Antibodies, Proteins, Kits and Reagents for Life Science

328 339 ₸ Product size

100 µl 328 339 ₸

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Anti-EpCAM antibody [EPR20532-222] (ab213500)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR20532-222] to EpCAM
Suitable for: IHC-P, WB, IP, ICC/IF
Knockout validated
Reacts with: Mouse, Rat, Human

Product name

Anti-EpCAM antibody [EPR20532-222]
See all EpCAM primary antibodies
Description

Rabbit monoclonal [EPR20532-222] to EpCAM
Host species

Rabbit

Tested Applications & Species

Application	Species
ICC/IF	Mouse Human
IHC-P	Mouse Rat Human
IP	Human
WB	Mouse Human

See all applications and species data

Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: A431, T-47D, SK-OV-3, HT-29, HCT 116 and 4T1 whole cell lysates; Human breast, colon and fetal kidney lysates; Mouse small intestine lysate. IHC-P: Human colon and endometrium cancer tissues; Mouse and rat colon tissues. ICC/IF: HCT 116 and 4T1 cells. IP: HCT 116 whole cell lysate.
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
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. 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...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR20532-222
Isotype

IgG
Research areas

Western blot - Anti-EpCAM antibody [EPR20532-222] (ab213500)

All lanes : Anti-EpCAM antibody [EPR20532-222] (ab213500) at 1/1000 dilution

Lane 1 : Wild-type A431 whole cell lysate
Lane 2 : EPCAM knockout A431 whole cell lysate
Lane 3 : HeLa whole cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 35 kDa
Observed band size: 35 kDa

Lanes 1 - 3: Merged signal (red and green). Green - ab213500 observed at 40 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.

ab213500 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 ab213500 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.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling EpCAM with ab213500 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Membranous staining on rat 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunofluorescent analysis of 100% methanol-fixed HCT 116 (Human colorectal carcinoma cell line) cells labeling EpCAM with ab213500 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing membranous staining on HCT 116 cells.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunohistochemical analysis of paraffin-embedded human colon tissue labeling EpCAM with ab213500 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-EpCAM antibody [EPR20532-222] (ab213500)

All lanes : Anti-EpCAM antibody [EPR20532-222] (ab213500) at 1/2000 dilution

Lane 1 : T-47D (Human ductal breast epithelial tumor cell line) whole cell lysate
Lane 2 : SK-OV-3 (Human ovarian cancer cell line) whole cell lysate
Lane 3 : HT-29 (Human colorectal adenocarcinoma cell line) whole cell lysate
Lane 4 : HCT 116 (Human colorectal carcinoma cell line) whole cell 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: 36,40 kDa
why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1: 15 seconds; Lane 2: 3 minutes; Lane 3: 10 seconds; Lane 4: 15 seconds.

The MW observed is consistent with the literature: PMID19136966; PMID 23618806.
Immunoprecipitation - Anti-EpCAM antibody [EPR20532-222] (ab213500)

EpCAM was immunoprecipitated from 0.35 mg of HCT 116 (Human colorectal carcinoma cell line) whole cell lysate with ab213500 at 1/40 dilution.

Western blot was performed from the immunoprecipitate using ab213500 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.

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

Lane 2: ab213500 IP in HCT 116 whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab213500 in HCT 116 whole cell lysate.

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

Exposure time: 30 seconds.
Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunofluorescent analysis of 100% methanol-fixed 4T1 (Mouse mammary gland carcinoma cell line) cells labeling EpCAM with ab213500 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing membranous staining on 4T1 cells.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunohistochemical analysis of paraffin-embedded human endometrium cancer tissue labeling EpCAM with ab213500 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Membranous staining on tumor cells of human endometrium 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-EpCAM antibody [EPR20532-222] (ab213500)

All lanes : Anti-EpCAM antibody [EPR20532-222] (ab213500) at 1/2000 dilution

Lane 1 : Human breast cancer lysate
Lane 2 : 4T1 (Mouse mammary gland carcinoma cell line) whole cell lysate
Lane 3 : Human colon lysate
Lane 4 : Mouse small intestine 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: 36,40 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1/2: 3 minutes; Lane 3: 10 seconds; Lane 4: 15 seconds.
Western blot - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Anti-EpCAM antibody [EPR20532-222] (ab213500) at 1/1000 dilution + Human fetal kidney lysate at 10 µg

Secondary
VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution

Predicted band size: 35 kDa
Observed band size: 36,40 kDa why is the actual band size different from the predicted?

Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20532-222] (ab213500)

Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling EpCAM with ab213500 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Membranous staining on mouse 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Anti-EpCAM antibody [EPR20532-222] (ab213500)

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