Recombinant
RabMAb

Recombinant Anti-PP2A alpha + beta antibody [E155] (ab32104)

Submit a review Q&A (3)References (36)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [E155] to PP2A alpha + beta
  • Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, Dot blot
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-PP2A alpha + beta antibody [E155]
    See all PP2A alpha + beta primary antibodies

  • Description

    Rabbit monoclonal [E155] to PP2A alpha + beta

  • Host species

    Rabbit

  • Specificity

    The immunogen sequence for ab32104 has 100% homology with the PP2A-beta protein.

  • Tested applications

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

  • Species reactivity

    Reacts with: Mouse, Rat, Human
    Predicted to work with: Zebrafish

  • Immunogen

    Synthetic peptide within Human PP2A alpha + beta aa 250-350. The exact sequence is proprietary.

  • Positive control

    • IHC-P: Human pancreas tissue. WB: lysate of A431 cells (serum starved overnight) treated with EGF (100ng/ml) for 5-10 minutes at 37C (BD Biosciences).

  • 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.

Properties

Applications

Our Abpromise guarantee covers the use of ab32104 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 1/5000. Predicted molecular weight: 35 kDa.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF Use a concentration of 5 µg/ml.
Flow Cyt Use at an assay dependent concentration.

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

 
Dot blot Use at an assay dependent concentration.

Target

  • Function

    PP2A can modulate the activity of phosphorylase B kinase casein kinase 2, mitogen-stimulated S6 kinase, and MAP-2 kinase. Cooperates with SGOL2 to protect centromeric cohesin from separase-mediated cleavage in oocytes specifically during meiosis I (By similarity). Can dephosphorylate SV40 large T antigen and p53/TP53. Dephosphorylates SV40 large T antigen, preferentially on serine residues 120, 123, 677, and perhaps 679. The C subunit was most active, followed by the AC form, which was more active than the ABC form, and activity of all three forms was strongly stimulated by manganese, and to a lesser extent by magnesium. Dephosphorylation by the AC form, but not C or ABC form is inhibited by small T antigen. Activates RAF1 by dephosphorylating it at 'Ser-259'.

  • Sequence similarities

    Belongs to the PPP phosphatase family. PP-1 subfamily.

  • Post-translational
    modifications

    Reversibly methyl esterified on Leu-309. Carboxyl methylation may play a role in holoenzyme assembly, enhancing the affinity of the PP2A core enzyme for some, but not all, regulatory subunits. It varies during the cell cycle.
    Phosphorylation of either threonine (by autophosphorylation-activated protein kinase) or tyrosine results in inactivation of the phosphatase. Auto-dephosphorylation has been suggested as a mechanism for reactivation.

  • Cellular localization

    Cytoplasm. Nucleus. Chromosome > centromere. Cytoplasm > cytoskeleton > spindle pole. In prometaphase cells, but not in anaphase cells, localizes at centromeres. During mitosis, also found at spindle poles. Centromeric localization requires the presence of SGOL2.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • PP2A A antibody
    • PP2A alpha antibody
    • PP2A B antibody
    • PP2A beta antibody
    • PP2A-alpha antibody
    • PP2A-beta antibody
    • PP2AA_HUMAN antibody
    • PP2Aalpha antibody
    • PP2AB_HUMAN antibody
    • PP2Abeta antibody
    • PP2Ac antibody
    • PP2CA antibody
    • PP2Calpha antibody
    • PP2CB antibody
    • PPP2CA antibody
    • PPP2CB antibody
    • Protein phosphatase 2 catalytic subunit alpha isoform antibody
    • Protein phosphatase 2 catalytic subunit beta isoform antibody
    • Protein phosphatase type 2A catalytic subunit antibody
    • Replication protein C antibody
    • RP C antibody
    • RP-C antibody
    • RPC antibody
    • Serine/threonine protein phosphatase 2A catalytic subunit alpha isoform antibody
    • Serine/threonine protein phosphatase 2A catalytic subunit beta isoform antibody
    • Serine/threonine-protein phosphatase 2A catalytic subunit alpha isoform antibody
    see all

Images

  • Western blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Western blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    All lanes : Anti-PP2A alpha + beta antibody [E155] (ab32104) at 1/5000 dilution

    Lane 1 : Untreated HeLa (human cervix adenocarcinoma) whole cell lysates
    Lane 2 : HeLa (human cervix adenocarcinoma) whole cell lysates,cells were treated with Pervanadate for 30 minutes at the final concentration of 1mg/ml
    Lane 3 : HeLa (human cervix adenocarcinoma) whole cell lysates,cells were treated with Pervanadate for 30 minutes at the final concentration of 1mg/ml Then the membrane was incubated with Alkaline phosphatase.

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG, (H+L), HRP conjugated)

    Predicted band size: 35 kDa



    Blocking and Diluting buffer and concentration:5% NFDM/TBST

    Exposure time:3 minutes

  • Immunocytochemistry/ Immunofluorescence - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Immunocytochemistry/ Immunofluorescence - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    ICC/IF image of ab32104 stained HeLa cells. The cells were 4% formaldehyde 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 (ab32104, 5µg/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.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PP2A alpha + beta antibody [E155] (ab32104)

    ab32104 (2µg/ml) staining PP2A alpha  in human pancreas using an automated system (DAKO Autostainer Plus). Using this protocol there is strong cytoplasmic staining of the islet of Langerhans.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

  • Flow Cytometry - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Flow Cytometry - Anti-PP2A alpha + beta antibody [E155] (ab32104)

    Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling PP2A alpha + beta with purified ab32104 at 1/20 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

  • Dot Blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Dot Blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)

    Dot blot was performed using ab32104 at 1/1000 dilution.Goat Anti-Rabbit IgG,(H+L),HRP conjugated (ab97051) was used as secondary antibody at 1/100000 dilution

    Lane 1:PP2A alpha + beta  phospho peptide

    Lane 2:PP2A alpha + beta non-phospho peptide

    Blocking and Diluting buffer and concentration:5% NFDM/TBST

    Exposure time:10 seconds

  • Western blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    Western blot - Anti-PP2A alpha + beta antibody [E155] (ab32104)
    All lanes : Anti-PP2A alpha + beta antibody [E155] (ab32104) at 1/5000 dilution

    Lane 1 : untreated A431 cell lysate
    Lane 2 : EGF treated A431 cell lysate (the specific EGF concentration and incubation time is confidential information)

    Predicted band size: 35 kDa
    Observed band size: 35 kDa

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

References

Publishing research using ab32104? Please let us know so that we can cite the reference in this datasheet.

ab32104 has been referenced in 36 publications.

  • Yuan C  et al. OAB-14, a bexarotene derivative, improves Alzheimer's disease-related pathologies and cognitive impairments by increasing ß-amyloid clearance in APP/PS1 mice. Biochim Biophys Acta Mol Basis Dis 1865:161-180 (2019). PubMed: 30389579
  • da Silva RA  et al. Laminar shear stress-provoked cytoskeletal changes are mediated by epigenetic reprogramming of TIMP1 in human primary smooth muscle cells. J Cell Physiol 234:6382-6396 (2019). PubMed: 30238981
  • Williams AP  et al. Investigation of PP2A and Its Endogenous Inhibitors in Neuroblastoma Cell Survival and Tumor Growth. Transl Oncol 12:84-95 (2019). PubMed: 30286326
  • Pinto TS  et al. c-Src kinase contributes on endothelial cells mechanotransduction in a heat shock protein 70-dependent turnover manner. J Cell Physiol 234:11287-11303 (2019). PubMed: 30565700
  • Tang S  et al. Hydrogen peroxide redistributes the localization of protein phosphatase methylesterase 1. Life Sci 213:166-173 (2018). PubMed: 30340029
View all Publications for this product

Customer reviews and Q&As

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1-3 of 3 Abreviews or Q&A

Question

Estimada ,
Esta es la secuencia C-terminal alrededor de la Tyr307 en mi proteína:XXXXXXXXXXXXXXXXXXX
Muchas gracias por su ayuda,

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Answer

Te pido disculpas por la espera.

Respecto al anticuerpo ab32104, hemos realizado un BLAST comparando ambas secuencias, y existe un 77% de homología entre ellas. No se prevé por tanto que el anticuerpo pueda reconocerla.


¿De qué especie se trata? ¿En qué aplicación lo quieres usar?

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Question

BATCH NUMBER -- NOT SPECIFIED -- ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM Multiple bands SAMPLE cone photoreceptor derived 661W cells PRIMARY ANTIBODY primary antibody added at 1 in 2500 dilution overnight at 4C DETECTION METHOD ECL 10 min exposure SAMPLE PREPARATION extracted in RIPA buffer on ice via standard protocol AMOUNT OF PROTEIN LOADED 20 ug ELECTROPHORESIS/GEL CONDITIONS reducing 12% gel TRANSFER AND BLOCKING CONDITIONS transfer 90 V for 50 mins blocked in 10% milk for 1 hour room temp SECONDARY ANTIBODY anti-rabbit 1 in 1000 dilution HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 1 HAVE YOU RUN A "NO PRIMARY" CONTROL? No ADDITIONAL NOTES the data sheet says that this antibody picks up only one band, as you can see on the attached file there are two definite bands, can you tell me what each band might be? Which band is the specific band?

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Answer

Thank you for your enquiry. I am sorry to hear you are having a problem with ab32104. To our knowledge, the antibody does not react with mouse. Therefore, we are unable to say, which of the two bands might be correct or if any of them is specific. I would recommend to use a positive control, as described on our datasheet and run it next to your sample. Doing this, you will be able to see, which band corresponds to the control band. Please let me know if this helps and do not hesitate to contact us for further advice.

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Question

Thank you for your quick answer. Concearning the WB image you show on the data sheet: Could you find out how long the cells were treated with EGF and at what concentration? Thank you for your help. Best regards,

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Answer

I have the regret to inform you that the scientist who tested the antibody informed me that the stimulation paradigm was confidential. I'm sorry I was unable to provide this information.

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