Overview

  • Product name

    Anti-N Cadherin antibody [EPR1791-4]
    See all N Cadherin primary antibodies
  • Description

    Rabbit monoclonal [EPR1791-4] to N Cadherin
  • Host species

    Rabbit
  • Specificity

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

  • Tested applications

    Suitable for: WB, IHC-Pmore details
    Unsuitable for: Flow Cyt or ICC/IF
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human N Cadherin aa 150-250 (extracellular). The exact sequence is proprietary.

  • Positive control

    • WB: A549, PC-3, HepG2, C6, Human brain, Mouse brain, and Rat brain lysates; IHC-P: Human liver, and Human cardiac muscle tissues;
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab76011 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 1/5000 - 1/20000. Predicted molecular weight: 100 kDa.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

  • Application notes
    Is unsuitable for Flow Cyt or ICC/IF.
  • 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. CDH2 may be involved in neuronal recognition mechanism. In hippocampal neurons, may regulate dendritic spine density.
    • Sequence similarities

      Contains 5 cadherin domains.
    • Cellular localization

      Cell membrane.
    • Information by UniProt
    • Database links

    • Alternative names

      • CADH2_HUMAN antibody
      • Cadherin 2 antibody
      • Cadherin 2 N cadherin neuronal antibody
      • Cadherin 2 type 1 antibody
      • Cadherin 2 type 1 N cadherin neuronal antibody
      • Cadherin 2, type 1, N-cadherin (neuronal) antibody
      • Cadherin-2 antibody
      • Cadherin2 antibody
      • Calcium dependent adhesion protein neuronal antibody
      • CD325 antibody
      • CD325 antigen antibody
      • CDH2 antibody
      • CDHN antibody
      • CDw325 antibody
      • CDw325 antigen antibody
      • N cadherin 1 antibody
      • N-cadherin antibody
      • NCAD antibody
      • Neural cadherin antibody
      • OTTHUMP00000066304 antibody
      • OTTHUMP00000067378 antibody
      see all

    Images

    • All lanes : Anti-N Cadherin antibody [EPR1791-4] (ab76011) at 1/1000 dilution

      Lane 1 : HeLa Whole Cell Lysate
      Lane 2 : HeLa Whole Cell Lysate (Scraped)
      Lane 3 : Human Brain Tissue Lysate
      Lane 4 : Mouse Brain Tissue Lysate
      Lane 5 : Rat Brain Tissue Lysate
      Lane 6 : MCF7 Whole Cell Lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 100 kDa
      Observed band size: 125 kDa
      why is the actual band size different from the predicted?



      This blot was produced using a 4-12% Bis-tris under the MOPS buffer system. The gel was run at 200V for 55 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was blocked for an hour using 3% milk before ab76011 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/1000 dilution and 1/20000 dilution respectively. Antibody binding was detected using Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human cardiac muscle tissue sections labeling N Cadherin with purified ab76011 at 1:50 dilution (1.94 µg/ml). Heat mediated antigen retrieval was performed Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control:PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Lane 1: HEK-293T wildtype cell lysate (20 µg)

      Lane 2: CDH2 HEK-293T knockout cell lysate (20 µg)

      Lanes 1 - 2: Merged signal (red and green). Green - ab76011 observed at 100 kDa. Red - loading control, ab8245 observed at 37 kDa.

      ab76011 was shown to react with N Cadherin in HEK-293T wildtype. Loss of signal was observed when knockout sample ab263843 was used. Wild-type and N Cadherin knockout samples were subjected to SDS-PAGE. ab76011 and Anti-GAPDH antibody [6C5] - Loading Control?(ab8245) were incubated overnight at 4°C at 1 in 5000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773)? and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
    • All lanes : ab76011, Anti-N Cadherin antibody [EPR1791-4] (Left) or ab207608, Anti-N Cadherin antibody [EPR19654] (Right) at 1/1000 dilution

      Lane 1 : A549 (Human lung carcinoma epithelial cell) whole cell lysates prepared in RIPA lysis method with 5% NFDM/TBST
      Lane 2 : PC-3 (Human prostate adenocarcinoma epithelial cell) whole cell lysates prepared in RIPA lysis method with 5% NFDM/TBST
      Lane 3 : HCT 116 (Human colorectal carcinoma epithelial cell) whole cell lysates prepared in RIPA lysis method with 5% NFDM/TBST
      Lane 4 : HCT 116 (Human colorectal carcinoma epithelial cell) whole cell lysates prepared in 1%SDS Hot lysis method with 5% NFDM/TBST
      Lane 5 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate prepared in RIPA lysis method with 5% NFDM/TBST
      Lane 6 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate prepared in 1%SDS Hot lysis method with 5% NFDM/TBST
      Lane 7 : C6 (Rat glial tumor glial cell) whole cell lysates prepared in RIPA lysis method with 5% NFDM/TBST
      Lane 8 : C6 (Rat glial tumor glial cell) whole cell lysates prepared in 1%SDS Hot lysis method with 5% NFDM/TBST
      Lane 9 : Human brain lysates prepared in 1%SDS Hot lysis method with 5% NFDM/TBST
      Lane 10 : Mouse brain lysates prepared in 1%SDS Hot lysis method with 5% NFDM/TBST
      Lane 11 : Rat brain lysates prepared in 1%SDS Hot lysis method with 5% NFDM/TBST

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 100 kDa
      Observed band size: 110-130 kDa why is the actual band size different from the predicted?



      The molecular weight observed is consistent with what has been described in the literature (PMID: 22553038). This antibody fails to detect N Cadherin in HCT 116 cell which is positive described in the literature (PMID: 23431386 and 26540342)

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human liver tissue sections labeling N Cadherin with purified ab76011 at 1:50 dilution (1.94 µg/ml). Heat mediated antigen retrieval was performed Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control:PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Immunohistochemistry of kidney carcinoma staining N Cadherin with ab76011 at 1μg/ml

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

    • ab76011 staining N Cadherin in Mouse pancreas tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA + 1% FBS for 2 hours at room temperature; antigen retrieval was by heat mediation in a citrate buffer pH6. Samples were incubated with primary antibody (1/500 in 1% BSA + 1% FBS) for 16 hours at 4°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:

    • Cheng B  et al. MicroRNA-122 inhibits epithelial-mesenchymal transition of hepatic stellate cells induced by the TGF-ß1/Smad signaling pathway. Exp Ther Med 17:284-290 (2019). Read more (PubMed: 30651793) »
    • Xu C  et al. NPTX2 promotes colorectal cancer growth and liver metastasis by the activation of the canonical Wnt/ß-catenin pathway via FZD6. Cell Death Dis 10:217 (2019). Read more (PubMed: 30833544) »
    See all 46 Publications for this product

    Customer reviews and Q&As

    1-10 of 18 Abreviews or Q&A

    This product is known to not work in this application or species.
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Mouse Cell (Brain)
    Permeabilization
    No
    Specification
    Brain
    Blocking step
    Serum as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 28°C
    Fixative
    Paraformaldehyde

    Abcam user community

    Verified customer

    Submitted Oct 09 2019

    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Cow Tissue sections (oviductal tissues)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Tris-EDTA pH9.0
    Permeabilization
    No
    Specification
    oviductal tissues
    Blocking step
    Serum as blocking agent for 20 minute(s) · Concentration: 2.5% · Temperature: 25°C
    Fixative
    Paraformaldehyde

    Dr. Yoshihiko Kobayashi

    Verified customer

    Submitted Sep 21 2016

    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (MCF-7 cell)
    Gel Running Conditions
    Reduced Denaturing (10%)
    Loading amount
    30 µg
    Specification
    MCF-7 cell
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C

    Abcam user community

    Verified customer

    Submitted Mar 11 2016

    Application
    Immunohistochemistry (Frozen sections)
    Sample
    Mouse Tissue sections (Embryonic mouse brain coronal cortical section)
    Permeabilization
    Yes - Triton X 100
    Specification
    Embryonic mouse brain coronal cortical section
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
    Fixative
    Paraformaldehyde

    Dr. Bhavin Shah

    Verified customer

    Submitted Oct 20 2015

    Application
    Western blot
    Loading amount
    50 µg
    Gel Running Conditions
    Reduced Denaturing (10)
    Sample
    Mouse Cell lysate - whole cell (MEFs)
    Specification
    MEFs
    Blocking step
    (agent) for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

    Abcam user community

    Verified customer

    Submitted Dec 16 2013

    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    1%BSA 1%FBS as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citrate buffer pH 6
    Sample
    Mouse Tissue sections (Pancreas)
    Specification
    Pancreas
    Permeabilization
    Yes - TBS-Tween 0,1%
    Fixative
    Formaldehyde

    Abcam user community

    Verified customer

    Submitted Jun 06 2013

    This product is known to not work in this application or species.
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Mouse Cell (MEFs wild type)
    Specification
    MEFs wild type
    Fixative
    Paraformaldehyde
    Permeabilization
    Yes - 0,3% Triton PBS
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: RT°C

    Abcam user community

    Verified customer

    Submitted Mar 18 2013

    Answer

    Thank you for contacting us and your interest in our products.
    The anti-N Cadherin antibody [8C11], ab19348, was raised against a bacterial fusion protein corresponding to the extracellular domain of N-cadherin (a domain between EC3 and EC4, amino acid residues 92-593 from human N-cadherin).
    The anti-N Cadherin antibody [EPR1791-4], ab76011, meanwhile was raised against a synthetic peptide taken from within the residues 100-200 of human N-cadherin , also within the extracellular domain of the protein.
    I hope this information has been of help. If you have any further questions, please do not hesitate to contact us again.

    Read More

    Answer

    Thank you for your reply.

    Can you please confirm what species your samples are from, including cell line or tissue type?

    Also what is the order number for this product? I will need to ensure that the order is not more than 6 months old before offering a replacement.

    What are the MWs of the bands in the WB image you had sent?

    Thank you for your cooperation and I look forward to your reply.

    Read More

    Question

    Thank-you for your help. I copied the answers to the questions you gave me bellow and have attached scanned images from our best western blots,

    1) Abcam product code: ab1791

    2) Abcam order reference number or product batch number

    3) Description of the problem: Multiple bands in WB

    4) Sample preparation:

    Type of sample (whole cell lysates, fraction, recombinant protein…): Cell lysates

    Lysis buffer: Boston BioProducts RIPA Buffer

    Protease inhibitors: Complete Mine (Roche)

    Phosphatase inhibitors: PhosSTOP (Roche)

    Reducing agent: 2-Mercaptoethanol

    Boiling for ≥5 min?: yes

    Protein loaded ug/lane or cells/lane:10 and 20 ug/lane

    Positive control: yes

    Negative control: yes

    5) Percentage of gel---10%

    Type of membrane: nitrocellulose

    Protein transfer verified: yes

    Blocking agent and concentration: 5% bovine serum albumin (BSA)

    Blocking time: 1hr

    Blocking temperature: Room Temp.

    6) Primary antibody (If more than one was used, describe in “additional notes”) : ab1791

    Concentration or dilution: 1:1000

    Diluent buffer: Tris-buffered saline with Tween (TBST)

    Incubation time: 12 hours

    Incubation temperature: Cold Room

    7) Secondary antibody:

    Species: rabbit

    Reacts against: ab1791

    Concentration or dilution: 1:1000

    Diluent buffer: TBST

    Incubation time: 1 hour

    Incubation temperature: Room Temp.

    Fluorochrome or enzyme conjugate:HRP

    8) Washing after primary and secondary antibodies:

    Buffer: TBST

    Number of washes: 3

    9)Detection method: Manuel exposure of X-ray films

    10) How many times have you run this staining?: Twice

    Do you obtain the same results every time?: Yes

    What steps have you altered to try and optimize the use of this antibody?:

    Protein concentration, running time, cutting NC film to desired molecular weight using a molecular weight marker

    Read More
    Answer

    Thank you for your reply.

    You're having trouble with the ab76011, anti-N cadherin antibody, correct? Not the ab1791 histone H3? You had included ab76011 in your initial email, but ab1791 in your reply.

    Do you have a lot # from the vial? I see an order, but that was from June 2010. Is this your order?

    What species are your cells from?

    Have you run a loading control like beta actin?

    From the WB image that you sent, it looks like you're seeing "reverse banding" which signifies that the primary and/or secondary antibodies are too concentrated. We recommend using the primary from 1/5000 - 1/20,000 and incubating this overnight at 4C with 5% BSA in TBST. For the secondary, you should try 1/10,000 - 1/30,000and incubate this for 1 hr at RT with 5% BSA in TBST (check the manufacturer's recommendations for dilution recommendations).

    Would you be able to label the MW of the bands in the images you had sent? I'm pretty sure that diluting the primary and secondary more will resolve a lot of the background issues, but I'd like to see if you're getting any bands at the predicted 100 kDa.

    I hope this information helps. Please let us know if the protocol recommendations do not improve the banding.

    Read More

    1-10 of 18 Abreviews or Q&A

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com

    Sign up