Anti-Estrogen Receptor beta antibody (ab3576)

Thank you for your enquiry and interest in our products.

I can confirm that ab3576: Anti-Estrogen Receptor beta has been used in different dilutions in IHC (P).

There are two protocols which were used the same buffer but slightly different temperature and both were successfully applied with this antibody:

A) Paraffin-embedded sections were dewaxed and rehydrated through graded alcohol. Heat-mediated antigen retrieval was performed at 100 C in 1 liter 0.01 M citrate buffer (pH 6.0). Concentration of antibody used: 5 µg/ml.

B) After washing, the sections were transferred to plastic jars containing 0.01 M citrate buffer (pH 6.0) and boiled in a microwave oven for 10 min at 700 W. Samples were allowed to cool for 20 min before washing in PBS. 1:500 dilution in PBS + 2% BSA.

If you need any further assistance in the future, please do not hesitate to contact me.

Thank you for your participation in Abcam's post complaint survey.

Please let me know if there is anything else I can do for you, or if you have questions to any of our products (e.g. primary antibodies, lysates, secondary antibodies, kits, controls, gels, proteins/peptides, slides, biochemicals, conjugation kits).

Thank you for taking the time to 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 ab3576 is tested and covered by our 6 month guarantee for use in WB and human, mouse and rat 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 ab3576.
1.) Antibodies will produce high background if they are used too concentrated. I suggest to further dilute the antibody.
2.) background is often generated by the secondary antibody and therefore I can recommend to rum a "no primary" control.
3.) Cell lines often produce artifact due to prolonged culturing. We have reports from customers using this antibody in tissue samples with good results. I suggest to run a positive control with a suitable tissue lysate.
4.) Since Estrogen Receptor beta is a nuclear protein, I can suggest to prepare nuclear lysates to clean up the blot.
I would also appreciate if you can confirm some further details:
1.) What lysis buffer was used?
2.) There are 8 isoforms of this protein. They range from 35kDa to 59kDa. Do you which one are expressed in the cell lines used?
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.

Thank you very much for you reply.

I'm sending a free of charge vial of ab3576 on the order ***, which we expect to arrive around August 24. Your purchasing agent will receive an email update if there are further delays to expect.

Please keep me updated about the results with this new antibody, and let me know if there is anything else that we can do for you. I look forward to hearing from you.

Have a nice day!