Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-Acid phosphatase antibody [EPR21787] (ab235448)

Overview

  • Product name

    Anti-Acid phosphatase antibody [EPR21787]
    See all Acid phosphatase primary antibodies
  • Description

    Rabbit monoclonal [EPR21787] to Acid phosphatase
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human Acid phosphatase aa 100 to the C-terminus. The exact sequence is proprietary.
    Database link: P24666

  • Positive control

    • WB: HepG2, SH-SY5Y, HCT 116, Jurkat, HeLa, C6 and NIH/3T3 whole cell lysates; human placenta and colon lysates. IP: HeLa whole cell lysate. IHC-P: Human colon cancer tissue; mouse colon tissue; rat colon tissue. ICC/IF: HeLa and HepG2 cells. FC: HeLa cells.
  • 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.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab235448 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. Detects a band of approximately 18 kDa (predicted molecular weight: 18 kDa).
IHC-P 1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF 1/100.
Flow Cyt 1/50.
IP 1/30.

Target

  • Relevance

    Acid phosphatases (AP) dephosphorylate phosphate groups from phosphate esters under acid conditions. Different acid phosphatase isozymes are found in different organs, and their serum levels are used as a diagnostic for disease in the corresponding organs. Elevated prostatic acid phosphatase levels may indicate the presence of prostate cancer and elevated tartrate-resistant acid phosphatase levels may indicate bone disease.
  • Cellular localization

    ACP1: Cytoplasm. ACP2: Lysosome membrane; Single-pass membrane protein. ACP5: Lysosome. ACPP: Isoform 1: Secreted. Isoform 2: Lysosome membrane; Single-pass type I membrane protein.
  • Database links

  • Alternative names

    • Acid phosphatase 1 soluble antibody
    • Acid phosphatase 2 lysosomal antibody
    • Acid phosphatase 5 tartrate resistant antibody
    • Acid phosphatase of erythrocyte antibody
    • Acid phosphatase prostate antibody
    • ACP1 antibody
    • ACP2 antibody
    • ACP3 antibody
    • ACP5 antibody
    • ACPP antibody
    • Adipocyte acid phosphatase antibody
    • HAAP antibody
    • LAP antibody
    • LMW PTP antibody
    • LMW PTPase antibody
    • Low molecular weight cytosolic acid phosphatase antibody
    • Low molecular weight phosphotyrosine protein phosphatase antibody
    • Lysosomal acid phosphatase antibody
    • PAP antibody
    • Prostatic acid phosphotase antibody
    • Red cell acid phosphatase 1 antibody
    • Tartrate resistant acid ATPase antibody
    • Tartrate resistant acid phosphatase 5 antibody
    • Tartrate resistant acid phosphatase type 5 antibody
    • TR AP antibody
    • TRAP antibody
    • TrATPase antibody
    see all

Images

  • All lanes : Anti-Acid phosphatase antibody [EPR21787] (ab235448) at 1/1000 dilution

    Lane 1 : Wild-type HEK293 whole cell lysate
    Lane 2 : ACP1 knockout HEK293 whole cell lysate
    Lane 3 : K562 whole cell lysate
    Lane 4 : HeLa whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 18 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab235448 observed at 18 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab235448 was shown to specifically react with ACP1 (Acid phosphatase 1) in wild-type HEK 293 cells as signal was lost in ACP1 knockout cells. Wild-type and ACP1 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% Milk. Ab235448 and ab8245 (Mouse 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-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.

  • Immunohistochemical analysis of paraffin-embedded human colon cancer tissue labeling Acid phosphatase with ab235448 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Positive staining in human colon cancer (PMID: 25811796) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

    Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Acid phosphatase with ab235448 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic and nuclear staining in HeLa cell line (PMID 26159288). The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

  • All lanes : Anti-Acid phosphatase antibody [EPR21787] (ab235448) at 1/1000 dilution

    Lane 1 : HepG2 (human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg
    Lane 2 : SH-SY5Y (human neuroblastoma cell line from bone marrow) whole cell lysate at 20 µg
    Lane 3 : HCT 116 (human colorectal carcinoma cell line) whole cell lysate at 20 µg
    Lane 4 : Jurkat (human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg
    Lane 5 : Human placenta lysate at 20 µg
    Lane 6 : Human colon lysate at 20 µg
    Lane 7 : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg
    Lane 8 : C6 (rat glioma cell line) whole cell lysate at 10 µg
    Lane 9 : NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate at 10 µg

    Secondary
    Lanes 1-6 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution
    Lanes 7-9 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 18 kDa
    Observed band size: 18 kDa



    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure times.

    Lane 1: 37 seconds, Lanes 2-9: 3 minutes.

    The molecular mass observed is consistent with what has been described in the literature (PMID:25811796).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cell line labeling Acid phosphatase with ab235448 at 1/50 dilution (red) compared with a Isotype control details (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

  • Acid phosphatase was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab235448 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab235448 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

    Lane 1: HeLa whole cell lysate lysate 10 μg (Input).
    Lane 2: ab235448 IP in HeLa whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab235448 in HeLa whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.
    Exposure time: 3 minutes.

  • Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling Acid phosphatase with ab235448 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Positive staining in rat colon (PMID: 25811796) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

    Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

     

  • Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling Acid phosphatase with ab235448 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Positive staining in mouse colon (PMID: 25811796) is observed. Counterstained with hematoxylin.
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

    Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling Acid phosphatase with ab235448 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic and nuclear staining in HepG2 cell line (PMID 26159288). The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

References

ab235448 has not yet been referenced specifically in any publications.

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