Anti-EpCAM antibody [RM1016]

• ICC

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Immunocytochemistry - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• Flow Cyt

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Flow Cytometry - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IP

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Immunoprecipitation - Anti-EpCAM antibody [RM1016] (AB282457)

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).

All lanes:

Immunoprecipitation - Anti-EpCAM antibody [RM1016] (ab282457)

Predicted band size: 35 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• ICC

Supplier Data

Immunocytochemistry - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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).

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-EpCAM antibody [RM1016] (AB282457)

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.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-EpCAM antibody [RM1016] (AB282457)

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).

• IP

Supplier Data

Immunoprecipitation - Anti-EpCAM antibody [RM1016] (AB282457)

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).

All lanes:

Immunoprecipitation - Anti-EpCAM antibody [RM1016] (ab282457)

Predicted band size: 35 kDa

false

• WB

Lab

Western blot - Anti-EpCAM antibody [RM1016] (AB282457)

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.

All lanes:

Western blot - Anti-EpCAM antibody [RM1016] (ab282457) at 1/1000 dilution

Lane 1:

HCT 116 (Human colorectal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

4T1 (Mouse mammary gland carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 5:

Human colon lysate at 20 µg

Lane 6:

Mouse colon lysate at 20 µg

Lane 7:

Rat colon lysate at 20 µg

Lane 8:

Mouse kidney lysate at 20 µg

Lane 9:

Rat kidney lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 35 kDa

Observed band size: 12 kDa,36 kDa,40 kDa

false