Overview

  • Product name

    Anti-EpCAM antibody [EPR20532-225]
    See all EpCAM primary antibodies
  • Description

    Rabbit monoclonal [EPR20532-225] to EpCAM
  • 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 EpCAM aa 1-300. The exact sequence is proprietary.
    Database link: P16422

  • Positive control

    • WB: HCT 116 and HT-29 cell lysates; Human breast cancer and fetal kidney lysates. IHC-P: Human colon and thyroid cancer tissues. ICC/IF: HCT 116 and HT-29 cells. Flow Cyt: HCT 116 cells. IP: HCT 116 cell lysate.
  • 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 ab223582 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 42 kDa (predicted molecular weight: 35 kDa).
IHC-P 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF 1/500.
Flow Cyt 1/500.
IP 1/30.

Target

  • Function

    May act as a physical homophilic interaction molecule between intestinal epithelial cells (IECs) and intraepithelial lymphocytes (IELs) at the mucosal epithelium for providing immunological barrier as a first line of defense against mucosal infection. Plays a role in embryonic stem cells proliferation and differentiation. Up-regulates the expression of FABP5, MYC and cyclins A and E.
  • Tissue specificity

    Highly and selectively expressed by undifferentiated rather than differentiated embryonic stem cells (ESC). Levels rapidly diminish as soon as ESC's differentiate (at protein levels). Expressed in almost all epithelial cell membranes but not on mesodermal or neural cell membranes. Found on the surface of adenocarcinoma.
  • Involvement in disease

    Defects in EPCAM are the cause of diarrhea type 5 (DIAR5) [MIM:613217]. It is an intractable diarrhea of infancy characterized by villous atrophy and absence of inflammation, with intestinal epithelial cell dysplasia manifesting as focal epithelial tufts in the duodenum and jejunum.
    Defects in EPCAM are a cause of hereditary non-polyposis colorectal cancer type 8 (HNPCC8) [MIM:613244]. HNPCC is a disease associated with marked increase in cancer susceptibility. It is characterized by a familial predisposition to early-onset colorectal carcinoma (CRC) and extra-colonic tumors of the gastrointestinal, urological and female reproductive tracts. HNPCC is reported to be the most common form of inherited colorectal cancer in the Western world. Clinically, HNPCC is often divided into two subgroups. Type I is characterized by hereditary predisposition to colorectal cancer, a young age of onset, and carcinoma observed in the proximal colon. Type II is characterized by increased risk for cancers in certain tissues such as the uterus, ovary, breast, stomach, small intestine, skin, and larynx in addition to the colon. Diagnosis of classical HNPCC is based on the Amsterdam criteria: 3 or more relatives affected by colorectal cancer, one a first degree relative of the other two; 2 or more generation affected; 1 or more colorectal cancers presenting before 50 years of age; exclusion of hereditary polyposis syndromes. The term 'suspected HNPCC' or 'incomplete HNPCC' can be used to describe families who do not or only partially fulfill the Amsterdam criteria, but in whom a genetic basis for colon cancer is strongly suspected. Note=HNPCC8 results from heterozygous deletion of 3-prime exons of EPCAM and intergenic regions directly upstream of MSH2, resulting in transcriptional read-through and epigenetic silencing of MSH2 in tissues expressing EPCAM.
  • Sequence similarities

    Belongs to the EPCAM family.
    Contains 1 thyroglobulin type-1 domain.
  • Post-translational
    modifications

    Hyperglycosylated in carcinoma tissue as compared with autologous normal epithelia. Glycosylation at Asn-198 is crucial for protein stability.
  • Cellular localization

    Lateral cell membrane. Cell junction > tight junction. Co-localizes with CLDN7 at the lateral cell membrane and tight junction.
  • Information by UniProt
  • Database links

  • Alternative names

    • 17 1A antibody
    • 323/A3 antibody
    • Adenocarcinoma associated antigen antibody
    • Adenocarcinoma-associated antigen antibody
    • Antigen identified by monoclonal antibody AUA1 antibody
    • AUA1 antibody
    • CD326 antibody
    • CD326 antigen antibody
    • Cell surface glycoprotein Trop 1 antibody
    • Cell surface glycoprotein Trop 2 antibody
    • Cell surface glycoprotein Trop-1 antibody
    • CO 17A antibody
    • CO17 1A antibody
    • CO17A antibody
    • DIAR5 antibody
    • EGP 2 antibody
    • EGP antibody
    • EGP2 antibody
    • EGP314 antibody
    • EGP40 antibody
    • Ep CAM antibody
    • Ep-CAM antibody
    • EPCAM antibody
    • EPCAM_HUMAN antibody
    • EpCAM1 antibody
    • Epithelial cell adhesion molecule antibody
    • Epithelial Cell Adhesion Molecule Intracellular Domain (EpCAM-ICD) antibody
    • Epithelial cell surface antigen antibody
    • Epithelial cellular adhesion molecule antibody
    • Epithelial glycoprotein 1 antibody
    • Epithelial glycoprotein 314 antibody
    • Epithelial glycoprotein antibody
    • ESA antibody
    • GA733 1 antibody
    • GA733 2 antibody
    • GA733-2 antibody
    • gastrointestinal tumor-associated antigen 2, 35-KD glycoprotein antibody
    • gp4 antibody
    • hEGP 2 antibody
    • hEGP314 antibody
    • HNPCC8 antibody
    • Human epithelial glycoprotein 2 antibody
    • KS 1/4 antigen antibody
    • KS1/4 antibody
    • KSA antibody
    • Ly74 antibody
    • Lymphocyte antigen 74 antibody
    • M1S 1 antibody
    • M1S2 antibody
    • M4S1 antibody
    • Major gastrointestinal tumor associated protein GA733 2 antibody
    • Major gastrointestinal tumor-associated protein GA733-2 antibody
    • mEGP314 antibody
    • Membrane component chromosome 4 surface marker (35kD glycoprotein) antibody
    • Membrane component, chromosome 4, surface marker 1 antibody
    • Membrane component, chromosome 4, surface marker antibody
    • MIC18 antibody
    • MK 1 antibody
    • Protein 289A antibody
    • TACD1 antibody
    • TACSTD1 antibody
    • TROP1 antibody
    • Tumor associated calcium signal transducer 1 antibody
    • Tumor associated calcium signal transducer 2 precursor antibody
    • Tumor-associated calcium signal transducer 1 antibody
    see all

Images

  • Immunohistochemical analysis of paraffin-embedded human colon tissue labeling EpCAM with ab223582 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Membranous staining on human colon is observed (PMID: 15637741). Counter stained 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.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunofluorescent analysis of 4% paraformaledehyde-fixed, 0.1% Triton X-100 permeabilized HCT 116 (human colorectal carcinoma cell line) cells labeling EpCAM with ab223582 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining on HT-29 cells.

    The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.

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

  • All lanes : Anti-EpCAM antibody [EPR20532-225] (ab223582) at 1/5000 dilution

    Lane 1 : HCT 116 (human colorectal carcinoma cell line) lysate
    Lane 2 : HT-29 (human colorectal adenocarcinoma cell line) lysate

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size: 35 kDa
    Observed band size: 42 kDa
    why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-EpCAM antibody [EPR20532-225] (ab223582) at 1/1000 dilution

    Lane 1 : Human breat cancer lysate
    Lane 2 : Human fetal kidney lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Lane 1 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution
    Lane 2 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution

    Predicted band size: 35 kDa
    Observed band size: 42 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunohistochemical analysis of paraffin-embedded human thyroid cancer tissue labeling EpCAM with ab223582 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Membranous staining on human thyroid cancer is observed (PMID: 15637741). Counter stained 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.

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunofluorescent analysis of 4% paraformaledehyde-fixed, 0.1% Triton X-100 permeabilized HT-29 (human colorectal adenocarcinoma cell line) cells labeling EpCAM with ab223582 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining on HT-29 cells.

    The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.

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

  • Flow cytometric analysis of HCT 116 (human colorectal carcinoma cell line) cell line labeling EpCAM with ab223582 at 1/500 (red) compared with a rabbit monoclonal IgG Isotype control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

    Total viable cells were gated for the final data.

  • EpCAM was immunoprecipitated from 0.35 mg of HCT 116 (human colorectal carcinoma cell line) whole cell lysate with ab223582 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab223582 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: HCT 116 lysate 10 μg (Input).

    Lane 2: ab223582 IP in HCT 116 lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab223582 in HCT 116 whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 30 seconds.

References

This product has been referenced in:

  • Xu P  et al. A New in Vitro Model of GDLD by Knocking Out TACSTD2 and Its Paralogous Gene EpCAM in Human Corneal Epithelial Cells. Transl Vis Sci Technol 7:30 (2018). Read more (PubMed: 30619650) »
See 1 Publication for this product

Customer reviews and Q&As

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Breast)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: ab93684 Tris-EDTA Buffer pH 9.0
Permeabilization
Yes - 0.05% Tween 20
Specification
Breast
Blocking step
sea block as blocking agent for 30 minute(s) · Concentration: 100% · Temperature: 22°C
Fixative
Formaldehyde

Mr. David Ivancic

Verified customer

Submitted Jun 21 2019

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