antibody
    • ICE LAP6 antibody
    • ICE like apoptotic protease 6 antibody
    • ICE-LAP6 antibody
    • ICE-like apoptotic protease 6 antibody
    • MCH6 antibody
    • PPP1R56 antibody
    • protein phosphatase 1, regulatory subunit 56 antibody
    • RNCASP9 antibody
    see all

Anti-Caspase-9 antibody [E23] 图像



  • Predicted band size : 46 kDa

    Lanes:
    Lane 1: Wild type HAP1 whole cell lysate (40 µg)
    Lane 2: CASP9 knockout  HAP1 whole cell lysate (40 µg)
    Lane 3: HeLa whole cell lysate (40 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab32539 observed at 45 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab32539 was shown to recognize Caspase-9 when Caspase-9 knockout samples were used, along with additional cross-reactive bands. Wild-type and Caspase-9 knockout samples were subjected to SDS-PAGE. Ab32539 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 500 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human skeletal muscle tissue labelling Caspase-9 with purified ab32539 at 1/250. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • Immunocytochemistry/Immunofluorescence analysis of HepG2 cells labelling Caspase-9 with purified ab32539 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

    Control 1: primary antibody (1/500) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).

    Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000).

  • Flow Cytometry analysis of K562 cells labelling Caspase-9 with purified ab32539 at 1/250 (red). Cells were fixed with 100% methanol. A FITC-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

  • ab32539 (purified) at 1/80 immunoprecipitating Caspase-9 in HeLa whole cell lysate.

    Lane 1 (input): HeLa whole cell lysate (10µg)

    Lane 2 (+): ab32539 + HeLa whole cell lysate.

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32539 in HeLa whole cell lysate.

    For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • All lanes : Anti-Caspase-9 antibody [E23] (ab32539) at 1/1000 dilution (purified)

    Lane 1 : HeLa whole cell lysate - treated with Camptothecin
    Lane 2 : HeLa whole cell lysate - untreated

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution

    Predicted band size : 46 kDa
    Observed band size : 35,46 kDa (why is the actual band size different from the predicted?)

    Blocking and dilution buffer: 5% NFDM/TBST.

  • Overlay histogram showing K562 cells stained with unpurified ab32539 (red line). The cells were fixed with methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab32539, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was anti-rabbit DyLight® 488 (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in K562 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween used under the same conditions.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue labelling Caspase 9 with unpurified ab32539 at a dilution of 1/50.

  • All lanes : Anti-Caspase-9 antibody [E23] (ab32539) at 1/20000 dilution (unpurified)

    Lane 1 : Jurkat cell lysate - untreated
    Lane 2 : Jurkat cell lysate - treated with Camptothecin


    Predicted band size : 46 kDa
    Observed band size : 48 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 28 kDa. We are unsure as to the identity of these extra bands.

Anti-Caspase-9 antibody [E23] (ab32539)参考文献

This product has been referenced in:
  • Zhang FQ  et al. Regulation of c-Jun N-Terminal Protein Kinase (JNK) Pathway in Apoptosis of Endothelial Outgrowth Cells Induced by Asymmetric Dimethylarginine. Med Sci Monit 23:2535-2542 (2017). WB ; Human . Read more (PubMed: 28546532) »
  • Razavi BM  et al. Protective effect of crocin against apoptosis induced by subchronic exposure of the rat vascular system to diazinon. Toxicol Ind Health 32:1237-45 (2016). Rat . Read more (PubMed: 27353299) »

See all 26 Publications for this product

Product Wall

Application
Western blot
Loading amount
25 µg
Gel Running Conditions
Reduced Denaturing
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: 23°C
Username

Abcam user community

Verified customer

提交于 Dec 22 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Fadu)
Loading amount
30 µg
Specification
Fadu
Treatment
RT8gray for 48hrs+Metformin400mM
Gel Running Conditions
Reduced Denaturing
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 Apr 24 2013


Full length caspase-9 will cleave into two units, caspase-9 p35 subunit and caspase-9 p10 subunit. The immunogen of ab32539 is a synthetic peptide corresponding to the C terminus of caspase-9 p35 subunit. Therefore, the antibody is able to detect ...

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Thank you for your inquiry.
I am happy to confirm that all these antibodies are tested and guaranteed to work with human samples.
You are correct that we do not have any reports yet about the use of these antibodies in muscle. I assume that i...

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La référence ab2014 n'a pas encore été testée chez le rat, c'est pour cela que j'ai préféré comparer les séquences.

Nous serions ravis si vous pouviez nous communiquer vos r&eacut...

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Merci pour votre réponse.

L'anticorps ab63488 est en effet garanti pour fonctionner en WB et IHC-P. Je suis d'accord avec vous, l'image du blotn'est pas très attractive.

Une autre référence qui a retenu m...

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Merci de nous avoir contactés de nouveau.

Je suis vraimentdésolé d'apprendre que le second lot de ab32359 ne fonctionne pas toujours pas comme attendu.

Afin de solutionnerce problème, je peux vous propo...

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As far as we know, rabbits do not have specific IgG subclasses like mice or
humans. All of our antibodies are simply considered to be rabbit IgG.
I hope this is helpful. Please contact me again if you have any further questions.

Je suis vraiment désolé d'apprendre que vous rencontrez un autre problème avec cet anticorps. Pourriez-vous s'il vous plait tester cet anticorps. Si il fonctionne je vous enverrai une nouvelle unité en prenant soin de faire vérifier le tube avant ex...

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Merci de nous avoir contactés. Nous sommes désolés que le produit ab32539 que vous avez reçu ne fonctionne pas comme attendu. Comme convenu j'ai mis en place l'envoi d'une unité de ab32539 d'un nouveau lot. Le numéro de commande de remplacement grat...

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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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