Overview

  • Product name
    Anti-WIPI2 antibody [2A2]
    See all WIPI2 primary antibodies
  • Description
    Mouse monoclonal [2A2] to WIPI2
  • Host species
    Mouse
  • Tested applications
    Suitable for: ICC/IF, IP, IHC-P, IHC-Fr, WB, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide:

    CSALRLDEDSEHPPMILRTD

    , corresponding to C terminal amino acids 418-436 of Human WIPI2 (Isoform WIPI2b).

  • Epitope
    EHPPM
  • Positive control
    • This antibody gave a positive signal in Human Skeletal Muscle, Human Placenta, Mouse Placenta, Mouse Testis as well as the following whole cell lysates: HeLa, and NIH3T3. IF/ICC: HeLa cells (50mM chloroquine for 24h) Flow Cyt: HeLa cells IHC-P: Human skeletal muscle (normal)
  • General notes

    This antibody clone is manufactured by Abcam. We welcome customer feedback and would appreciate any comments regarding this product and the data presented.

    If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
  • Storage buffer
    Preservative: 0.02% Sodium azide
    Constituent: PBS
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Clonality
    Monoclonal
  • Clone number
    2A2
  • Isotype
    IgG1
  • Light chain type
    kappa

Applications

Our Abpromise guarantee covers the use of ab105459 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
ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.
IHC-Fr Use at an assay dependent concentration. Fix with Acetone.
WB Use at an assay dependent concentration. Predicted molecular weight: 49 kDa.
Flow Cyt Use 0.1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 

Target

  • Function
    Probable early component of the autophagy machinery being involved in formation of preautophagosomal structures and their maturation into mature phagosomes in response to PtdIns3P. May bind PtdIns3P.
  • Tissue specificity
    Ubiquitously expressed (at protein level). Highly expressed in heart, skeletal muscle and pancreas. Expression is down-regulated in pancreatic and in kidney tumors.
  • Sequence similarities
    Belongs to the WD repeat SVP1 family.
    Contains 3 WD repeats.
  • Cellular localization
    Preautophagosomal structure membrane. Enriched at preautophagosomal structure membranes in response to ptdIns3P.
  • Information by UniProt
  • Database links
  • Alternative names
    • ATG18B antibody
    • Atg21 antibody
    • CGI 50 antibody
    • DKFZp434J154 antibody
    • DKFZp686P02188 antibody
    • FLJ12979 antibody
    • FLJ14217 antibody
    • FLJ42984 antibody
    • WD repeat domain phosphoinositide-interacting protein 2 antibody
    • WD repeat domain, phosphoinositide interacting 2 antibody
    • WD40 repeat protein interacting with phosphoinositides 2 antibody
    • WIPI 2 antibody
    • WIPI-2 antibody
    • WIPI2 antibody
    • WIPI2_HUMAN antibody
    • WIPI49 like protein 2 antibody
    • WIPI49-like protein 2 antibody
    see all

Images

  • All lanes : Anti-WIPI2 antibody [2A2] (ab105459) at 1 µg/ml

    Lane 1 : Human skeletal muscle tissue lysate
    Lane 2 : Human placenta tissue lysate
    Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 4 : NIH 3T3 (Mouse) Whole Cell Lysate
    Lane 5 : Pancreas (Mouse) Tissue Lysate
    Lane 6 : Testis (Mouse) Tissue Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) (ab65485) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 49 kDa


    Exposure time: 1 minute


    Abcam recommends using milk as the blocking agent.

  • IHC image of WIPI2 staining in a section of formalin-fixed paraffin-embedded normal human skeletal muscle* performed on a Leica BONDTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab105459, 1ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • All lanes : Anti-WIPI2 antibody [2A2] (ab105459) at 1/1000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : Wipi2 knockout HAP1 whole cell lysate
    Lane 3 : Saos2 whole cell lysate
    Lane 4 : CACO2 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 49 kDa
    Observed band size: 49 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab105459 observed at 49 kDa. Red - loading control, ab181602, observed at 37 kDa.

    ab105459 was shown to recognize WIPI2 in wild-type HAP1 cells as signal was lost at the expected MW in Wipi2 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and Wipi2 knockout samples were subjected to SDS-PAGE. Ab105459 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

  • ab105459 staining WIPI2 in HeLa cells. The cells were treated with 50uM chloroquine for 24h and then fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab105459 at 1ugml then detected with an Alexa Fluor® 488 goat anti-rabbit secondary antibody (ab150081) at a 1/1000 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue), and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at a 1/250 dilution (shown in red).

  • Overlay histogram showing HeLa cells stained with ab105459 (red line). The cells were fixed with 80% 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 (ab105459, 0.1μg/1x106 cells) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H+L) (ab150113) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

References

This product has been referenced in:
  • Lee MY  et al. Peroxisomal protein PEX13 functions in selective autophagy. EMBO Rep 18:48-60 (2017). Human . Read more (PubMed: 27827795) »
  • Jung J  et al. Multiplex image-based autophagy RNAi screening identifies SMCR8 as ULK1 kinase activity and gene expression regulator. Elife 6:N/A (2017). WB, ICC/IF ; Human . Read more (PubMed: 28195531) »

See all 14 Publications for this product

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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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