Overview

  • Product name
    Anti-Corticotropin Releasing Factor antibody [4H9]
    See all Corticotropin Releasing Factor primary antibodies
  • Description
    Mouse monoclonal [4H9] to Corticotropin Releasing Factor
  • Host species
    Mouse
  • Specificity
    Reacts with CRF. Does not react with bovine serum albumin, insulin, ovalbumin or epidermal growth factor.
  • Tested applications
    Suitable for: ELISA, IHC-P, IHC-Fr, Inhibition Assaymore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Rat
  • Immunogen

    Full length protein (Human)

  • Epitope
    The unique lysine residue (Lys-36) is likely to be involved in the antigenic determinant of ab35748. Lysine modification by CA blocks binding completely.

Properties

Applications

Our Abpromise guarantee covers the use of ab35748 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
ELISA Use at an assay dependent concentration.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval via the microwave method before commencing with IHC staining protocol.
IHC-Fr 1/100 - 1/7000.
Inhibition Assay Use at an assay dependent concentration. Human serum, containing CRF-binding protein, inhibits the binding of CRF to this antibody.

Target

  • Relevance
    CRF precursor is a 41-amino acid peptide derived from a 196-amino acid preprohormone. CRF is a neuroendocrine hormone secreted by the paraventricular nucleus (PVN) of the hypothalamus and is a key player in the hypothalamic pituitary adrenal axis. It is also synthesized in peripheral tissues, such as T lymphocytes and is highly expressed in the placenta. Neuronal CRF is important in a variety of diverse roles including, memory, depression, stress, kidney function, parturition, metabolism and feeding (to mention a few).
  • Cellular localization
    Cytoplasmic and Secreted
  • Database links
  • Alternative names
    • Corticoliberin antibody
    • Corticotropin releasing factor antibody
    • Corticotropin releasing hormone antibody
    • CRF antibody
    • Crh antibody
    • CRH1 antibody
    see all

Images

  • ab35748 staining Corticotrophin Releasing Factor in Rat brain tissue by Immunohistochemistry (Frozen sections). The sections were fixed in methanol prior to blocking with PBS with 1.5% goat serum for 20 minutes at 22°C. The primary antibody was diluted 1/100 in the blocking buffer and incubated with the sample for 10 hours at 4°C. A Dylight® 594-conjugated polyclonal antibody was used as the secondary antibody, diluted 1/80.

    See Abreview

References

This product has been referenced in:
  • Kravchenko IV & Furalev VA Monoclonal antibodies directed against two different corticotropin-releasing factor determinants. Hybridoma 13:59-64 (1994). Read more (PubMed: 7515375) »
See 1 Publication for this product

Customer reviews and Q&As

1-10 of 11 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Rat Tissue sections (Paraventricular nucleus brain)
Antigen retrieval step
None
Permeabilization
Yes - 0.1% Triton-X 100
Specification
Paraventricular nucleus brain
Blocking step
Serum as blocking agent for 45 minute(s) · Concentration: 10% · Temperature: 22°C
Fixative
Paraformaldehyde

Dr. Fiona Shenton

Verified customer

Submitted Jun 26 2017

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Rat Tissue sections (Brain)
Specification
Brain
Fixative
Paraformaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: citrate
Permeabilization
Yes - Triton-x
Blocking step
Casein as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Oct 15 2012

Answer

Thank you for contacting us.

As we discussed on the phone, adding a BSA block and trying a shorter antigen retrieval time may help to reduce the background for ab7963, ab59023, and ab75168, while trying a 1:100 dilution for ab35748 may help to improve your signal. Please let me know how your results turn out. If you these antibodies continue to give poor results, I will be happy to offer you free of charge replacements, credits, or refunds to the antibodies that are guaranteed for use on rat samples. Please let me know your original order or PO numbers and how you would like to proceed and I will be happy to help you further.

Read More

Answer

Thank you for contacting us and your patience while I double checked with the laboratory.

The antibody will recognize an epitope around the Lysine 36 of the mature protein, as the lysine modification blocks the binding completely. The lysine is in position 36 of the corticoliberin - using the full length sequence, it would be the lysine in position 189. http://www.uniprot.org/uniprot/P06850

For information regarding the immunogen used to raise this antibody please see the Kravchenko reference, aslisted on the datasheet, PMID: 7515375 . It is unclear if a peptide or full-length Protein was used as the immunogen; I do not have access to the journal to verify which. As this is a monoclonal antibody, the antibody has only one epitope (the above mentioned).

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Read More

Answer

Merci pour ces informations. Je devine qu'un démasquage de l'antigène a également été tenté à pH 6.0 comme recommandé par le laboratoire qui produit cet anticorps, voir mon email du 21 novembre 2011 ci-après. Pourriez-vous s'il vous plait me confirmer ceci? Le laboratoire vient de m'informer que le tampon recommandé pour le démasquage de l'antigène est un tampon citrate pH 6,0. Placer les lames dans le tampon et chauffer au micro-ondes pendant 10 minutes. Laisser refroidir pendant 15 minutes et rincer avec du tampons TBS. Préparation du tampon de citrate pH 6.0 (solution 20x): 10,5 g acide citrique dans 250 ml d'eau distillée. Préparation d'une solution 1x : 25 ml de solution 20x 475 ml d'eau distillée Ajuster à pH 6,0 avec NaOH 1M.

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Question
Answer

Le laboratoire vient de m'informer que le tampon recommandé pour le démasquage de l'antigène est un tampon citrate pH 6,0. Placer les lames dans le tampon et chauffer au micro-ondes pendant 10 minutes. Laisser refroidir pendant 15 minutes et rincer avec du tampons TBS. Préparation du tampon de citrate pH 6.0 (solution 20x): 10,5 g acide citrique dans 250 ml d'eau distillée. Préparation d'une solution 1x : 25 ml de solution 20x 475 ml d'eau distillée Ajuster à pH 6,0 avec NaOH 1M. Si vous rencontrez toujours des problèmes avec cet anticorps, nous serions ravis si vous pouviez nous communiquer quelques détails concernant votre protocole en remplissant le questionnaire ci-joint. Ces informations sont très importantes pour nous car elles nous permettent d'investiguer la source du problème rencontré avec cet anticorps et de prendre les dispositions nécessaires pour assurer une bonne qualité de nos produits.

Read More

Answer

Thank you for contacting Abcam. Regarding ab35748, this monoclonal antibody was generated using a full length protein as the immunogen.  Unfortunately, epitope mapping has not been performed, although K36 is likely to be a part of the immunogen.  We do not have a specific blocking peptide or recommended protein in our catalog to use for blocking. I apologize for any inconvenience this may cause.  Please do not hesitate to contact us if you have any additional questions.

Read More

Question

Dear Technical support Group! I have a customer who is not successful with the following antibody and needs some advise: AB35748,  anti  CRF Corticotropin relaesing factor Here is her mail: Hope you are well. I am following up on the ab products bought in June/July this year including ab35748. We have not been able to get this antibody to work at all. I have detailed below the variations that Marilyn in the lab has tried without success. Do you think you could please get a technical opinion or suggestion from Abcam for us? Or let me know who to contact and I can do that? If there are no technical issues, maybe the ab itself is the problem in which case we would like to find out about replacing it. Here is a brief report of what I have done did not put in all the steps as I followed the AbCam protocol, what I have reported is the slight variations according to the detection used.   CRF Corticotropin releasing factor [4H9} ab 35748   aliquoted and froze at -20degrees Celcius   First tried on 27-07-2011   Immunohistochemistry as per AbCam protocol   These are my variations   Hydrogen peroxide/methanol 15mins   Blocked with normal goat serum 25mins   Antigen retrieval with 0.01M Citrate buffer in a)pressure cooker  2mins and b)microwave 10mins   Antibody dilutions 1/100, 1/50, 1/20  o/n @ 4 deg. C   Secondary antibody Envision goat anti mouse (Dakocytomation)   Detection with DAB   Tried several times  NO STAINING   Control tissue:Human gastric,small intestine,colon. Pig tissue: brain,intestine The tissue was fixed using cold 4% paraformaldehyde. Once fixed for 24 hours it was transferred to 70% ethanol and refridgerated until it was processed over a 24 hour cycle through 10% buffered formalin, 70% to absolute alcohol, xylol and finally to paraffin wax embedding. Dont think the antibody is working.   Do you have any suggestions   Sorry cannot get it to work dont know what else to try. Do you want to contact the company and maybe they can offer some other suggestions   Thank you in advance for your help, it is very much appreciated.   Best Regards

Read More
Answer

Thank you very much for contacting me again and for passing some useful information. Although, pig has not been tested for cross-reaction; ab35748 should recognize Corticotropin Releasing Factor in human samples. I can offer your troubled customer either a new vial as free of charge replacement or a credit note which can be used in future purchase. Please do let me know how you wish to proceed. I look forward to hearing from you soon.

Read More

Answer

Thank you for your enquiry regarding ab35748 and for taking the time to provide some useful details of the experiments. I am very sorry to hear that you are having problems with this antibody. Dr. Nitsan Kozlovsky has submitted an Abreview in March 2011. I am sorry we do not have any additional information regarding the protocol; the information we have is already published on the datasheet under their Abreview. Alternatively you can request protocol from them by sending them an email. To send him an email clicks on their name and send them a message by logging into your Abcam account. Alternatively I can also help you with troubleshooting. I would appreciate if you could fill the attached questionnaire and send it to me as early as possible. I will look forward to hearing from you soon.

Read More
Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (brain)
Specification
brain
Fixative
Methanol
Blocking step
Serum as blocking agent for 20 minute(s) · Concentration: 1.5% · Temperature: 22°C

Dr. Nitsan Kozlovsky

Verified customer

Submitted Mar 02 2011

1-10 of 11 Abreviews or Q&A

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