Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

Thanks so much for taking the time to complete this survey. I am glad to hear the replacement antibody is working as expected. If you have any questions, please do not hesitate to contact us.

Best of luck in your research.

Thank you for contacting us.

The link to our indirect ELISA protocol is:
https://www.abcam.com/index.html?pageconfig=resource&rid=11389

I have also attached the protocol as pdf-file.

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

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https://www.abcam.com/abreviews

Thank youf or your inquiry.

We have many goat anti rabbit IgG HRP conjugated antibodies:

Please use the links below to review the datasheets of three of them:

ab97051

https://www.abcam.com/Goat-Rabbit-IgG-HL-HRP-ab97051.html (or use the following: https://www.abcam.com/Goat-Rabbit-IgG-HL-HRP-ab97051.html).

ab97080

https://www.abcam.com/Goat-Rabbit-IgG-HL-HRP-preadsorbed-ab97080.html (or use the following: https://www.abcam.com/Goat-Rabbit-IgG-HL-HRP-preadsorbed-ab97080.html).

ab97200

https://www.abcam.com/Goat-Rabbit-IgG-Fc-HRP-ab97200.html (or use the following: https://www.abcam.com/Goat-Rabbit-IgG-Fc-HRP-ab97200.html).

Unfortunately, we do not have any rabbit anti mouse IgG1 HRP conjugated antibodies in our catalog. But do have anti mouse IgG1 antibodies from other host animals like goat and rat:

ab97240

https://www.abcam.com/Goat-Mouse-IgG1-HRP-ab97240.html (or use the following: https://www.abcam.com/Goat-Mouse-IgG1-HRP-ab97240.html).

ab20043

https://www.abcam.com/Goat-Mouse-IgG-Fc-HRP-ab20043.html (or use the following: https://www.abcam.com/Goat-Mouse-IgG-Fc-HRP-ab20043.html).

ab99603

https://www.abcam.com/Rat-monoclonal-SB77e-Mouse-IgG1-HL-HRP-preadsorbed-ab99603.html (or use the following: https://www.abcam.com/Rat-monoclonal-SB77e-Mouse-IgG1-HL-HRP-preadsorbed-ab99603.html).

I hope this information is helpful. Please do not hesitate to contact me again with any further questions.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement for one vial of ab6721 (which can be used as secondary with ab65166).

To check the status of the order please contact our Customer Service team and reference this number.

Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

I wish you the best of luck with your research.

Gracias por tu respuesta.

Acabo de realizar un pedido totalmente gratuito del anticuerpo ab97051. El número de pedido es XXXX. Si necesitáis contactarnos en referencia al mismo, no olvidéis facilitar dicho número.

Recibiréis una confirmación automática del mismo informando sobre la fecha estimada de entrega y detalles del pedido.

Si te parece, en lugar de enviarte los dos anticuerpos en el mismo pedido, os mandamos solo uno, y lo usáis en vuestra aplicación. Si os funciona perfectamente me respondéis a este correo para que os mande la segunda unidad.

Creo que es más razonable hacerlo así, por si tuvierais algún problema con el anticuerpo.

Perdona por las molestias, y mucha suerte en vuestra investigación.

Thank you for your enquiry. I appreciate theamount of detail you have provied andI am sorry to hearof the difficulties you are experiencing regarding ab97051.

You have done a good job of troubleshooting which leads me to believe you have received a faulty vial of ab97051. I am happy to offer a replacement vial, either of another vial of ab97051 or alternatively a different anti-rabbit IgG (HRP) antibody.

Please let me know how you wish to proceed. In your reply, please include the PO number or Abcam order reference number associated with this purchase.

Thank you for your email.

I am sorry to hear that the problem indeed is coming from the secondary antibody.
As I learned from my colleague, a free of charge replacement has already been offered to you and is on its way to you, for delivery tomorrow.

I hope this replacement product will work as expected. Please let me know should itnot, or if you have any further questions.

Thank you for contacting us.

I'm sorry to hear that you are obtaining high background. As we discussed over the phone,I would suggest to try a no primary control so that we candetermine if the primary or the secondary antibody is causing the problem.

Please let me know what results you have obtained and we can discuss this case further.

I wish you good luck and look forward to hear back from you.

Thank you for getting back to me and for specifying the sample types.
As you have kindly explained to me in your previous e-mail, the primary and the secondary antibodies worked with the positive control but the signal was very weak using lysates from sheep cardiac tissues.
This antibody (ab3583) should recognize sheep Leptin, the cross-reactivity with this species has been tested and confirmed.
After reading through the detailed protocol you kindly forwarded to Abcam, I would like to make the following comments/suggestions:
1) Lysis buffer:
Leptin is secreted so potentially it can be degraded very quickly. It is very important to prepare the samples from fresh tissues and use lysis buffer which contains proteinase inhibitors. The whole process should be carried out on ice. I understand from the initial e-mail that Laemmli buffer was used for lysing the samples. I would rather suggest using RIPA buffer to prevent any unwanted protein degradation in the sheep cardiac tissues. The recipe of RIPA can be found at this website:
https://www.abcam.com/index.html?pageconfig=resource&rid=11379#A1
2) Loading amount:
Try to load at least 25-30 ug total tissue lysate per lane onto the gel.
3) Dilution:
You may need to optimize the dilutions of the primary antibody to find out the most suitable working dilution range for sheep cardiac tissue lysate. Brain tissue has high levels of leptin but cardiac tissue may have significantly lower. I have tried to find some information for you regarding the expressed levels of leptin in sheep tissue, but unfortunately not too much data are available on the public sites compared to human, rat our mouse (i.e. Swiss-Prot, Unigen).
http://www.ncbi.nlm.nih.gov/UniGene/clust.cgi?ORG=Oar&CID=524
http://www.ncbi.nlm.nih.gov/UniGene/ESTProfileViewer.cgi?uglist=Rn.44444
The expression levels of leptin may vary from cell type to cell type or from tissue to tissue. It would be worth testing 1/500 or 1/1000 (for 1 hr at room temperature or overnight at 4oC).
I hope this will be useful for you. Should you still have any problem with this antibody after following these suggestions, then please do not hesitate to contact me again. I would be delighted to help you further.
Have a nice day!

Thank you for confirming the order dfetails.
Would you be so kind to clarify what samples you have been using:
- species,
- cell or tissue types,
- lysate (total cell lysates, or cytoplasmic fraction etc)
I look forward to hearing from you soon.