Overview

  • Product nameAnti-Bad antibody [Y208]
    See all Bad primary antibodies
  • Description
    Rabbit monoclonal [Y208] to Bad
  • SpecificityThe antibody does not cross-react with other Bcl-2 members.
  • Tested applicationsSuitable for: WB, IHC-P, ICC/IF, Flow Cyt, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Dog, Human
  • Immunogen

    A synthetic peptide corresponding to residues in the N-term of human Bad.

  • Positive control
    • Hela cell lysate, human lymphoma.
  • General notes

    This product is a recombinant rabbit monoclonal antibody.

     

    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.

    Alternative versions available:

    Anti-Bad antibody (Biotin) [Y208] (ab199754)

Properties

Applications

Our Abpromise guarantee covers the use of ab32445 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 1/2000 - 1/5000. Detects a band of approximately 23 kDa (predicted molecular weight: 18 kDa).
IHC-P Use at an assay dependent concentration.
ICC/IF 1/250 - 1/500.
Flow Cyt 1/20 - 1/50.

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

IP 1/100.

Target

  • FunctionPromotes cell death. Successfully competes for the binding to Bcl-X(L), Bcl-2 and Bcl-W, thereby affecting the level of heterodimerization of these proteins with BAX. Can reverse the death repressor activity of Bcl-X(L), but not that of Bcl-2 (By similarity). Appears to act as a link between growth factor receptor signaling and the apoptotic pathways.
  • Tissue specificityExpressed in a wide variety of tissues.
  • Sequence similaritiesBelongs to the Bcl-2 family.
  • DomainIntact BH3 motif is required by BIK, BID, BAK, BAD and BAX for their pro-apoptotic activity and for their interaction with anti-apoptotic members of the Bcl-2 family.
  • Post-translational
    modifications
    Phosphorylated on one or more of Ser-75, Ser-99, Ser-118 and Ser-134 in response to survival stimuli, which blocks its pro-apoptotic activity. Phosphorylation on Ser-99 or Ser-75 promotes heterodimerization with 14-3-3 proteins. This interaction then facilitates the phosphorylation at Ser-118, a site within the BH3 motif, leading to the release of Bcl-X(L) and the promotion of cell survival. Ser-99 is the major site of AKT/PKB phosphorylation, Ser-118 the major site of protein kinase A (CAPK) phosphorylation. Ser-75 is phosphorylated by AKT/PKB, protein kinase A and PIM2.
  • Cellular localizationMitochondrion outer membrane. Cytoplasm. Upon phosphorylation, locates to the cytoplasm.
  • Information by UniProt
  • Database links
  • Alternative names
    • AI325008 antibody
    • BAD antibody
    • BAD_HUMAN antibody
    • BBC 2 antibody
    • BBC2 antibody
    • BBC6 antibody
    • Bcl 2 Antagonist of Cell Death antibody
    • Bcl 2 Binding Component 6 antibody
    • BCL X / BCL 2 Binding Protein antibody
    • BCL X Binding Protein antibody
    • Bcl XL/Bcl 2 Associated Death Promoter antibody
    • Bcl-2-binding component 6 antibody
    • Bcl-2-like protein 8 antibody
    • Bcl-XL/Bcl-2-associated death promoter antibody
    • Bcl2 antagonist of cell death antibody
    • BCL2 antagonist of cell death protein antibody
    • BCL2 associated agonist of cell death antibody
    • Bcl2 Associated Death Promoter antibody
    • BCL2 binding component 6 antibody
    • BCL2 binding protein antibody
    • Bcl2 Like 8 Protein antibody
    • Bcl2-L-8 antibody
    • BCL2L8 antibody
    • Proapoptotic BH3 Only Protein antibody
    see all

Anti-Bad antibody [Y208] images

  • Anti-Bad antibody [Y208] (ab32445) at 1/5000 dilution + Hela cell lysate

    Predicted band size : 18 kDa
    Observed band size : 23 kDa (why is the actual band size different from the predicted?)
  • Immunohistochemical staining of paraffin-embedded human lymphoma using ab32445 at 1/100 dilution.
  • ICC/IF image of ab32445 stained MCF7 cells. The cells were 4% PFA fixed (10 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 (ab32445, 1µg/ml) 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.

  • Overlay histogram showing MCF-7 cells stained with ab32445 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab32445, 1/20 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (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 decreased signal in MCF-7 cells fixed with methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

References for Anti-Bad antibody [Y208] (ab32445)

This product has been referenced in:
  • Sun Y  et al. Up-regulation of eEF1A2 promotes proliferation and inhibits apoptosis in prostate cancer. Biochem Biophys Res Commun 450:1-6 (2014). WB ; Human . Read more (PubMed: 24853801) »
  • Sachewsky N  et al. Cyclosporin A enhances neural precursor cell survival in mice through a calcineurin-independent pathway. Dis Model Mech 7:953-61 (2014). WB ; Mouse . Read more (PubMed: 25056698) »

See all 12 Publications for this product

Product Wall

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Application Immunocytochemistry
Sample Mouse Cultured Cells (RAW 264.7)
Specification RAW 264.7
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton-X100 in 2% BSA for 15min
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 22°C
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted May 19 2012

Thank you for contacting us.
There is a wide range of secondaries suitable to detect either ab32445 or ab33862 in Flow Cytometry. I have conducted a search through our Secondaries search tool in the Abcam Website and found more than 100 matches, d...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (Glioblastoma neurospheres cells (primary))
Loading amount 30 µg
Specification Glioblastoma neurospheres cells (primary)
Gel Running Conditions Reduced Denaturing (12.5% gel)
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Username

Abcam user community

Verified customer

Submitted Dec 06 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (Platelets)
Loading amount 20 µg
Specification Platelets
Treatment Calcium Ionophore for 1 h
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
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted Nov 14 2011

Thank you for contacting us. I can confirm that this antibody does recongize the phosphorylated and unphosphorylated form of BAD. Indeed, there is no phosphorylation site on the immunogen sequences used for this antibody. I hope this inf...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Flow Cytometry
Sample Human Cell (Mononuclear Cell Preparations)
Specification Mononuclear Cell Preparations
Fixation Paraformaldehyde
Permeabilization Yes - 0.1% Triton-X100 in 2% BSA for 15min
Gating Strategy Monocytes Lymphocytes
Username

Dr. Mahesh Shivananjappa

Verified customer

Submitted Mar 30 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (PMN)
Specification PMN
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton-X100 in 2% BSA for 30min
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Dec 20 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunoprecipitation
Sample Human Cell lysate - whole cell (293T)
Total protein in input 200 µg
Specification 293T
Immuno-precipitation step Protein A
Username

Abcam user community

Verified customer

Submitted Dec 31 2007

1-10 of 13 Abreviews or Q&A

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