Overview

  • Product name
  • Description
    Rabbit polyclonal to E Cadherin
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, ICC/IF, IHC-P, IHC-Frmore details
  • Species reactivity
    Reacts with: Human, Pig
  • Immunogen

    Recombinant fragment within Human E Cadherin aa 600-750. The exact sequence is proprietary.
    Database link: P12830

  • Positive control
    • Skin

Properties

Applications

Our Abpromise guarantee covers the use of ab15148 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/500. Detects a band of approximately 120 kDa.
ICC/IF Use at an assay dependent concentration.
IHC-P 1/30. for 10 min at RT. Staining of formalin-fixed tissues requires boiling tissue sections in 10mM citrate buffer, pH 6.0 for 10 min followed by cooling at RT for 20 min.
IHC-Fr 1/50.

Target

  • Function
    Cadherins are calcium-dependent cell adhesion proteins. 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. 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.
  • Tissue specificity
    Non-neural epithelial tissues.
  • Involvement in disease
    Defects in CDH1 are the cause of hereditary diffuse gastric cancer (HDGC) [MIM:137215]. An autosomal dominant cancer predisposition syndrome with increased susceptibility to diffuse gastric cancer. Diffuse gastric cancer is a malignant disease characterized by poorly differentiated infiltrating lesions resulting in thickening of the stomach. Malignant tumors start in the stomach, can spread to the esophagus or the small intestine, and can extend through the stomach wall to nearby lymph nodes and organs. It also can metastasize to other parts of the body. Note=Heterozygous germline mutations CDH1 are responsible for familial cases of diffuse gastric cancer. Somatic mutations in the has also been found in patients with sporadic diffuse gastric cancer and lobular breast cancer.
    Defects in CDH1 are a cause of susceptibility to endometrial cancer (ENDMC) [MIM:608089].
    Defects in CDH1 are a cause of susceptibility to ovarian cancer (OC) [MIM:167000]. Ovarian cancer common malignancy originating from ovarian tissue. Although many histologic types of ovarian neoplasms have been described, epithelial ovarian carcinoma is the most common form. Ovarian cancers are often asymptomatic and the recognized signs and symptoms, even of late-stage disease, are vague. Consequently, most patients are diagnosed with advanced disease.
  • Sequence similarities
    Contains 5 cadherin domains.
  • Post-translational
    modifications
    During apoptosis or with calcium influx, cleaved by a membrane-bound metalloproteinase (ADAM10), PS1/gamma-secretase and caspase-3 to produce fragments of about 38 kDa (E-CAD/CTF1), 33 kDa (E-CAD/CTF2) and 29 kDa (E-CAD/CTF3), respectively. Processing by the metalloproteinase, induced by calcium influx, causes disruption of cell-cell adhesion and the subsequent release of beta-catenin into the cytoplasm. The residual membrane-tethered cleavage product is rapidly degraded via an intracellular proteolytic pathway. Cleavage by caspase-3 releases the cytoplasmic tail resulting in disintegration of the actin microfilament system. The gamma-secretase-mediated cleavage promotes disassembly of adherens junctions.
  • Cellular localization
    Cell junction. Cell membrane. Endosome. Golgi apparatus > trans-Golgi network. Colocalizes with DLGAP5 at sites of cell-cell contact in intestinal epithelial cells. Anchored to actin microfilaments through association with alpha-, beta- and gamma-catenin. Sequential proteolysis induced by apoptosis or calcium influx, results in translocation from sites of cell-cell contact to the cytoplasm. Colocalizes with RAB11A endosomes during its transport from the Golgi apparatus to the plasma membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • Arc 1 antibody
    • CADH1_HUMAN antibody
    • Cadherin 1 antibody
    • cadherin 1 type 1 E-cadherin antibody
    • Cadherin1 antibody
    • CAM 120/80 antibody
    • CD 324 antibody
    • CD324 antibody
    • CD324 antigen antibody
    • cdh1 antibody
    • CDHE antibody
    • E-Cad/CTF3 antibody
    • E-cadherin antibody
    • ECAD antibody
    • Epithelial cadherin antibody
    • epithelial calcium dependant adhesion protein antibody
    • LCAM antibody
    • Liver cell adhesion molecule antibody
    • UVO antibody
    • Uvomorulin antibody
    see all

Images

  • ab15148 staining E Cadherin in Human breast cancer MDA-MB-231 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were treated with ehanol (vehicle) as control or Origanummarjorana extract for 24 hours. Cells were fixed with 4% paraformaldehyde, permeabilized with 0.1% Triton X-100 in PBS and blocked with 5% milk for 30 minutes at room temperature. Samples were incubated with primary antibody overnight at 4°C. An Alexa Fluor 488-conjugated Goat anti-rabbit IgG (H+L) polyclonal (1/200) was used as the secondary antibody.

  • All lanes : Anti-E Cadherin antibody (ab15148) at 1/500 dilution (for 16 hours at 4°C)

    All lanes : Human OE33 cell - whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/1000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Observed band size: 120 kDa
    why is the actual band size different from the predicted?


    Exposure time: 1 minute


    Blocking Step: 5% BSA fro 1 hour at 23°C

    See Abreview

  • ab15148 staining human MCF10A cells by ICC/IF.  Cells were fixed with paraformaldehyde and blocked using 10% serum for 30 minutes at 25 °C.  The primary antibody was diluted 1/25 in TBST and incubated for 1 hour at 25 °C.  An Alexa Fluor® 555 goat anti-rabbit was used as the secondary antibody.

    See Abreview

  • ab15148 staining E Cadherin in Human AGS Gastric Carcinoma tissue sections by Immunohistochemistry (Frozen sections). The sections were acetone fixed and blocked in 5% serum for 1 hour at 23°C. The primary antibody was diluted 1/50 in blocking buffer and incubated with the sample for 1 hours at 23°C.  An HRP-conjugated Goat polyclonal to Rabbit IgG, diluted 1/200, was used as the secondary.

    See Abreview

  • ab15148 staining E Cadherin in Pig Cervix uteri tissue sections by IHC-P (Formaldehyde-fixed, Paraffin-embedded sections). Tissue samples were fixed with formaldehyde and blocked with 10% goat serum for 1 hour at 37°C; antigen retrieval was by heat mediation in 10mM citrate at pH 6 for 2 minutes. The sample was incubated with primary antibody (1/50) at 4°C for 12 hours. An HRP-conjugated goat polyclonal to rabbit IgG (undiluted) was used as secondary antibody.

    See Abreview

  • Immunohistochemical staining of formalin fixed paraffin embedded human skin using ab15148.
  • ab15148 staining E Cadherin in human stomach tissue section by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue underwent formaldehyde fixation before heat mediated antigen retrieval in Citrate pH 6.0 and then blocking with 5% serum for 1 hour at 23°C was performed. The primary antibody was used diluted 1/50 and incubated with sample for 1 hour at 23°C. A HRP conjugated goat polyclonal to rabbit IgG was used undiluted as secondary antibody.

    See Abreview

  • ab15148 staining E cadherin in Human AGS Gastric carcinoma cultured cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilzed with 0.025% Trton X-100 in TBS and blocked with 5% serum for 1 hour at 23°C. Samples were incubated with primary antibody (1/50 in blocking buffer) for 1 hour at 23°C. An undiluted HRP-conjugated goat anti-rabbit IgG polyclonal was used as the secondary antibody.

    See Abreview

References

This product has been referenced in:
  • Zhu Y  et al. Overexpression of microRNA-612 Restrains the Growth, Invasion, and Tumorigenesis of Melanoma Cells by Targeting Espin. Mol Cells 41:119-126 (2018). Read more (PubMed: 29385671) »
  • Gooskens SL  et al. TCF21 hypermethylation regulates renal tumor cell clonogenic proliferation and migration. Mol Oncol 12:166-179 (2018). WB ; Human . Read more (PubMed: 29080283) »
See all 86 Publications for this product

Customer reviews and Q&As

1-10 of 22 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Pig Cell (Swine testicle cells)
Permeabilization
Yes - Triton, 0.1%, 2min
Specification
Swine testicle cells
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Dec 15 2015

Application
ELISA
Sample
Human Cell (H9 embryonic stem cells)
Specification
H9 embryonic stem cells
Type
Indirect
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Oct 11 2013

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 10 minute(s) · Concentration: 5% · Temperature: 25°C
Sample
Human Cell (H9 embryonic stem cells)
Specification
H9 embryonic stem cells
Permeabilization
Yes - 0.01% Triton X100
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Oct 04 2013

Application
Western blot
Sample
Human Cell lysate - whole cell (MCF-7)
Loading amount
100000 cells
Specification
MCF-7
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Nov 19 2012

Question

Dear Sir/Madam,
Please find the completed form below and relevant image attached.
WB Questionnaire
1) Abcam product code ab15148

2) Abcam order reference number or product batch number

3) Description of the problem: Non-specific bands detected in positive and negative control, I selected the antibody as the reviews displayed a single band detected for E-cadherin which is what I need for my experiments.
4) Sample preparation:
Type of sample (whole cell lysates, fraction, recombinant protein…): Whole cell lysates
Lysis buffer RIPA buffer
Protease inhibitors: Protease cocktail inhibitor tablets (Roche).
Phosphatase inhibitors N/A
Reducing agent Laemmli buffer
Boiling for ≥5 min? yes
Protein loaded ug/lane or cells/lane 100,000 cells/well
Positive control MCF-7
Negative control MDA-MB-231
5) Percentage of gel 10%
Type of membrane Hybond ECL Nitrocellulose (GE Healthcare)
Protein transfer verified Yes (α-tubulin)
Blocking agent and concentration 5% milk in TBS
Blocking time 30 minutes
Blocking temperature Room temp.
6) Primary antibody (If more than one was used, describe in “additional notes”) :
Concentration or dilution 1/200
Diluent buffer 5% milk in TBS
Incubation time 20h
Incubation temperature: 4oC
7) Secondary antibody: Goat anti-rabbit IgG HRP (Dako)
Species: Goat
Reacts against: Rabbit
Concentration or dilution 1/2000
Diluent buffer 5% Milk in TBS
Incubation time 1h
Incubation temperature: Room temp
Fluorochrome or enzyme conjugate: HRP
8) Washing after primary and secondary antibodies:
Buffer TBST
Number of washes 3x 10min
9)Detection method
10) How many times have you run this staining? 4
Do you obtain the same results every time? yes
What steps have you altered to try and optimize the use of this antibody? Tested primary antibody concentration at 1/200-1/1000 and increased blocking time from 30 minutes to 60 minutes.

Read More
Answer

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.
The details you have kindly provided will enable us to investigate this case for you and this is also helpful in our records for monitoring of quality.
I would like to reassure you that ab15148 is tested and covered by our 6 month guarantee for use in WB and human samples. In the event that a product is not functioning in the applications cited on the product data sheet, we will be pleased to provide a credit note or free of charge replacement.
Reviewing this case, I would like to offer some suggestions to help optimise the results from ab15148.
1.) I recommend to run a no primary control, to make sure that it is not the secondary antibody producing the extra bands at 50 and 90kDa.
2.) When testing our antibodies, our lab uses 5% BSA as a blocking reagent, so I recommend switching to this instead of milk. Some antibodies give stronger, more specific signals on blots blocked with BSA instead of milk, so doing this may improve the results you are seeing, and reduce the non-specific bands. An example of the above is the western blot obtained with the Abcam GAPDH antibody ab9385 : www.abcam.com/ab9385
3.) Since the smallest extra bands are only in the positive control and it is smaller than the expected band, this could be a degradation band. I therefore suggest to review the proteinase inhibitors and always freshly prepare them.
I hope this information is helpful, thank you for your cooperation. Should the suggestions not improve the results, please do not hesitate to contact me again with the further requested details.

Read More

Answer

Thank you for contacting us.
The immunogen of ab53033 corresponds to the internal sequence of E-cadherin so this may not suitable as per your requirements.
The immunogens of antibodies ab15148, ab77287, ab128804, and ab133597 are located within the extra cellular domain of E-cadherin so these antibodies might be suitable. ab133597 has been tested in ICC/IF with mouse and rat cells so it is the most suitable candidate. However the other antibodies can be offered with either mouse or rat or with ICC/IF 100% Abreview discount.
The Abpromise guarantee is with ICC/IF when performed on fixed cells we do not know which antibody would be suitable for live cell imaging so the suitability has t o be determined by the end user.
I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.
Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More

Answer

Thank you for your phone call. Our current E Cadherin antibodies that bind to an extracellular domain of the human protein are:

ab8050
ab14015
ab15148
ab15150
ab22585
ab40772
ab45990
ab77287

I hope this helps, please let me know if you would like me to narrow down the list by any additional criteria.

Read More

Answer

Thank you for confirming this information and for your help and cooperation with this case. As requested, I have asked our accounting department to issue you with a credit note. This can then be redeemed against the invoice of a future order. As usual if you have any further questions regarding this credit note, please contact the accounts department by email at creditcontrol@abcam.com. The reference number, or credit ID, is xxxx. Please refer to this number in any correspondence with the accounting department. I would like to wish the customer good luck with their research. The technical team is always at your service, should you require further expert advice.

Read More

Answer

Thank you for confirming this information and for your help and cooperation with this case. As mentioned before, the protocol looks absolutely fine to me and the results with ab53033 are excellent. I can suggest the customer has regrettably received a bad vial. Therefore, I would be pleased to offer your customer a replacement or a credit note. I look forward to your reply.  

Read More

Question

LOT NUMBER gr46326-1 ORDER NUMBER xxxxDESCRIPTION OF THE PROBLEM No signal or weak signal SAMPLE •Species: human cell line •What’s cell line or tissue: human HCC cell line ex:huh7 hucct1 •Cell extract or Nuclear extract: cell extract PRIMARY ANTIBODY •Reacts against:anti-E cadherin •At what dilution(s) have you tested this antibody:1:500 •What dilution buffer was used: 5% skim milk in PBST •Incubation time: overnight >16hr •Incubation temperature: 4℃ •What washing steps were done:4 times, 10min every time DETECTION METHOD ECL ANTIBODY STORAGE CONDITIONS -20℃ SAMPLE PREPARATION •What lysis buffer was used: TPER+phospho stop+EDTA free •What protease inhibitors were used: phospho stop+EDTA free •What loading buffer was used: •Phosphatase inhibitors •Did you heat the samples: temperature and time: 95℃ 5min AMOUNT OF PROTEIN LOADED 66μg ELECTROPHORESIS/GEL CONDITIONS •Gel percentage : 6 TRANSFER AND BLOCKING CONDITIONS •Transfer conditions:PVDF, 100volt 90min •Buffer:5% skim milk in PBST •Blocking agent: milk, BSA, serum, what percentage: 5% skim milk •Incubation time:1hr •Incubation temperature:room temperature SECONDARY ANTIBODY •Reacts against:anti-rabbit IgG •At what dilution(s) have you tested this antibody: 1:10000 •Incubation time: 1hr •Wash steps: 4 times, 10min every time •Fluorochrome or enzyme conjugate:HRP HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? Dilution (from 1:1000 to 1:500), but results are still the same result

Read More
Answer

Thank you for contacting us. I am sorry to hear you have been experiencing problems with one of our antibodies. The quality of our products is important to us and I would like to reassure you that we investigate all customer complaints. I appreciate the time taken to complete our questionnaire, and after reading the answers provided I would like to ask additional questions: 1) Could you please confirm which positive control you have used? 2) May I ask whether you have checked your cell lines for the transcription of E-Cadherin, e.g. by RT-PCR? As your protocol looks absolutely fine to me, I would be pleased to offer you a replacement or a credit note, once I have received your reply. I look forward to hearing from you again.

Read More

1-10 of 22 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Sign up