Anti-Nodal antibody [5C3] (ab55676)

Reviews (5)Q&A (7)References (15)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nodal antibody [5C3] (ab55676)
  • Western blot - Anti-Nodal antibody [5C3] (ab55676)
  • Immunocytochemistry/ Immunofluorescence - Anti-Nodal antibody [5C3] (ab55676)
  • Flow Cytometry - Anti-Nodal antibody [5C3] (ab55676)

Key features and details

  • Mouse monoclonal [5C3] to Nodal
  • Suitable for: WB, IHC-P, ICC/IF, Flow Cyt
  • Reacts with: Mouse, Human
  • Isotype: IgG1

Overview

  • Product name

    Anti-Nodal antibody [5C3]
    See all Nodal primary antibodies

  • Description

    Mouse monoclonal [5C3] to Nodal

  • Host species

    Mouse

  • Tested applications

    Suitable for: WB, IHC-P, ICC/IF, Flow Cytmore details

  • Species reactivity

    Reacts with: Mouse, Human

  • Immunogen

    Recombinant fragment: RCEGECPNPV GEEFHPTNHA YIQSLLKRYQ PHRVPSTCCA PVKTKPLSML YVDNGRVLLD HHKDMIVEEC GC, corresponding to amino acids 275-347 of Human Nodal

    Run BLAST with BLAST the sequence with ExPASy Run BLAST with BLAST the sequence with NCBI

  • General notes

    This product was changed from ascites to tissue culture supernatant on 28th May 2019. Please note that the dilutions may need to be adjusted accordingly. If you have any questions, please do not hesitate to contact our scientific support team.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. 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, along with publications, customer reviews and Q&As

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.

  • Storage buffer

    pH: 7.4
  • Concentration information loading...

  • Purity

    Tissue culture supernatant

  • Clonality

    Monoclonal

  • Clone number

    5C3

  • Isotype

    IgG1

  • Light chain type

    kappa

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab55676 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 (3)
Use at an assay dependent concentration. Predicted molecular weight: 40 kDa.
IHC-P (1)
Use at an assay dependent concentration.
ICC/IF (1)
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 
Notes
WB
Use at an assay dependent concentration. Predicted molecular weight: 40 kDa.
IHC-P
Use at an assay dependent concentration.
ICC/IF
Use at an assay dependent concentration.
Flow Cyt
Use at an assay dependent concentration.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 

Target

  • Function

    Essential for mesoderm formation and axial patterning during embryonic development.

  • Involvement in disease

    Defects in NODAL are the cause of visceral heterotaxy autosomal type 5 (HTX5) [MIM:270100]. A form of visceral heterotaxy, a complex disorder due to disruption of the normal left-right asymmetry of the thoracoabdominal organs. It results in an abnormal arrangement of visceral organs, and a wide variety of congenital defects. Clinical features of visceral heterotaxy autosomal type 5 include situs inversus viscerum or situs ambiguus, congenital heart defect, transposition of the great vessels ventricular septal defect, atrial septal defect, truncuscommunis, and dextrocardia.

  • Sequence similarities

    Belongs to the TGF-beta family.

  • Cellular localization

    Secreted.
  • Information by UniProt

  • Database links

    • Alternative names

      • HTX5 antibody
      • MGC138230 antibody
      • Nodal antibody
      • Nodal Growth Differentiation Factor antibody
      • nodal homolog (mouse) antibody
      • Nodal homolog antibody
      • Nodal, Mouse, Homolog antibody
      • NODAL_HUMAN antibody
      see all

    Images

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nodal antibody [5C3] (ab55676)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nodal antibody [5C3] (ab55676)

      Nodal antibody (ab55676) used in immunohistochemistry at 3ug/ml on formalin fixed and paraffin embedded human endometrium cancer.

      This image was generated using the ascites version of the product.

    • Western blot - Anti-Nodal antibody [5C3] (ab55676)
      Western blot - Anti-Nodal antibody [5C3] (ab55676)

      Nodal antibody (ab55676) at 1ug/lane + HeLa cell lysate at 25ug/lane.

      This image was generated using the ascites version of the product.

    • Immunocytochemistry/ Immunofluorescence - Anti-Nodal antibody [5C3] (ab55676)
      Immunocytochemistry/ Immunofluorescence - Anti-Nodal antibody [5C3] (ab55676)

      ICC/IF image of ab55676 stained mouse embryonic stem cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab55676, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

      This image was generated using the ascites version of the product.

    • Flow Cytometry - Anti-Nodal antibody [5C3] (ab55676)
      Flow Cytometry - Anti-Nodal antibody [5C3] (ab55676)

      Overlay histogram showing HEK293 cells stained with ab55676 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab55676, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HEK293 cells fixed with 100% methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

      This image was generated using the ascites version of the product.

    References (15)

    Publishing research using ab55676? Please let us know so that we can cite the reference in this datasheet.

    ab55676 has been referenced in 15 publications.

    • Harpelunde Poulsen K  et al. Influence of Nodal signalling on pluripotency factor expression, tumour cell proliferation and cisplatin-sensitivity in testicular germ cell tumours. BMC Cancer 20:349 (2020). PubMed: 32326899
    • Margaryan NV  et al. The Stem Cell Phenotype of Aggressive Breast Cancer Cells. Cancers (Basel) 11:N/A (2019). PubMed: 30857267
    • Li Z  et al. Nodal Facilitates Differentiation of Fibroblasts to Cancer-Associated Fibroblasts that Support Tumor Growth in Melanoma and Colorectal Cancer. Cells 8:N/A (2019). PubMed: 31167491
    • Walewska E  et al. The Interaction Between Nodal, Hypoxia-Inducible Factor 1 Alpha, and Thrombospondin 1 Promotes Luteolysis in Equine Corpus Luteum. Front Endocrinol (Lausanne) 10:667 (2019). PubMed: 31632347
    • Jiang Y  et al. ALDH enzyme activity is regulated by Nodal and histamine in the A549 cell line. Oncol Lett 14:6955-6961 (2017). PubMed: 29181106
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
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    1-10 of 12 Abreviews or Q&A

    Question

    Since in the Data sheet of the ab55676 a IF picture is showed, I would say that the antibody was already tested for IF.

    I am working with male gonads at embryonic stage 13.5, the protocol I'm using is in the attached file.

    I have tried 3 different concentration of ab55676, 1:500, 1:1000 and 1:2000. The antibody is diluted in Block solution

    As secondary antibody I have tested 2: alexa fluor 488 donkey anti-mouse IgG (H+L) from Invitrogen and Donkey Anti-Mouse IgG (H+L) Cy3 from Jackson Immunoresearch and in my last experiment the incubation with the secondary Ab was for 15 and 30 min, with the hope to decrease the background. But I didn't get good results.


    I tried the M.O.M kit from Vector, but I didn't have success.

    I have attached one IF picture, the negative control (-) was made at the same time under the same conditions, the only difference is that the Nodal antibody was omitted and just the secondary Ab was added. Based in this result the next step will be to buy a new secondary antibody we are thinking in get this one

    http://products.invitrogen.com/ivgn/product/A21240

    Read More
    Answer

    Thank you for your reply.

    I am sorry about the confusion whether ab55676 was tested in IF or not, as you say the image on the datasheet clearly shows that is does work in IF, I am sorry I missed that.

    Having, looked over your protocol, I am in agreement with you that the issue seems to be non-specific binding of your secondary antibody. The only part of the protocol that I would say to be careful about is your block, as you should match the block to the host species of your secondary. I only mention this as you say you use goat serum for blocking but one of the secondary’s you used was raised in donkey. Also you do not need to incubate secondary antibodies overnight at 4C, in most cases 60 minutes at room temp is more than sufficient and any longer can cause high background.

    Therefore before you purchase a different secondary antibody, I would recommend that you repeat the experiment as before but alter the incubation time of your secondary antibody.

    Please let me know if there is anything else I can help you with

    Read More

    Western blot abreview for Anti-Nodal antibody

     Average
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Horse Tissue lysate - whole (Endometrium)
    Gel Running Conditions
    Reduced Denaturing (6% Stacking Gel, 10% Resolving Gel)
    Loading amount
    6 µg
    Specification
    Endometrium
    Blocking step
    BSA in PBS+Tween-20 as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
    Read More

    Abcam user community

    Verified customer

    Submitted Apr 12 2018

    Immunocytochemistry/ Immunofluorescence abreview for Anti-Nodal antibody

     Good
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (1 week Retinoic acid differentiation of NTERA-2)
    Specification
    1 week Retinoic acid differentiation of NTERA-2
    Blocking step
    Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C
    Fixative
    Paraformaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Oct 21 2015

    Western blot abreview for Anti-Nodal antibody

     Good
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    60 µg
    Gel Running Conditions
    Reduced Non-Denaturing (Native) (12)
    Sample
    Human Cell lysate - whole cell (LNCaP cell line)
    Specification
    LNCaP cell line
    Blocking step
    Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 26°C
    Read More

    Dr. Francesco Elia Marino

    Verified customer

    Submitted May 15 2014

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Nodal antibody

     Good
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    CAS BLOCK as blocking agent for 20 minute(s) · Concentration: 100% · Temperature: 26°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Sodium Citrate Buffer
    Sample
    Human Tissue sections (Human Prostate)
    Specification
    Human Prostate
    Permeabilization
    No
    Fixative
    Paraformaldehyde
    Read More

    Dr. Francesco Elia Marino

    Verified customer

    Submitted May 15 2014

    Question

    I already did some modification to our regular protocol. As I wrote in my previous e-mail I also did short incubation with secondary antibodies (If you read my e-mail again with more carefull you will note that). Could you recommend a secondary antibody?, that would be helpful.

    Read More
    Answer

    Thank you for your reply.

    I am sorry that I misread your original email with the protocol information, I know see that you tried the secondary antibody with both 15 and 30 minutes incubation times but that did not make a difference.

    Below are links to antibodies that I believe will work well for you, there are 3, all with a different fluorescent tag on them so that you can pick one based on which color you want to use:

    https://www.abcam.com/Goat-polyclonal-Secondary-Antibody-to-Rabbit-IgG-H-L-DyLight-488-pre-adsorbed-ab96899.html
    https://www.abcam.com/Goat-polyclonal-Secondary-Antibody-to-Rabbit-IgG-H-L-DyLight-650-pre-adsorbed-ab96898.html
    https://www.abcam.com/Donkey-polyclonal-Secondary-Antibody-to-Rabbit-IgG-H-L-DyLight-594-pre-adsorbed-ab98500.html



    Under our Abpromise, all of the above antibodies are guaranteed to work in IF and if for whatever reasons the antibody fails to work as stated then we will be happy to either to replace or refund the antibody.

    If there is anything else I can help you with, please let me know.

    Read More

    Question


    Thanks for your answer,



    In our Laboratory we study the expression and level of different molecules through immunofluorescense, it mean we have a significant number of primary and secondary antibodies and that I have tried different ways to make the Nodal ab55676 works without success. Additionally this antibody doesn't work in our system in vitro. I'm asking you for a sample of the Anti-Nodal antibody (ab39242) because we trust in Abcam products and I'm almost sure that the ab39242 will work better so, Is possible to get a sample of this antibody? Please, just take your time to answer my question, don't waste your time sending me others links.

    Thanks,

    Read More
    Answer

    Thank you for taking the time to respond and for contacting Abcam.

    With over 70,000 products it is logistically impossible for us to offer free samples. I hope you understand.

    Please let me know if you have any questions.

    Read More

    Question

    Recently I bought the antibody mouse monoclonal to Nodal ab55676. Still I don't have results in which I can trust because with the recommended dilusion and even more diluted I got a lot of background (IF). I'm wondering (and I wil be very happy) if is possible to get a sample of the Anti-Nodal antibody (ab39242).

    Read More
    Answer

    Thank you for contacting Abcam.

    I am sorry that you are having problems with ab55676 in immunofluorescence. As we had not tested this antibody in IF before, we were not sure how the antibody would work in that application. I would be very happy to see if I can give any protocol suggestions that would improve the staining that you are getting.

    Could you provide me with a description of the protocol that you are using, sample types, type of fix, blocking agent used, primary antibody concentration and incubations times. Also do you use any type of detergent to permeablilze your samples.

    I look forward to your reply and helping you to achieve better staining.

    Read More

    Question

    Hello,



    Recently I bought the Mouse monoclonal antibody ab55676. I have tried this antibody several times in a mouse model to study the role that Nodal could play in teramota incidence, a kind of cancer. Until now I don't have any good result because this antibody show a lot of unspecific binding. I'm writing to you with the hope to get a sample of the Anti-Nodal antibody ab3924 which is rabbit, and I think could work better in my samples..





    I will be waiting for your answer,

    Read More
    Answer

    Thank you for contacting Abcam.

    I am sorry to hear that Ab55676 has not performed well for you. Although the peptide sequence has excellent identity with the mouse homolog, this product is not guaranteed to work in mouse tissues. High background is a very common issue when working in tissues of the same species as the host speciesof the primary anti-body. These may be reduced by using a secondary antibody that has been pre-absorbed or by direct conjugation of your label to the primary antibody.

    Our EasyLink conjugation kits offer an effective and simple way to directly conjugate labels to your primary antibody. More information may be found at the following link:

    https://www.abcam.com/index.html?pageconfig=resource&rid=13148&source=pagetrap&viapagetrap=easylink

    We have 12 secondary antibodies against mouse IgG1, they may be viewed at the following link:

    https://www.abcam.com/index.html?pageconfig=searchresults&search=IgG1&pt=7&sk=targsp&sv=2&sn=Mouse&l=2&fViewMore=1

    Please let me know if you have any questions.

    Read More

    Question

    LOT NUMBER GR7047-4 ORDER NUMBER 884826 DESCRIPTION OF THE PROBLEM No signal or weak signal SAMPLE Human cell MAD-MB231 medium and cell extraction PRIMARY ANTIBODY Dilution 1:500,1:200, overnight ,wash three times DETECTION METHOD ECL POSITIVE AND NEGATIVE CONTROLS USED YES ANTIBODY STORAGE CONDITIONS Aliquoted and stored -20℃ SAMPLE PREPARATION RIPA buffer AMOUNT OF PROTEIN LOADED 50ug and 100ug ELECTROPHORESIS/GEL CONDITIONS 12% reducing and non-reducing (both tried) TRANSFER AND BLOCKING CONDITIONS 100 V 40 min, PBS-T SECONDARY ANTIBODY Goat anti mouse santa cruz (1:1000) 2 hours HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes

    Read More
    Answer

    Further to my previous response to your complaint, I would like to pass some more information which you may find useful. The antibody is (NODAL monoclonal antibody (ab55676) has been tested and characterized in Western blot on several cell lines, such as HeLa, PC-12, and NIH/3T3. I would suggest using one of these cell lines (if you have access to them) as good well characterized positive control alongside with the samples. Please find the cell lysate preparation (denatured) protocol below: Step-by-step procedure: 1. Prepare cell lysate by homogenization in ice-cold modified RIPA Lysis Buffer. And add sufficient volume of protease Inhibitor (add 100 ul every 1mL RIPA Lysis Buffer), then vortex. 2. After centrifugation, determine the protein concentration of the cell lysate. 3. Boil the lysate for 10 minutes in 1X SDS sample buffer. 4. The lysate is ready to load onto SDS-PAGE for Western blotting. (20~50 μg per lane is recommended for mini gel.) Tested Applications: Western blot Reagents: _ RIPA Lysis Buffer: 50mM Tris-HCl pH 7.4,150mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1mM PMSF( PMSF should be added before use) _ 1X SDS sample buffer: 2% SDS, 300mM, 300mM _-mercaptoethanol, 50mM Tris-HCl pH 6.8,10% Glycerol, 0.01% Bromophenol blue If you need any further assistance in the future, please do not hesitate to contact me.

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    1-10 of 12 Abreviews or Q&A

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