Recombinant Anti-EpCAM antibody [RM1016] (ab282457)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit recombinant multiclonal [RM1016] to EpCAM
- Suitable for: Flow Cyt, IP, WB, IHC-P, IHC-Fr, ICC
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-EpCAM antibody [RM1016]
See all EpCAM primary antibodies -
Description
Rabbit recombinant multiclonal [RM1016] to EpCAM -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt, IP, WB, IHC-P, IHC-Fr, ICCmore details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
This product was produced with the following immunogens:
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers. -
Positive control
- WB: HCT 116, 4T1 and NIH/3T3 whole cell lysate. Human colon lysate. Mouse and rat colon and kidney lysate. IHC-P: Human colon and colon carcinoma tissue. Mouse and rat colon tissue. Mouse colon carcinoma tissue. Human liver tissue. IHC-Fr: Mouse and rat colon tissue. ICC: HCT 116 and 4T1 cells. Flow Cyt: HCT-116 and 4T1 cells. IP: Human colon lysate. 4T1 whole cell lysate.
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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
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
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Recombinant Multiclonal -
Clone number
RM1016 -
Isotype
IgG -
Research areas
Associated products
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Compatible Secondaries
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab282457 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 |
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Flow Cyt |
1/50.
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IP |
1/30.
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WB |
1/1000. Predicted molecular weight: 35 kDa.
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IHC-P |
1/4000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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IHC-Fr |
1/100.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20) |
|
ICC |
1/250.
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Notes |
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Flow Cyt
1/50. |
IP
1/30. |
WB
1/1000. Predicted molecular weight: 35 kDa. |
IHC-P
1/4000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
IHC-Fr
1/100. Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20) |
ICC
1/250. |
Target
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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
modificationsHyperglycosylated 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
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Database links
- Entrez Gene: 4072 Human
- Entrez Gene: 17075 Mouse
- Entrez Gene: 171577 Rat
- Omim: 185535 Human
- SwissProt: P16422 Human
- SwissProt: Q99JW5 Mouse
- SwissProt: O55159 Rat
- Unigene: 542050 Human
see all -
Alternative names
- 17 1A antibody
- 323/A3 antibody
- Adenocarcinoma associated antigen antibody
see all
Images
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All lanes : Anti-EpCAM antibody [RM1016] (ab282457) at 1/1000 dilution
Lane 1 : HCT 116 (Human colorectal carcinoma epithelial cell) whole cell lysate
Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 3 : 4T1 (Mouse mammary gland carcinoma epithelial cell) whole cell lysate
Lane 4 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate
Lane 5 : Human colon lysate
Lane 6 : Mouse colon lysate
Lane 7 : Rat colon lysate
Lane 8 : Mouse kidney lysate
Lane 9 : Rat kidney 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: 12, 36, 40 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST.
Negative control: HeLa (PMID: 31806375), NIH/3T3 (PMID: 23264216).
The MW observed is consistent with the literature: PMID 19136966; PMID 23618806. The band between 12-15kDa has been documented in literature as the cleaved form of EpCAM (PMID: 23409978).
Exposure time: 8 seconds.
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Immunohistochemical analysis of paraffin-embedded human colon tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on human colon. The section was incubated with ab282457 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on human colon carcinoma. The section was incubated with ab282457 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on mouse colon. The section was incubated with ab282457 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on rat colon. The section was incubated with ab282457 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded mouse colon carcinoma tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on mouse colon carcinoma. The section was incubated with ab282457 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded human liver tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Membranous staining on the bile duct in human liver, and no staining on the hepatocytes. The section was incubated with ab282457 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded human skeletal muscle tissue labeling EpCAM with ab282457 at 1/4000 (0.124 µg/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Negative control: no staining on human skeletal muscle. The section was incubated with ab282457 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HCT 116 and HeLa cells labelling EpCAM with ab282457 at 1/250 (1.976 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing membranous staining in HCT 116 cell line.
Negative control: HeLa (PMID: 31806375).
ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized 4T1 and NIH/3T3 cells labelling EpCAM with ab282457 at 1/250 (1.976 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing membranous staining in 4T1 cell line.
Negative control: NIH/3T3 (PMID:31806375).
ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.
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Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat colon tissue labeling EpCAM with ab282457 at 1/100 (4.94 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Membranous staining on rat colon is observed. The nuclear counterstain was DAPI (Blue).
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
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Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse colon tissue labeling EpCAM with ab282457 at 1/100 (4.94 µg/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution (Green). Membranous staining on mouse colon is observed. The nuclear counterstain was DAPI (Blue).
Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).
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Flow cytometric analysis of HeLa (Human cervix adenocarcinoma epithelial cell, Left) / HCT-116 (Human colorectal carcinoma epithelial cell, Right) cells labelling EpCAM with ab282457 at 1/500 dilution (0.1 µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled 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.
Negative control: HeLa. (PMID: 31806375).
Gated on viable cells.
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Flow cytometric analysis of NIH/3T3 (Mouse embryonic fibroblast, Left) / 4T1 (Mouse mammary gland carcinoma epithelial cell, Right) cells labelling EpCAM with ab282457 at 1/50 dilution (1 µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled 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.
Negative control: NIH/3T3 (PMID:23264216).
Gated on viable cells.
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EpCAM was immunoprecipitated from 0.35 mg human colon lysate 10 µg with ab282457 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab282457 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: Human colon lysate 10 µg
Lane 2: ab282457 IP in human colon lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab282457 in human colon lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 8 seconds.
The MW observed is consistent with the literature: PMID 19136966; PMID 23618806. The band between 12-15kDa has been documented in literature as the cleaved form of EpCAM (PMID: 23409978).
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EpCAM was immunoprecipitated from 0.35 mg 4T1 (Mouse mammary gland carcinoma epithelial cell) whole cell lysate 10 µg with ab282457 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab282457 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: 4T1 whole cell lysate 10 µg
Lane 2: ab282457 IP in 4T1 whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab282457 in 4T1 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 8 seconds.
The MW observed is consistent with the literature: PMID 19136966; PMID 23618806. The band between 12-15kDa has been documented in literature as the cleaved form of EpCAM (PMID: 23409978).
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.
Datasheets and documents
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SDS download
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Datasheet download
References (1)
ab282457 has been referenced in 1 publication.
- Yu Z et al. Single-cell sequencing reveals the heterogeneity and intratumoral crosstalk in human endometrial cancer. Cell Prolif 55:e13249 (2022). PubMed: 35560676