Recombinant
RabMAb

Anti-PABPN1 antibody [EP3000Y] (ab75855)

Overview

  • Product name
    Anti-PABPN1 antibody [EP3000Y]
    See all PABPN1 primary antibodies
  • Description
    Rabbit monoclonal [EP3000Y] to PABPN1
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ICC/IF, WB, IP, IHC-P, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human PABPN1 aa 1-100 (N terminal). The exact sequence is proprietary.

  • Positive control
    • WB: Raw264.7, MCF-7, 293T, Mouse spleen, Rat brain and HeLa cell lysates. ICC/IF: MCF-7 cells. Flow Cyt: MCF-7 cells. IHC-P: Squamous cell cervical carcinoma tissue, human bladder carcinoma, mouse kidney, rat kidney.
  • General notes

     

     

    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.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab75855 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 1/100 - 1/250.
WB 1/1000 - 1/10000. Predicted molecular weight: 33 kDa.
IP 1/30.

Fur unpurified use at 1:50.

IHC-P 1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.
For unpurified use at 1/100 -1/250.

Flow Cyt 1/40.

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

For unpurified use at 1:80.

Target

  • Function
    Involved in the 3'-end formation of mRNA precursors (pre-mRNA) by the addition of a poly(A) tail of 200-250 nt to the upstream cleavage product. Stimulates poly(A) polymerase (PAPOLA) conferring processivity on the poly(A) tail elongation reaction and controls also the poly(A) tail length. Increases the affinity of poly(A) polymerase for RNA. Is also present at various stages of mRNA metabolism including nucleocytoplasmic trafficking and nonsense-mediated decay (NMD) of mRNA. Cooperates with SKIP to synergistically activate E-box-mediated transcription through MYOD1 and may regulate the expression of muscle-specific genes. Binds to poly(A) and to poly(G) with high affinity. May protect the poly(A) tail from degradation.
  • Tissue specificity
    Ubiquitous.
  • Involvement in disease
    Defects in PABPN1 are the cause of oculopharyngeal muscular dystrophy (OPMD) [MIM:164300]. OPMD is a form of late-onset slowly progressive myopathy characterized by eyelid ptosis, dysphagia and, sometimes by other cranial and limb-muscle involvement.
  • Sequence similarities
    Contains 1 RRM (RNA recognition motif) domain.
  • Domain
    The RRM domain is essential for specific adenine bases recognition in the poly(A) tail but not sufficient for poly(A) binding.
  • Post-translational
    modifications
    Arginine dimethylation is asymmetric and involves PRMT1 and PRMT3. It does not influence the RNA binding properties.
  • Cellular localization
    Nucleus. Cytoplasm. Localized in cytoplasmic mRNP granules containing untranslated mRNAs. Shuttles between the nucleus and the cytoplasm but predominantly found in the nucleus. Its nuclear import may involve the nucleocytoplasmic transport receptor transportin and a RAN-GTP-sensitive import mechanism. Is exported to the cytoplasm by a carrier-mediated pathway that is independent of mRNA traffic. Nucleus; nuclear speckle. Colocalizes with SKIP and poly(A) RNA in nuclear speckles.
  • Information by UniProt
  • Database links
  • Alternative names
    • Nuclear poly(A)-binding protein 1 antibody
    • OPMD antibody
    • PAB2 antibody
    • PABII antibody
    • PABP 2 antibody
    • pABP-2 antibody
    • PABP2 antibody
    • PABP2_HUMAN antibody
    • PABPII antibody
    • Pabpn1 antibody
    • poly(A) binding protein nuclear 1 antibody
    • Poly(A)-binding protein 2 antibody
    • Poly(A)-binding protein II antibody
    • PolyA binding protein II antibody
    • Polyadenylate-binding nuclear protein 1 antibody
    • Polyadenylate-binding protein 2 antibody
    see all

Images

  • Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling PABPN1 with purified ab75855 at 1:40 dilution (10 ug/ml) (red). Cells were fixed with 80% Methanol and permeabilized with 0.1% Tween-20. A Goat anti rabbit IgG (Alexa Fluor®488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

  • Immunocytochemistry/ Immunofluorescence analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling PABPN1 with purified ab75855 at 1:100 dilution (4.1μg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% tritonX-100. ab150077 Goat anti rabbit IgG(Alexa Fluor® 488) was used as the secondary antibody at 1:1000 dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

  • ab75855 (purified) at 1:30 dilution (5ug) immunoprecipitating PABPN1 in MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate.
    Lane 1 (input): MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate 10ug
    Lane 2 (+): ab75855 & MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab75855 in MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate
    For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used as the secondary antibody at 1:10,000 dilution.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue sections labeling PABPN1 with Purified ab75855 at 1:1000 dilution (0.41 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse kidney tissue sections labeling PABPN1 with Purified ab75855 at 1:1000 dilution (0.41 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human bladder carcinoma tissue sections labeling PABPN1 with Purified ab75855 at 1:1000 dilution (0.41 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.

  • Immunocytochemistry/Immunofluorescence analysis of MCF-7 cells labelling PABPN1 with unpurified ab75855 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.
    Control: PBS only.
    Nuclear counter stain: DAPI.

  • Anti-PABPN1 antibody [EP3000Y] (ab75855) at 1/2000 dilution (purified) + Rat brain lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 33 kDa



    Blocking and diluting buffer: 5% NFDM/TBST

  • All lanes : Anti-PABPN1 antibody [EP3000Y] (ab75855) at 1/2000 dilution (purified)

    Lane 1 : RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
    Lane 2 : Mouse spleen lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 33 kDa



    Blocking and diluting buffer: 5% NFDM/TBST

  • All lanes : Anti-PABPN1 antibody [EP3000Y] (ab75855) at 1/2000 dilution (purified)

    Lane 1 : 293T (Human embryonic kidney epithelial cell) whole cell lysates
    Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 33 kDa
    Observed band size: 49 kDa (why is the actual band size different from the predicted?)



    Blocking and diluting buffer: 5% NFDM/TBST

  • Lanes 1-2 : Anti-PABPN1 antibody [EP3000Y] (ab75855) at 1/200000 dilution (unpurified)
    Lane 3 : Anti-PABPN1 antibody [EP3000Y] (ab75855) at 1/1000000 dilution (unpurified)

    Lane 1 : Raw264.7 cell lysate
    Lane 2 : MCF-7 cell lysate
    Lane 3 : HeLa cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : HRP-conjugated goat anti-rabbit IgG at 1/1000 dilution

    Predicted band size: 33 kDa
    Observed band size: 49 kDa (why is the actual band size different from the predicted?)

  • Overlay histogram showing MCF-7 cells stained with unpurified ab75855 (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 (ab75855, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • Unpurified ab75855, at 1/100 dilution, staining PABPN1 in squamous cell cervical carcinoma, by Immunohistochemistry using formalin-fixed, paraffin-embedded tissue.

References

This product has been referenced in:

See all 13 Publications for this product

Customer reviews and Q&As

Abcam has not validated the combination of species/application used in this Abreview.
Application
CLIP
Sample
Human Cell lysate - whole cell (HEK 293T)
Specification
HEK 293T
Type
iCLIP
Irradiation Energy Level (mJ/cm2): 150
Wavelength (nm): 254
Positive control
Anti-hnRNP C antibody (sc-15386) bound protein G dynabeads, 0.1 mg/ml (antibody/beads)
Negative control
Protein G dynabeads without antibody bound
Immuno-precipitation step
Other - Protein G dynabeads
Username

Abcam user community

Verified customer

Submitted Mar 21 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (293T)
Loading amount
30000 cells
Specification
293T
Gel Running Conditions
Reduced Denaturing (4-20% Tris-Glycine)
Blocking step
5% Milk, 1% BSA as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Username

Abcam user community

Verified customer

Submitted Feb 26 2010

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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