Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-BIN1 antibody [EPR13463] (ab182562)

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Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR13463] to BIN1
  • Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, IP
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-BIN1 antibody [EPR13463]
    See all BIN1 primary antibodies

  • Description

    Rabbit monoclonal [EPR13463] to BIN1

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, IPmore details

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment within Human BIN1 aa 400 to the C-terminus (C terminal). The exact sequence is proprietary.
    Database link: O00499

  • Positive control

    • IHC-P: human ovarian carcinoma, skeletal muscle, clear cell kidney carcinoma and brain tissue; ICC/IF: HeLa cells; Flow Cyt: HeLa cells; IP: HeLa and U87-MG cells. WB: A431, HeLa, HAP1, Human fetal kidney and skeletal muscle cell lysates ICC/IF: U87-MG and HeLa cells

  • 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.

    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.

Properties

Applications

Our Abpromise guarantee covers the use of ab182562 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 1/1000 - 1/10000. Detects a band of approximately 65, 70 kDa (predicted molecular weight: 65 kDa).
IHC-P 1/100 - 1/500.

Perform heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0).
ICC/IF 1/250.
Flow Cyt 1/30 - 1/40.

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

 
IP 1/20 - 1/40.

Target

  • Function

    May be involved in regulation of synaptic vesicle endocytosis. May act as a tumor suppressor and inhibits malignant cell transformation.

  • Tissue specificity

    Ubiquitous. Highest expression in the brain and muscle. Isoform IIA is expressed only in the brain where it is concentrated in axon initial segments and nodes of Ranvier. Isoform BIN1 is widely expressed with highest expression in skeletal muscle.

  • Involvement in disease

    Defects in BIN1 are the cause of centronuclear myopathy autosomal recessive (ARCNM) [MIM:255200]; also known as autosomal recessive myotubular myopathy. Centronuclear myopathies are congenital muscle disorders characterized by progressive muscular weakness and wasting involving mainly limb girdle, trunk, and neck muscles. It may also affect distal muscles. Weakness may be present during childhood or adolescence or may not become evident until the third decade of life. Ptosis is a frequent clinical feature. The most prominent histopathologic features include high frequency of centrally located nuclei in muscle fibers not secondary to regeneration, radial arrangement of sarcoplasmic strands around the central nuclei, and predominance and hypotrophy of type 1 fibers.

  • Sequence similarities

    Contains 1 BAR domain.
    Contains 1 SH3 domain.

  • Post-translational
    modifications

    Phosphorylated by protein kinase C.

  • Cellular localization

    Cytoplasm and Nucleus.
  • Information by UniProt

  • Database links

    • Alternative names

      • AMPH 2 antibody
      • AMPH2 antibody
      • Amphiphysin 2 antibody
      • Amphiphysin II antibody
      • Amphiphysin like protein antibody
      • amphiphysin-like antibody
      • Amphiphysin-like protein antibody
      • AMPHL antibody
      • Bin1 antibody
      • BIN1_HUMAN antibody
      • Box Dependant MYC Interacting Protein 1 antibody
      • Box-dependent myc-interacting protein 1 antibody
      • Bridging integrator 1 antibody
      • DKFZp547F068 antibody
      • MGC10367 antibody
      • MGC105358 antibody
      • Myc box dependent interacting protein 1 antibody
      • Myc box-dependent-interacting protein 1 antibody
      • SH3P9 antibody
      see all

    Images

    • Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)

      Lane 1: Wild type HAP1 whole cell lysate (20 µg)
      Lane 2: BIN1 (KO) knockout HAP1 whole cell lysate (20 µg)
      Lane 3: HeLa whole cell lysate (20 µg)
      Lane 4: U-87MG whole cell lysate (20 µg)

      Lanes 1 - 4: Merged signal (red and green). Green - ab182562 observed at 50-65 kDa. Red - loading control, ab181602, observed at 37 kDa.

      ab182562 (unpurified) was shown to specifically react with BIN1 when BIN1 knockout samples were used. Wild-type and BIN1 knockout samples were subjected to SDS-PAGE. Ab182562 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1 µg/ml 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 1/20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human ovarian carcinoma tissue sections labeling BIN1 with purified ab182562 at 1/500 dilution (0.52 µg/ml). Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

    • Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      All lanes : Anti-BIN1 antibody [EPR13463] (ab182562) at 1/10000 dilution (Purified)

      Lane 1 : Human brain lysates
      Lane 2 : Human muscle lysates

      Lysates/proteins at 15 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

      Predicted band size: 65 kDa
      Observed band size: 50-65 kDa
      why is the actual band size different from the predicted?



      The multiple bands are isoforms produced by alternative splicing

    • Immunoprecipitation - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunoprecipitation - Anti-BIN1 antibody [EPR13463] (ab182562)
      ab182562 (purified) at 1/20 dilution (1 µg) immunoprecipitating BIN1 in HeLa whole cell lysate.
      Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10 µg
      Lane 2 (+): ab182562 & HeLa whole cell lysate
      Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab182562 in HeLa whole cell lysate
      For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was at 1/1000 dilution.
      Blocking and diluting buffer: 5% NFDM/TBST.
    • Flow Cytometry - Anti-BIN1 antibody [EPR13463] (ab182562)
      Flow Cytometry - Anti-BIN1 antibody [EPR13463] (ab182562)
      Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling BIN1 with purified ab182562 at 1/30 dilution (10 µg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
    • Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling BIN1 with purified ab182562 at 1:250 dilution (1 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
    • Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunofluorescent analysis of HeLa cells labeling BIN1 with ab182562 (unpurified) at 1/250 and Goat anti rabbit IgG(Alexa Fluor®555) at 1/200.  Cells were fixed with -20℃ Acetone. Image at the right stained with DAPI.

    • Immunoprecipitation - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunoprecipitation - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunoprecipitation. ab182562 (unpurified) at 1/10000 staining BIN1 in U87-MG cell lysate immunoprecipitated using ab182562 at 1/50.
      Lane 2: negative control.

    • Flow Cytometry - Anti-BIN1 antibody [EPR13463] (ab182562)
      Flow Cytometry - Anti-BIN1 antibody [EPR13463] (ab182562)

      Flow Cytometrical analysis of 2% paraformaldehyde fixed HeLa cells labeling BIN1 with ab182562 (unpurified) at 1/40.

    • Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunocytochemistry/ Immunofluorescence - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunofluorescent analysis of U87-MG cells labeling BIN1 with ab182562 (unpurified) at 1/250 and Goat anti rabbit IgG(Alexa Fluor®555) at 1/200. Cells were fixed with 4% paraformaldehyde. Image at the right stained with DAPI.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunohistochemical analysis of paraffin embedded Human brain tissue labeling BIN1 with ab182562 (unpurified) at 1/100.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunohistochemical analysis of paraffin embedded Human clear cell carcinoma of kidney tissue labeling BIN1 with ab182562 (unpurified) at 1/100.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BIN1 antibody [EPR13463] (ab182562)

      Immunohistochemical analysis of paraffin embedded Human skeletal muscle tissue labeling BIN1 with ab182562 (unpurified) at 1/100.

    • Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      All lanes : Anti-BIN1 antibody [EPR13463] (ab182562) at 1/5000 dilution (unpurified)

      Lane 1 : Human skeletal muscle
      Lane 2 : Human fetal kidney

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG HRP (ab136636) at 1/500 dilution

      Predicted band size: 65 kDa
      Additional bands at: 56, 65, 70 kDa. We are unsure as to the identity of these extra bands.

    • Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      Western blot - Anti-BIN1 antibody [EPR13463] (ab182562)
      All lanes : Anti-BIN1 antibody [EPR13463] (ab182562) at 1/5000 dilution (unpurified)

      Lane 1 : A431 cell lysate
      Lane 2 : HeLa cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

      Predicted band size: 65 kDa
      Additional bands at: 65, 70 kDa. We are unsure as to the identity of these extra bands.

    References

    Publishing research using ab182562? Please let us know so that we can cite the reference in this datasheet.

    ab182562 has been referenced in 1 publication.

    • Ge Y  et al. The splicing factor RBM25 controls MYC activity in acute myeloid leukemia. Nat Commun 10:172 (2019). PubMed: 30635567

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    Immunocytochemistry/ Immunofluorescence abreview for Anti-BIN1 antibody [EPR13463]

     Excellent
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (iPS cell derived neurons)
    Permeabilization
    Yes - see blocking buffer
    Specification
    iPS cell derived neurons
    Blocking step
    Serum as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: rt°C
    Fixative
    Paraformaldehyde
    Read More

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    Submitted Feb 02 2016

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