Anti-E Cadherin 抗体 [EP700Y] - Low endotoxin, Azide free

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40772).

Flow cytometry overlay histogram showing MCF7 positive cells (left) and negative MDA-MB-231 cells (right) stained with 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 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.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Formalin/PFA-fixed paraffin-embedded human colonic adenocarcinoma tissue stained for E Cadherin with unpurified 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

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 (ab150077) was used as the secondary antibody at 1/1000 dilution. Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (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 (ab40772).

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

PMA induced cell fusion, DYSF expression, and activation of PKC in BeWo cells while 4αPMA was inactive

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 (ab40772).

Image from Omata W. et al PLoS One. 2013 Nov 13;8(11):e81003. doi: 10.1371/journal.pone.0081003. eCollection 2013.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Formalin-fixed, paraffin-embedded human lung adenocarcinoma tissue stained for E Cadherin with unpurified 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Overlay histogram showing A431 (Human epidermoid carcinoma cell line) cells stained with unpurified 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 (ab40772, 1/1000 dilution) for 30 minute at 22°C. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (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 (ab40772).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Formalin-fixed, paraffin-embedded human papillary carcinoma of thyroid gland tissue stained for E Cadherin with unpurified 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Formalin-fixed, paraffin-embedded human transitional cell carcinoma of kidney tissue stained for E Cadherin with unpurified 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Intracellular Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling E Cadherin with purified 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 (ab40772).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Formalin-fixed, paraffin-embedded human breast carcinoma tissue stained for E Cadherin with unpurified 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

This IHC data was generated using the same anti-E Cadherin antibody clone, EP700Y, in a different buffer formulation (cat# ab40772).

Fluorescent immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using ab40772. Green-E-Cadherin red-PI

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma epithelial) cells labeling E Cadherin with 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) (ab150077) secondary antibody at a 1/1000 dilution (green). The nuclear counter stain is DAPI (blue). Counterstained with 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 (ab40772).

• IHC-P

PubMed

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Mesenchymal cancer cells show increased metastasis while not requiring MET for solid tumor formation.

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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Image from Roca H. et al PLoS One. 2013 Oct 4;8(10):e76773. doi: 10.1371/journal.pone.0076773. eCollection 2013.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Immunofluorescence staining of E-Cadherin using 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 ab40772 at 0.2 µg/mL and 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 (ab150081) (shown in green) and 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 (ab40772).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Immunofluorescence staining of E-Cadherin using 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 ab40772 at 1 µg/mL and 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 (ab150081) (shown in green) and 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 (ab40772).

• WB

Lab

Western blot - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

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 (ab40772).

All lanes:

Western blot - Anti-E Cadherin antibody [EP700Y] - Intercellular Junction Marker (<a href='/products/primary-antibodies/e-cadherin-antibody-ep700y-intercellular-junction-marker-ab40772'>ab40772</a>) at 1/1000 dilution

All lanes:

MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 97 kDa

Observed band size: 80-125 kDa

false

• WB

Lab

Western blot - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40772). Western blot : Anti-CDH1 antibody [EP700Y] (ab40772) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, 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 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 (<a href='/products/primary-antibodies/e-cadherin-antibody-ep700y-intercellular-junction-marker-ab40772'>ab40772</a>) 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 (<a href='/products/cell-lines/human-cdh1-e-cadherin-knockout-a-431-cell-line-ab273747'>ab273747</a>)

Lane 2:

CDH1 knockout A431 cell lysate at 20 µg

Lane 3:

Caco-2 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 97 kDa

false

• WB

Lab

Western blot - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

This data was developed using the same antibody clone in a different buffer formulation (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] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, 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 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 (<a href='/products/primary-antibodies/e-cadherin-antibody-ep700y-intercellular-junction-marker-ab40772'>ab40772</a>) 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 (<a href='/products/primary-antibodies/e-cadherin-antibody-ep700y-bsa-and-azide-free-ab256580'>ab256580</a>) 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 (<a href='/products/cell-lines/human-cdh1-e-cadherin-knockout-a-431-cell-line-ab273747'>ab273747</a>)

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

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 105 kDa,130 kDa

false

• WB

Lab

Western blot - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40772). Blocking and diluting buffer and concentration : 5% NFDM/TBST. 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 (<a href='/products/primary-antibodies/e-cadherin-antibody-ep700y-intercellular-junction-marker-ab40772'>ab40772</a>) 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

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 97 kDa

false

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

Immunohistochemistry of breast carcinoma staining E Cadherin with 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 (ab40772).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-E Cadherin antibody [EP700Y] - Low endotoxin, Azide free (AB201499)

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.