Antibodies, Proteins, Kits and Reagents for Life Science

284 784 ₸ Product size

100 µl 284 784 ₸

Order now and get it on Tuesday March 02, 2021

Anti-EpCAM antibody [EPR20533-63] (ab221552)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR20533-63] to EpCAM
Suitable for: IP, Flow Cyt, ICC/IF, WB, IHC-P
Reacts with: Mouse

Product name

Anti-EpCAM antibody [EPR20533-63]
See all EpCAM primary antibodies
Description

Rabbit monoclonal [EPR20533-63] to EpCAM
Host species

Rabbit

Tested Applications & Species

Application	Species
Flow Cyt	Mouse
ICC/IF	Mouse
IHC-P	Mouse
IP	Mouse
WB	Mouse

See all applications and species data

Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: 4T1 whole cell lysate; Mouse colon, kidney and small intestine lysates. IHC-P: Mouse colon and lung tissues. ICC/IF: 4T1 cells. Flow Cyt: 4T1 cells. IP: Mouse kidney lysate; 4T1 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.
Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

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: 40% Glycerol (glycerin, glycerine), 0.05% BSA, PBS
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR20533-63
Isotype

IgG
Research areas

Flow Cytometry - Anti-EpCAM antibody [EPR20533-63] (ab221552)

Flow cytometry overlay histogram showing left M158 positive cells and right negative NIH3T3 cells stained with ab221552 (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 (ab221552) (1x10⁶ in 100 µL at 0.1 µg/ml) for 30 min on ice.
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The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor ^® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min on ice.
Isotype control antibody (black line) was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.
Western blot - Anti-EpCAM antibody [EPR20533-63] (ab221552)

All lanes : Anti-EpCAM antibody [EPR20533-63] (ab221552) at 1/5000 dilution

Lane 1 : Mouse colon lysate
Lane 2 : Mouse kidney lysate
Lane 3 : 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

Developed using the ECL technique.

Predicted band size: 35 kDa
Observed band size: 39 kDa
why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1/2: 3 seconds; Lane 3: 10 seconds.

The MW observed is consistent with the literature (PMID 23409978; PMID 23618806).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20533-63] (ab221552)

Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling EpCAM with ab221552 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. 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) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-EpCAM antibody [EPR20533-63] (ab221552)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized 4T1 (mouse mammary gland carcinoma cell line) and NIH/3T3 (mouse embyro fibroblast cell line) cells labeling EpCAM with ab221552 at 1/100 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 4T1 cells.

Negative control: NIH/3T3 (PMID:23264216).

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.
Immunoprecipitation - Anti-EpCAM antibody [EPR20533-63] (ab221552)

EpCAM was immunoprecipitated from 0.35 mg of mouse kidney lysate with ab221552 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab221552 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1: Mouse kidney lysate 10 µg (Input).

Lane 2: ab221552 IP in mouse kidney lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab221552 in mouse kidney lysate.

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

Exposure time: 3 seconds.
Flow Cytometry - Anti-EpCAM antibody [EPR20533-63] (ab221552)

Flow cytometric analysis of 4% paraformaldehyde-fixed 4T1 (mouse mammary gland carcinoma cell line) and NIH/3T3 (mouse embyro fibroblast cell line) cells labeling EpCAM with ab221552 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

Flow cytometry was performed with fresh cells without fixation and permeabilization.

Negative control: NIH/3T3 (PMID:23264216).
Western blot - Anti-EpCAM antibody [EPR20533-63] (ab221552)

All lanes : Anti-EpCAM antibody [EPR20533-63] (ab221552) at 1/5000 dilution

Lane 1 : 4T1 (mouse mammary gland carcinoma cell line) whole cell lysate
Lane 2 : NIH/3T3 (mouse embyro fibroblast 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

Developed using the ECL technique.

Predicted band size: 35 kDa
Observed band size: 39 kDa why is the actual band size different from the predicted?

Exposure time: 1 second

Blocking/Dilution buffer: 5% NFDM/TBST.

The MW observed is consistent with the literature (PMID 23409978; PMID 23618806).

Negative control: NIH/3T3 (PMID:23264216).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EpCAM antibody [EPR20533-63] (ab221552)

Immunohistochemical analysis of paraffin-embedded mouse lung tissue labeling EpCAM with ab221552 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Membranous staining on mouse lung 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.
Western blot - Anti-EpCAM antibody [EPR20533-63] (ab221552)

All lanes : Anti-EpCAM antibody [EPR20533-63] (ab221552) at 1/5000 dilution

Lane 1 : 4T1 (mouse mammary gland carcinoma cell line) whole cell lysate
Lane 2 : NIH/3T3 (mouse embyro fibroblast 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

Developed using the ECL technique.

Predicted band size: 35 kDa
Observed band size: 12,39 kDa why is the actual band size different from the predicted?

Exposure time: 10 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

The MW observed is consistent with the literature (PMID 23618806).

The band between 12-15kDa has been documented in literature as the cleaved form of EpCAM (PMID: 23409978).

Negative control: NIH/3T3 (PMID:23264216).
Immunoprecipitation - Anti-EpCAM antibody [EPR20533-63] (ab221552)

EpCAM was immunoprecipitated from 0.35 mg of 4T1 (mouse mammary gland carcinoma cell line) whole cell lysate with ab221552 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab221552 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1: 4T1 (Mouse mammary gland carcinoma) whole cell lysate 10 µg (Input).

Lane 2: ab221552 IP in 4T1 whole cell lysate .

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab221552 in 4T1 whole cell lysate .

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

Exposure time: 30 seconds.

The band between 12-15kDa has been documented in literature as the cleaved form of EpCAM (PMID: 23409978).
Anti-EpCAM antibody [EPR20533-63] (ab221552)

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