Knockout Tested Rabbit Monoclonal E Cadherin antibody. Carrier free. Suitable for Flow Cyt, IHC-P, WB, ICC/IF, Flow Cyt (Intra), mIHC and reacts with Human samples. Cited in 7 publications.
pH: 7.2 - 7.4
Constituents: PBS
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Species Human | Dilution info - | Notes Perform heat-mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
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Cadherins are calcium-dependent cell adhesion proteins (PubMed:11976333). They preferentially interact with themselves in a homophilic manner in connecting cells; cadherins may thus contribute to the sorting of heterogeneous cell types. CDH1 is involved in mechanisms regulating cell-cell adhesions, mobility and proliferation of epithelial cells (PubMed:11976333). Promotes organization of radial actin fiber structure and cellular response to contractile forces, via its interaction with AMOTL2 which facilitates anchoring of radial actin fibers to CDH1 junction complexes at the cell membrane (By similarity). Has a potent invasive suppressor role. It is a ligand for integrin alpha-E/beta-7. E-Cad/CTF2 promotes non-amyloidogenic degradation of Abeta precursors. Has a strong inhibitory effect on APP C99 and C83 production. (Microbial infection) Serves as a receptor for Listeria monocytogenes; internalin A (InlA) binds to this protein and promotes uptake of the bacteria.
CD324, CDHE, UVO, CDH1, Cadherin-1, CAM 120/80, Epithelial cadherin, Uvomorulin, E-cadherin
Knockout Tested Rabbit Monoclonal E Cadherin antibody. Carrier free. Suitable for Flow Cyt, IHC-P, WB, ICC/IF, Flow Cyt (Intra), mIHC and reacts with Human samples. Cited in 7 publications.
pH: 7.2 - 7.4
Constituents: PBS
E-cadherin contains a number of cleavage sites which may yield a complex fragmentation pattern in WB. Multiple bands between ~80-120 kDa may be observed. This antibody has been tested on human samples in both WB and IHC. Customer feedback (see Abreview) suggests the antibody does not perform well in IHC on mouse tissue.
ab201499 is the carrier-free version of Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
This product is a recombinant monoclonal antibody, which offers several advantages including:
For more information, read more on recombinant antibodies.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
Our Low endotoxin, azide-free formats have low endotoxin level (1 EU/mg, determined by the TAL assay) and are free from azide, to achieve consistent experimental results in functional assays.
E-Cadherin sometimes called CDH1 or Cadherin-1 is a protein that plays a role in cell-to-cell adhesion. This transmembrane protein has a molecular weight of approximately 120 kDa. E-Cadherin is mainly expressed in epithelial tissues of various organs including the skin and gut. Its adhesive function is made possible by its extracellular domain that facilitates homophilic binding between cells contributing to the maintenance of tissue architecture and cellular integrity.
E-Cadherin participates in establishing and maintaining adherens junctions which are vital for tissue structure. E-Cadherin operates as a core component of the cadherin-catenin complex which links the protein to the actin cytoskeleton. Through this linkage E-Cadherin plays an important role in signaling pathways that influence cellular growth and differentiation. The protein's ability to mediate intercellular connections also regulates cellular motility and supports basic aspects of cell behavior in epithelial tissues.
E-Cadherin influences both the Wnt signaling pathway and the epithelial-to-mesenchymal transition (EMT). Within the Wnt signaling pathway E-Cadherin partners with β-catenin a significant player in transcription regulation and cell signaling. Disruption in E-Cadherin's adhesive functionality can lead to increased β-catenin availability affecting downstream transcriptional control. In the EMT process E-Cadherin loss characterizes an important step in which cells gain migratory and invasive properties typically seen during metastasis in cancer progression.
E-Cadherin's role is prominent in cancer particularly in the context of gastric and breast cancers. Mutations or altered expression of E-Cadherin lead to diminished cell adhesion promoting tumorigenesis and metastatic spread. In gastric cancer for instance mutated E-Cadherin often associates with loss of epithelial function facilitating cancer cell dissemination. Additionally its loss or dysfunction may correlate with proteins such as β-catenin further impacting cancer progression and pathology.
We have tested this species and application combination and it works. It is covered by our product promise.
We have not tested this specific species and application combination in-house, but expect it will work. It is covered by our product promise.
This species and application combination has not been tested, but we predict it will work based on strong homology. However, this combination is not covered by our product promise.
We do not recommend this combination. It is not covered by our product promise.
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ZEB1 or E-cadherin staining of metastases in ICI-mice. Note the higher E-cad and lower ZEB1 expression in the metastatic cells expressing OVOL1 or ZEB1-shRNA (sh4). Scale bar represents 100 μm.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 staining E Cadherin in HT-29 (Human colorectal adenocarcinoma) cells by ICC/IF (Immunocytochemistry/Immunofluorescence). Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% tritonX-100. Samples were incubated with primary antibody at 1/500 dilution. An Alexa Fluor® 488 Goat anti-Rabbit (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used as the secondary antibody at 1/1000 dilution. Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889) at 1/200 dilution was used as a counterstain. DAPI was used as a nuclear counterstain. This is a confocal image showing membranous staining on HT-29 cell line.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Immunofluorescence analysis of BeWo cells treated with 0.25% DMSO (controls), 10 nM PMA, or 10 nM 4αPMA for 72 h. The cells were then fixed and subsequently double-labeled for detection of DYSF (red) and E-cadherin (green). Nuclei were labeled with DAPI. While there can be a low level of spontaneous fusion in control cells (in our hands this ranges from about 4 to 9%), most cells are not fused and have at their borders intact E-cadherin labeling. Moreover, DYSF labeling was not detectable in non-fused BeWo cells. However, treatment of BeWo cells with 10 nM PMA for 72 h led to increased levels of cell fusion as indicated by the breakdown of E-cadherin labeling and the expression of DYSF in fused cells. When BeWo cells were treated with 10 nM 4αPMA for 72 h there was no detectable increase in cell fusion or DYSF expression. Arrows indicate areas enlarged and placed in insets. Bar = 50 µm.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Intracellular Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling E Cadherin with purified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at 1/30 dilution (10 μg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody was used at 1/2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue). This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Immunohistochemistry of breast carcinoma staining E Cadherin with Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at 1μg/ml
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma epithelial) cells labeling E Cadherin with Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% tritonX-100. Samples were then incubated with the primary antibody at a 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) secondary antibody at a 1/1000 dilution (green). The nuclear counter stain is DAPI (blue). Counterstained with Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at a 1/200 dilution (red).
Confocal image shows membranous staining on MCF7 cell line.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Overlay histogram showing A431 (Human epidermoid carcinoma cell line) cells stained with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 (red line). The cells were fixed with 80% methanol (5 minutes) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772, 1/1000 dilution) for 30 minute at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed ab96899) at 1/500 dilution for 30 minutes at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1 μg/1x106 cells) used under the same conditions. Unlabeled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Formalin-fixed, paraffin-embedded human breast carcinoma tissue stained for E Cadherin with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at a 1/500 dilution in immunohistochemical analysis.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Formalin/PFA-fixed paraffin-embedded human colonic adenocarcinoma tissue stained for E Cadherin with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at a 1/500 dilution in immunohistochemical analysis.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Formalin-fixed, paraffin-embedded human papillary carcinoma of thyroid gland tissue stained for E Cadherin with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at a 1/500 dilution in immunohistochemical analysis.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Formalin-fixed, paraffin-embedded human transitional cell carcinoma of kidney tissue stained for E Cadherin with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at a 1/500 dilution in immunohistochemical analysis.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Formalin-fixed, paraffin-embedded human lung adenocarcinoma tissue stained for E Cadherin with unpurified Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at a 1/500 dilution in immunohistochemical analysis.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody.
Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.
This IHC data was generated using the same anti-E Cadherin antibody clone, EP700Y, in a different buffer formulation (cat# Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Fluorescent immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772. Green-E-Cadherin red-PI
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Blocking and diluting buffer and concentration: 5% NFDM/TBST.
Anti-GAPDH antibody [EPR16891] - Loading Control ab181602 was as GAPDH loading control.
Exposure time: Lane1: 3 seconds; Lane 2-5: 40 seconds.
A375, HeLa and HT-1080 were reported as negative or express low level of E cadherin (PMID: 30393081, PMID: 16980628, PMID: 34715746), PMID: 25411788).
All lanes: Western blot - Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772) at 1/1000 dilution
Lane 1: MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg
Lane 2: HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg
Lane 3: A375 (Human malignant melanoma epithelial cell) whole cell lysate at 20 µg
Lane 4: HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg
Lane 5: HT-1080 (Human fibrosarcoma epithelial cell) whole cell lysate at 20 µg
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 97 kDa
This data was developed using the same antibody clone in a different buffer formulation (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
False colour image of Western blot: Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 was shown to bind specifically to E Cadherin. A band was observed at 105/130 kDa in wild-type Raji cell lysates with no signal observed at this size in CDH1 knockout cell line Human CDH1 (E Cadherin) knockout A-431 cell line ab273747 (knockout cell lysate ab273781). To generate this image, wild-type and CDH1 knockout Raji cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
Lanes 1 - 4: Western blot - Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772) at 1/10000 dilution
Lanes 1 - 4: Western blot - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (ab201499) at 1/10000 dilution
Lanes 1 - 4: Western blot - Anti-E Cadherin antibody [EP700Y] - BSA and Azide free (Anti-E Cadherin antibody [EP700Y] - BSA and Azide free ab256580) at 1/10000 dilution
Lane 1: Wild-type Raji cell lysate at 20 µg
Lane 2: Western blot - Human CDH1 (E Cadherin) knockout A-431 cell line (Human CDH1 (E Cadherin) knockout A-431 cell line ab273747)
Lane 2: Western blot - Human CDH1 (E Cadherin) knockout A-431 cell lysate (ab273781)
Lane 2: CDH1 knockout Raji cell lysate at 20 µg
Lane 3: MCF7 cell lysate at 20 µg
Lane 4: MDA-MB-231 cell lysate at 20 µg
All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Observed band size: 105 kDa, 130 kDa
Immunofluorescence staining of E-Cadherin using Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 in wild-type A431 cells (top panel) and CDH1 knockout A431 cells (bottom panel). 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 with Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at 1 µg/mL and Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 at 1 µg/mL overnight at +4°C, followed by a further incubation at room temperature for 1h with Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) (shown in green) and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) (shown in red), both at 1/1000. Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a confocal section is shown.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Immunofluorescence staining of E-Cadherin using Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 in wild-type A431 cells (top panel) and CDH1 knockout A431 cells (bottom panel). 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 with Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 at 0.2 µg/mL and Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 at 1 µg/mL overnight at +4°C, followed by a further incubation at room temperature for 1h with Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) (shown in green) and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) (shown in red), both at 1/1000. Nuclear DNA was labelled with DAPI (shown in blue).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a confocal section is shown.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Exposure time: 3.25 seconds.
Blocking and diluting buffer: 5% NFDM/TBST.
Full-length E Cadherin has a molecular weight of approximately 125 kDa. Other molecular weights between 80-100 kDa could also be observed depending on cell types or cell conditions.
PMID: 27274359, PMID: 26983597, PMID: 18478055, PMID: 22375065.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
All lanes: Western blot - Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772) at 1/1000 dilution
All lanes: MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 97 kDa
Observed band size: 80-125 kDa
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Western blot: Anti-CDH1 antibody [EP700Y] (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 was shown to bind specifically to CDH1. A band was observed at 130, 110, 80, 55, 40 kDa in wild-type A431 cell lysates with no signal observed at this size in CDH1 knockout cell line Human CDH1 (E Cadherin) knockout A-431 cell line ab273747 (knockout cell lysate ab273781). To generate this image, wild-type and CDH1 knockout A431 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
All lanes: Western blot - Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772) at 1/1000 dilution
Lane 1: Wild-type A431 cell lysate at 20 µg
Lane 2: Western blot - Human CDH1 (E Cadherin) knockout A-431 cell lysate (ab273781)
Lane 2: Western blot - Human CDH1 (E Cadherin) knockout A-431 cell line (Human CDH1 (E Cadherin) knockout A-431 cell line ab273747)
Lane 2: CDH1 knockout A431 cell lysate at 20 µg
Lane 3: Caco-2 cell lysate at 20 µg
All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 97 kDa
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772).
Flow cytometry overlay histogram showing MCF7 positive cells (left) and negative MDA-MB-231 cells (right) stained with Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 (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 Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker ab40772 (1x 106 in 100μl at 5 μg/ml (1/400)) for 30min on ice.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min on ice
Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control 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 525/40 bandpass filter.
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