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  1. Link

    smacdiablo-antibody-y12-ab32023.pdf

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Cell Biology Apoptosis Receptors Death Receptors
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RecombinantRabMAb

Recombinant Anti-Smac/Diablo antibody [Y12] (ab32023)

  • Datasheet
Reviews (5)Q&A (1)References (20)

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Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)
  • Anti-Smac/Diablo antibody [Y12] (ab32023)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y12] to Smac/Diablo
  • Suitable for: WB, IHC-P, IP, ICC/IF
  • Reacts with: Human

Conjugates logo Related conjugates and formulations

Alexa Fluor® 488 Alexa Fluor® 555 Alexa Fluor® 647 Carrier Free

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Overview

  • Product name

    Anti-Smac/Diablo antibody [Y12]
    See all Smac/Diablo primary antibodies
  • Description

    Rabbit monoclonal [Y12] to Smac/Diablo
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IHC-P, IP, ICC/IFmore details
  • Species reactivity

    Reacts with: Human
    Predicted to work with: Mouse, Rat
  • Immunogen

    Synthetic peptide within Human Smac/ Diablo (C terminal). The exact sequence is proprietary.

  • Epitope

    ab32023 reacts with an epitope located in the C terminal region of Smac / Diablo.
  • Positive control

    • WB: Jurkat whole cell lysate (ab7899). IHC-P: Human skin cancer tissue. ICC/IF: HeLa cells. IP: Jurkat.
  • General notes

    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.

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: 49% PBS, 50% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    Y12
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Apoptosis
    • Receptors
    • Death Receptors
    • Cell Biology
    • Apoptosis
    • Mitochondrial
    • Cancer
    • Invasion/microenvironment
    • Apoptosis
    • Death receptors & ligands
    • IAPs
    • Metabolism
    • Pathways and Processes
    • Mitochondrial Metabolism
    • Mitochondrial markers
    • Metabolism
    • Pathways and Processes
    • Metabolism processes
    • Apoptosis
    • Cancer
    • Cell Death
    • Apoptosis
    • Receptors
    • Death receptors & ligands
    • IAPs
    • Cancer
    • Cell Death
    • Apoptosis
    • Mitochondrial
    • Cancer
    • Cell Death
    • Apoptosis
    • Metabolism

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-Smac/Diablo antibody [Y12] (ab199317)
    • Alexa Fluor® 647 Anti-Smac/Diablo antibody [Y12] (ab199503)
    • Alexa Fluor® 555 Anti-Smac/Diablo antibody [Y12] (ab214622)
    • Anti-Smac/Diablo antibody [Y12] - BSA and Azide free (ab239797)
  • Isotype control

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

    • Jurkat whole cell lysate (ab7899)
  • Recombinant Protein

    • Recombinant Human Smac/Diablo protein (His tag) (ab219716)
    • Recombinant Human Smac/Diablo protein (ab87721)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab32023 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
WB (3)
1/1000. Detects a band of approximately 21 kDa (predicted molecular weight: 27 kDa).
IHC-P
1/10. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
IP (1)
1/10 - 1/80.
ICC/IF
1/500.
Notes
WB
1/1000. Detects a band of approximately 21 kDa (predicted molecular weight: 27 kDa).
IHC-P
1/10. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
IP
1/10 - 1/80.
ICC/IF
1/500.

Target

  • Function

    Promotes apoptosis by activating caspases in the cytochrome c/Apaf-1/caspase-9 pathway. Acts by opposing the inhibitory activity of inhibitor of apoptosis proteins (IAP).
  • Tissue specificity

    Ubiquitously expressed with highest expression in testis. Expression is also high in heart, liver, kidney, spleen, prostate and ovary. Low in brain, lung, thymus and peripheral blood leukocytes.
  • Domain

    The mature N-terminus mediates interaction with XIAP.
  • Cellular localization

    Mitochondrion. Released into the cytosol when cells undergo apoptosis.
  • Target information above from: UniProt accession Q9NR28 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 56616 Human
    • Entrez Gene: 66593 Mouse
    • Entrez Gene: 288753 Rat
    • Omim: 605219 Human
    • SwissProt: Q9NR28 Human
    • SwissProt: Q9JIQ3 Mouse
    • Unigene: 169611 Human
    • Unigene: 46716 Mouse
    • Alternative names

      • 0610041G12Rik antibody
      • DBLOH_HUMAN antibody
      • DBOH antibody
      • DFNA64 antibody
      • diablo antibody
      • Diablo homolog (Drosophila) antibody
      • Diablo homolog antibody
      • Diablo homolog mitochondrial antibody
      • Diablo IAP binding mitochondrial protein antibody
      • Diablo like protein antibody
      • DIABLO S antibody
      • Direct IAP binding protein with low pI antibody
      • Direct IAP-binding protein with low pI antibody
      • FLJ10537 antibody
      • FLJ25049 antibody
      • mitochondrial antibody
      • Mitochondrial Smac protein antibody
      • Second mitochondria derived activator of caspase antibody
      • Second mitochondria-derived activator of caspase antibody
      • second mitochondrial activator of caspases antibody
      • SMAC 3 antibody
      • Smac antibody
      • Smac protein antibody
      • SMAC3 antibody
      see all

    Images

    • Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Anti-Smac/Diablo antibody [Y12] (ab32023) at 1/1000 dilution + Jurkat cell lysate

      Predicted band size: 27 kDa
      Observed band size: 21 kDa why is the actual band size different from the predicted?

    • Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)

      Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling Smac / Diablo with ab32023 at 1/500. Cells were fixed with 100% methanol. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.
      Control: PBS only.
      Nuclear counter stain: DAPI.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (ab32023)

      ab32023, at a 1/10 dilution, staining human Smac/Diablo in skin cancer tissue by immunohistochemistry, paraffin-embedded tissue.

      Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

    • Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (ab32023)Image from Lamers F et al., BMC Cancer. 2012 Jul 12;12:285. doi: 10.1186/1471-2407-12-285. Fig 3.; 12 July 2012, BMC Cancer 2012, 12:285

      Immunofluorescence analysis of IMR32 cells, staining Smac / Diablo with ab32023. Cells were fixed with 4% paraformaldehyde before incubating with primary antibody. An Alexa Fluor® conjugated anti-rabbit IgG was used as the secondary antibody.

    • Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)

      Smac/Diablo was immunoprecipitated from 0.35 mg Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg with ab32023 at 1/50 dilution (2μg). VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution. 

      Lane 1: Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg

      Lane 2: ab32023 IP in Jurkat whole cell lysate

      Lane 3:Rabbit monoclonal IgG (ab172730) instead of ab32023 in Jurkat whole cell lysate

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

    • Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)
      Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)This image is courtesy of an anonymous Abreview.

      Smac / Diablo immunprecipitated from mitochondria isolated from mice brains.
      Total protein in input: 800 µg.
      Immunoprecipitation step: Dynabeads protein G.
      Incubation: 16 hours, 4°C.
      Diluent: TNE buffer.

      For western blot, ab32023 was used at a 1/1000 dilution.

      Lanes:
      1. BioRad Precision Plus Protein standards, bands from top to bottom: 250, 150, 100, 75, 50, 37, 25, 20, 15, 10 kDa
      2. Mitochondria isolated from mice brains, lysate
      3. IP with anti-SMAC/DIABLO ab32023

      See Abreview

    • Anti-Smac/Diablo antibody [Y12] (ab32023)
      Anti-Smac/Diablo antibody [Y12] (ab32023)

    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

    • Datasheet download

      Download

    References (20)

    Publishing research using ab32023? Please let us know so that we can cite the reference in this datasheet.

    ab32023 has been referenced in 20 publications.

    • McKenna S  et al. BAX and SMAC regulate bistable properties of the apoptotic caspase system. Sci Rep 11:3272 (2021). PubMed: 33558564
    • Zhou H  et al. Upconversion NIR-II fluorophores for mitochondria-targeted cancer imaging and photothermal therapy. Nat Commun 11:6183 (2020). PubMed: 33273452
    • Xian H & Liou YC Loss of MIEF1/MiD51 confers susceptibility to BAX-mediated cell death and PINK1-PRKN-dependent mitophagy. Autophagy 15:2107-2125 (2019). PubMed: 30894073
    • Wu B  et al. Combination of Sodium Selenite and Doxorubicin Prodrug Ac-Phe-Lys-PABC-ADM Affects Gastric Cancer Cell Apoptosis in Xenografted Mice. Biomed Res Int 2019:2486783 (2019). PubMed: 31531348
    • Wei Q  et al. The mitochondrial pathway is involved in sodium fluoride (NaF)-induced renal apoptosis in mice. Toxicol Res (Camb) 7:792-808 (2018). PubMed: 30310657
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-6 of 6 Abreviews or Q&A

    Immunoprecipitation abreview for Anti-Smac / Diablo antibody [Y12]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunoprecipitation
    Sample
    Mouse Tissue lysate - other (Mitochondria isolated from mice brains, lysed by t)
    Total protein in input
    800 µg
    Immuno-precipitation step
    Other - Dynabeads protein G
    Specification
    Mitochondria isolated from mice brains, lysed by t
    Read More

    Abcam user community

    Verified customer

    Submitted Mar 05 2019

    Western blot abreview for Anti-Smac / Diablo antibody [Y12]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    20 µg
    Gel Running Conditions
    Reduced Denaturing (15% Gel)
    Sample
    Mouse Tissue lysate - other (Skeletal muscle cytosolic fractions)
    Specification
    Skeletal muscle cytosolic fractions
    Blocking step
    Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
    Read More

    Abcam user community

    Verified customer

    Submitted Feb 25 2015

    Western blot abreview for Anti-Smac / Diablo antibody [Y12]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    20 µg
    Gel Running Conditions
    Reduced Denaturing (15% Gel)
    Sample
    Rat Tissue lysate - other (Skeletal muscle cytosolic fractions)
    Specification
    Skeletal muscle cytosolic fractions
    Blocking step
    Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
    Read More

    Abcam user community

    Verified customer

    Submitted Feb 19 2015

    Western blot abreview for Anti-Smac / Diablo antibody [Y12]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (Platelets)
    Loading amount
    20 µg
    Specification
    Platelets
    Treatment
    ADP for 30 min
    Gel Running Conditions
    Reduced Denaturing (4-20%)
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Read More

    Abcam user community

    Verified customer

    Submitted Nov 23 2011

    Flow Cytometry abreview for Anti-Smac / Diablo antibody [Y12]

    Average
    Abreviews
    Abreviews
    abreview image
    Application
    Flow Cytometry
    Sample
    Human Cell (Platelets)
    Specification
    Platelets
    Preparation
    Cell harvesting/tissue preparation method: PL were isolated spinning Platelet rich plasma on Histopaque
    Sample buffer: PBS
    Fixation
    Paraformaldehyde
    Permeabilization
    Yes - 0.1% Triton-X100 in 2% BSA for 15min
    Gating Strategy
    Platelets
    Read More

    Abcam user community

    Verified customer

    Submitted Dec 31 2010

    Question

    Dear sirs

    Please I need a secondary antibody working with ab32023 on flowcytometer by dye (FITC and PE) please suggest



    Sincerely yours,

    Read More

    Abcam community

    Verified customer

    Asked on Mar 05 2012

    Answer

    Thank you for your inquiry.

    I chose two secondary antibodies for each conjugate. If you would like to review the whole selection of possible secondary antibodies,please use our secondary antibody search tool:

    https://www.abcam.com/index.html?pageconfig=productmap&cl=918

    anti rabbit conjugated with FITC:

    ab98502
    https://www.abcam.com/Donkey-Rabbit-IgG-HL-FITC-preadsorbed-ab98502.html (or use the following: https://www.abcam.com/Donkey-Rabbit-IgG-HL-FITC-preadsorbed-ab98502.html).

    ab98466
    https://www.abcam.com/Goat-Rabbit-IgG-Fc-FITC-preadsorbed-ab98466.html (or use the following: https://www.abcam.com/Goat-Rabbit-IgG-Fc-FITC-preadsorbed-ab98466.html).

    anti rabbit conjugated with PE:

    ab97070
    https://www.abcam.com/Goat-Rabbit-IgG-HL-PE-preadsorbed-ab97070.html (or use the following: https://www.abcam.com/Goat-Rabbit-IgG-HL-PE-preadsorbed-ab97070.html).

    ab98494
    https://www.abcam.com/Donkey-Fab2-Rabbit-IgG-HL-PE-pre-adsorbed-ab98494.html (or use the following: https://www.abcam.com/Donkey-Fab2-Rabbit-IgG-HL-PE-pre-adsorbed-ab98494.html).

    I can recommend to also review the new conjugates which have are much more stable than the old conjugates FITC and PE:

    https://www.abcam.com/index.html?pageconfig=resource&rid=12934

    For more general information please follow the following links:

    https://www.abcam.com/index.html?pageconfig=resource&rid=12922

    https://www.abcam.com/index.html?pageconfig=resource&rid=12825

    https://www.abcam.com/index.html?pageconfig=resource&rid=12722

    https://www.abcam.com/index.html?pageconfig=resource&rid=12956

    I hope this information is helpful. Please do not hesitate to contact me again with any further questions.

    Read More

    Abcam Scientific Support

    Answered on Mar 05 2012

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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