Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

This data was developed using the same antibody clone in a different buffer formulation (ab223582). ab223582 staining EpCAM in T47D positive cells (top panel) and HeLa negative cells (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab223582 at 0.1μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI. This antibody performed similarly using 100% methanol fixation. Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a single confocal section is shown.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

This data was developed using the same antibody clone in a different buffer formulation (ab223582). ab223582 staining EpCAM in wild-type A431 cells (top panel) and EpCAM knockout A431 cells (bottom panel). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab223582 at 1/5000 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

This data was developed using the same antibody clone in a different buffer formulation (ab223582). ab223582 staining EpCAM in wild-type A431 cells (top panel) and EpCAM knockout A431 cells (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% PBS-Tween for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab223582 at 0.1μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI. This antibody performed similarly using 100% methanol fixation. Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a single confocal section is shown.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Clone EPR20532-225 (ab225894) has been successfully conjugated by Abcam. This image was generated using Anti-EpCAM antibody [EPR20532-225] (Alexa Fluor® 647). Please refer to ab237396 for protocol details.

ab237396 staining EpCAM in HT-29 cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab237396 at 1/100 dilution (shown in red) and ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor^® 488), at 1/250 dilution (shown in ). Nuclear DNA was labeled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

This product also gave a positive signal under the same testing conditions in HT-29 cells fixed with 100% methanol (5 min).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223582).

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

• Flow Cyt

Lab

Flow Cytometry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

This data was developed using the same antibody clone in a different buffer formulation (ab225894)

Flow cytometry overlay histogram showing wild-type A431 (green line) and EPCAM knockout A431 cells stained with ab223582 (red line). The cells were incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab223582) (1x10⁶ in 100μl at 0.2 μg/ml) for 30 min at 4°C.

The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor^® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 4°C.

Isotype control antibody was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody (wild-type A431 - black line; EPCAM knockout A431 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223582).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Clone EPR20532-225 (ab225894) has been successfully conjugated by Abcam. This image was generated using Anti-EpCAM antibody [EPR20532-225] (Alexa Fluor® 488). Please refer to ab237395 for protocol details.

ab237395 staining EpCAM in HT-29 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab237395 at 1/1000 dilution (shown in green) and ab202272, Rabbit monoclonal to alpha Tubulin (Alexa Fluor^® 594), at 1/250 dilution (shown in red). Nuclear DNA was labeled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223582).

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223582).

• Flow Cyt

Unknown

Flow Cytometry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Clone EPR20532-225 (ab225894) has been successfully conjugated by Abcam. This image was generated using Anti-EpCAM antibody [EPR20532-225] (PE). Please refer to ab237397 for protocol details.

Overlay histogram showing HCT116 cells stained with ab237397 (red line). The cells were then incubated in 1x PBS / 10% normal Goat serum to block non-specific protein-protein interactions followed by the antibody (ab237397, 1/6250 dilution) for 30 min at 4°C. Isotype control antibody (black line) was Rabbit IgG (monoclonal) Phycoerythrin (ab209478) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 50 mW Yellow/Green laser (561nm) and 586/15 bandpass filter.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

This data is courtesy of ImmunoAtlas and it can be found here.

This image is courtesy of ImmunoAtlas.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data is courtesy of ImmunoAtlas and it can be found here.

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

This image is courtesy of ImmunoAtlas.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of the human endometrium (Formalin/PFA-fixed paraffin-embedded sections). Panel A : merged staining of anti-EpCAM (ab223582, magenta; Opal™690), anti-NKG2A (ab260035, green; Opal™520) and anti-CD31 (ab182981, red; Opal™570) on human endometrium. Panel B : anti-NKG2A stained on NK cells. Panel C : anti-CD31 stained on endothelial cells. Panel D : anti-EpCAM stained on glandular cells. Opal Polymer HRP Ms + Rb was used as a secondary antibody. The section was incubated in three rounds of staining : in the order of ab223582 at 1/500 dilution (1.008 μg/ml), ab260035 at 1/2000 dilution (0.262 μg/ml) and ab182981 at 1/4000 dilution (0.137 μg/ml) for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins. DAPI (blue) was used as a nuclear counter stain. This data was developed using the same antibody clone in a different buffer formulation (ab223582).

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

Fluorescence multiplex immunohistochemical analysis of Human breast cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of Anti-PD-L1 (ab251611; cyan; Opal™ 520), Anti-Granzyme B (ab219803; yellow; Opal™ 540), Anti-PD1 (ab251613; magenta; Opal™ 570), Anti-pan Cytokeratin (ab264485; red; Opal™ 620), Anti-EpCAM (ab225894; red; Opal™ 620), Anti-CD8 alpha (ab251596; green; Opal™ 650) and Anti-FOXP3 (ab96048; orange; Opal™ 690). EpCAM and pan-cytokeratin share the same dye and color. Dyes are pseudo-colored for better contrast of the markers.

The immunostaining was performed on a Leica Biosystems BOND^® MAX instrument with an Opal™ 6-Plex Detection Kit (NEL821001KT, Akoya Biosciences^®).

The section was incubated in six rounds of staining; sequentially for ab251611 (1/750 dilution), ab219803 (1/250 dilution), ab251613 (1/750 dilution), ab264485 (0.5 μg/ml), ab225894 (1/1250 dilution), ab251596 (1/1500 dilution) and ab96048 (10 μg/ml); each using a separate fluorescent tyramide signal amplification system. EDTA based antigen retrieval (Leica Biosystems BOND^® Epitope Retrieval Solution 2, pH 9.0, 20 minutes) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity. DAPI (dark blue) was used as a nuclear counter stain.
Microscopy and pseudocoloring of individual Opal™ dyes was performed using a Vectra 3 Imaging System (Akoya Biosciences^®).

This data is courtesy of ImmunoAtlas and it can be found here.

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

This image is courtesy of ImmunoAtlas.

• IP

Supplier Data

Immunoprecipitation - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

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.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab223582).

All lanes:

Immunoprecipitation - Anti-EpCAM antibody [EPR20532-225] (<a href='/en-us/products/primary-antibodies/epcam-antibody-epr20532-225-ab223582'>ab223582</a>)

Predicted band size: 35 kDa

Observed band size: 42 kDa

false

Exposure time: 30s

• WB

Lab

Western blot - Anti-EpCAM antibody [EPR20532-225] - BSA and Azide free (AB225894)

This data was developed using the same antibody clone in a different buffer formulation (ab223582).

Lanes 1 - 3 : Merged signal (red and green). Green - ab223582 observed at 40 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.

ab223582 was shown to react with EpCAM in A431 wild-type cells in Western blot. Loss of signal was observed when EpCAM knockout sample was used. A431 wild-type and EpCAM knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% Milk in TBS-T (0.1% Tween^®) before incubation with ab223582 and ab7291 (Mouse anti-Alpha Tubulin [DM1A] overnight at 4°C at a 1 in 1000 dilution and a 1 in 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 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-EpCAM antibody [EPR20532-225] (<a href='/en-us/products/primary-antibodies/epcam-antibody-epr20532-225-ab223582'>ab223582</a>) at 1/1000 dilution

Lane 1:

Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

EPCAM knockout A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human EPCAM knockout A-431 cell line (<a href='/en-us/products/cell-lines/human-epcam-knockout-a-431-cell-line-ab261902'>ab261902</a>)

Lane 3:

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

Predicted band size: 35 kDa

Observed band size: 40 kDa

false