Validated using a knockout cell line
Recombinant
RabMAb

Anti-Bcl-2 antibody [E17] - BSA and Azide free (ab185002)

Overview

  • Product name
    Anti-Bcl-2 antibody [E17] - BSA and Azide free
    See all Bcl-2 primary antibodies
  • Description
    Rabbit monoclonal [E17] to Bcl-2 - BSA and Azide free
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, ICC/IF, IP, IHC-Pmore details
    Unsuitable for: Flow Cyt
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human Bcl-2 aa 50-150. The exact sequence is proprietary.

  • Positive control
    • Jurkat whole cell lysate (ab7899) and human breast carcinoma.
  • General notes

    The formulation and the concentration of this product is compatible for metal-conjugation for mass cytometry (CyTOF®).

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    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.

Applications

Our Abpromise guarantee covers the use of ab185002 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 Use at an assay dependent concentration. Predicted molecular weight: 26 kDa.

Please check the parent abID, ab32124, for a recommended dilution.

ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

  • Application notes
    Is unsuitable for Flow Cyt.
  • Target

    • Function
      Suppresses apoptosis in a variety of cell systems including factor-dependent lymphohematopoietic and neural cells. Regulates cell death by controlling the mitochondrial membrane permeability. Appears to function in a feedback loop system with caspases. Inhibits caspase activity either by preventing the release of cytochrome c from the mitochondria and/or by binding to the apoptosis-activating factor (APAF-1). May attenuate inflammation by impairing NLRP1-inflammasome activation, hence CASP1 activation and IL1B release (PubMed:17418785).
    • Tissue specificity
      Expressed in a variety of tissues.
    • Involvement in disease
      A chromosomal aberration involving BCL2 has been found in chronic lymphatic leukemia. Translocation t(14;18)(q32;q21) with immunoglobulin gene regions. BCL2 mutations found in non-Hodgkin lymphomas carrying the chromosomal translocation could be attributed to the Ig somatic hypermutation mechanism resulting in nucleotide transitions.
    • Sequence similarities
      Belongs to the Bcl-2 family.
    • Domain
      BH1 and BH2 domains are required for the interaction with BAX and for anti-apoptotic activity.
      The BH4 motif is required for anti-apoptotic activity and for interaction with RAF1 and EGLN3.
      The loop between motifs BH4 and BH3 is required for the interaction with NLRP1.
    • Post-translational
      modifications
      Phosphorylation/dephosphorylation on Ser-70 regulates anti-apoptotic activity. Growth factor-stimulated phosphorylation on Ser-70 by PKC is required for the anti-apoptosis activity and occurs during the G2/M phase of the cell cycle. In the absence of growth factors, BCL2 appears to be phosphorylated by other protein kinases such as ERKs and stress-activated kinases. Phosphorylated by MAPK8/JNK1 at Thr-69, Ser-70 and Ser-87, wich stimulates starvation-induced autophagy. Dephosphorylated by protein phosphatase 2A (PP2A).
      Proteolytically cleaved by caspases during apoptosis. The cleaved protein, lacking the BH4 motif, has pro-apoptotic activity, causes the release of cytochrome c into the cytosol promoting further caspase activity.
      Monoubiquitinated by PARK2, leading to increase its stability. Ubiquitinated by SCF(FBXO10), leading to its degradation by the proteasome.
    • Cellular localization
      Mitochondrion outer membrane. Nucleus membrane. Endoplasmic reticulum membrane.
    • Information by UniProt
    • Database links
    • Alternative names
      • Apoptosis regulator Bcl 2 antibody
      • Apoptosis regulator Bcl-2 antibody
      • Apoptosis regulator Bcl2 antibody
      • AW986256 antibody
      • B cell CLL/lymphoma 2 antibody
      • B cell leukemia/lymphoma 2 antibody
      • Bcl-2 antibody
      • Bcl2 antibody
      • BCL2_HUMAN antibody
      • C430015F12Rik antibody
      • D630044D05Rik antibody
      • D830018M01Rik antibody
      • Leukemia/lymphoma, B-cell, 2 antibody
      • Oncogene B-cell leukemia 2 antibody
      • PPP1R50 antibody
      • Protein phosphatase 1, regulatory subunit 50 antibody
      see all

    Images

    • Anti-Bcl-2 antibody [E17] - BSA and Azide free (ab185002) + MCF-7 (human breast carcinoma) whole cell lysate at 10 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

      Predicted band size: 26 kDa


      Exposure time: 3 minutes


      Blocking buffer and concentration: 5% NFDM/TBST

      Diluting buffer and concentration: 5% NFDM/TBST

    • Methanol-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained for Bcl-2 using ab32124 at 1/200 dilution in ICC/IF, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor­ 488) preadsorbed (ab150081). Counter-stained with DAPI.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Formaldehyde-fixed, paraffin-embedded human DLBCL U2932 cell line xenograft tissue stained for Bcl-2 using ab32124 at 1/200 dilution in immunohistochemical analysis, followed by Goat anti-Rabbit IgG Alexa Fluor® 555.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Bcl-2 expression determined by immunohistochemical analyses of the 4 human UM xenografts (between 3 to 5 tumors have been studied per condition).

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • ab32124 (purified) at 1/30 immunoprecipitating Bcl-2 in Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate (Lane 1). Lane 2 - PBS. 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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma cell line) cells labelling Bcl-2 with purified ab32124 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. The cells were co-stained with ab7291, a mouse anti-tubulin (1/500) using ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500) as the secondary antibody. Nuclei counterstained with DAPI (blue). 

      Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).

      Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human B-cell lymphoma tissue labelling Bcl-2 with purified ab32124 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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma cell line) cells labelling Bcl-2 with unpurified ab32124 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. The cells were co-stained with ab7291, a mouse anti-tubulin antibody (1/500) using ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500) as the secondary. Nuclei counterstained with DAPI (blue).

      Control: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling Bcl-2 with unpurified ab32124 at 1/200 dilution.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human B cell lymphoma tissue labelling Bcl-2 with unpurified ab32124.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Immunohistochemical analysis of Human salivary glands taken from patients with primary Sjögren's syndrome, staining Bcl-2 with unpurified ab32124.
      Antigen retrieval was performed via heat mediation in a citrate buffer (pH 6). Sections were blocked using 2% BSA, 10% normal serum and permeabilized with 0.5% Triton X-100. Samples were incubated with primary antibody (1/100) for one hour at room temperature. An Alexa Fluor® 594-conjugated anti-rabbit IgG was used as the secondary antibody.
      N.B. Panels B and D are higher magnifications of panels A and C, respectively.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    • Equilibrium disassociation constant (KD)

      Click here to learn more about KD

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

    References

    ab185002 has not yet been referenced specifically in any publications.

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