Okay so after searching through my notes here it goes: I have tested over 20 ERalpha antibodies and the best one by far (in my hands) no matter which company used is that produced by the sp1 clone but you can pick and choose companies. I usually use Abcam though the one from ThermoFisher was also excellent. For Progesterone Receptor I use the C89F7 Rabbit mAb clone #3153. I have tried several others and they'll work for cells that are very high in PR Receptor like T47D but when you want to detect it in MCF7 which are fairly low on it (50X less than T47D), this antibody was the best by far. The transfer buffer I use is 20mM sodium phosphate, pH 6.8, 20% MeOH, and 0.05% SDS. Since there's SDS in the buffer, the transfer is slower than what we are used to nowadays and it is performed at 0.4 Amp for 4-5 hrs (though shorter times are also possible depending on your transfer apparatus size etc so you'll have to titrate a teensy bit); temperature is of the essence so I normally put my transfer apparatus in the cold room and I place it inside an ice bath to ensure that the temperature stays constant. You can find a detailed version of this protocol on Estes et al. 1987. I do not follow the rest of the protocol as described and instead block in milk/tbst and incubate the primary in 5% bsa/tbst but it'll at least give you a good sense of why the phosphate buffer is used and why it tends to work so much better than towbin buffer for this protein. I have tried the clone described in this article and it gave me some non-specific bands however the one I've pointed you to here is perfectly clean. I hope your steroid receptor woes are over soon. Best of luck!

They both are good companies. The tissues are from human or rodent, Just make sure to by the right kind of Antibodies usually they all work well. Even if I share my experience, it may not work well with you, so make a choice and it will work.

The best progesterone receptor antibody hails from cell signaling. It is an anti rabbit antibody; I will look up the clone in a little while but please be aware that because the isoelectric point of the progesterone receptor is about the same as that of the regular tris glycine buffer used for transfer you may have issues detecting it. For these purposes, a phosphate based transfer buffer works best. I will paste my protocol shortly. For estrogen receptor, the best antibody is the one generate by the sp1 clone and I have always purchased abcam's but I believe that this clone is also available from cell signaling or samara cruz. Will get back to you later on the specifics.

Hi Mariana,

You should be able to get antibodies from Abcam through http://www.biogenbr.com.br/

We also do anti-Estrogen receptor and anti-Progesterone antibodies (both of which have over 11 references):

http://www.abcam.com/Estrogen-Receptor-alpha-antibody-33-ab2746-references.html

http://www.abcam.com/progesterone-receptor-antibody-pr-at-414-chip-grade-ab2764.html

Hi Mariana,

we usually use the AER 311 mouse monoclonal anti-ERalpha antibody (provided by Dr Ciro Abbondanza at Department of General Pathology - II University of Naples). For ER detection in WB, we use the HC-20 anti-ERalpha antibody (Santa-Cruz).

I think, however, that the AER 311 MAb is now commercially available.

In ChIp analysis we used the anti-ERalpha antibody (154-171, from Sigma) .

We usually detect PgR using the mouse monoclonal anti-PgR antibody (clone 6A1 from Cell Signaling).

Good luck with your experiments!

The ER alpha antibody I have been using for a long time is from Santa Cruz, Cat.# SC-8002. It gives me very clean bands on western blot. But I also think there is a lot of good ERalpha antibody available in different companies because people have been doing research on this protein for so long.

Okay so after searching through my notes here it goes: I have tested over 20 ERalpha antibodies and the best one by far (in my hands) no matter which company used is that produced by the sp1 clone but you can pick and choose companies. I usually use Abcam though the one from ThermoFisher was also excellent. For Progesterone Receptor I use the C89F7 Rabbit mAb clone #3153. I have tried several others and they'll work for cells that are very high in PR Receptor like T47D but when you want to detect it in MCF7 which are fairly low on it (50X less than T47D), this antibody was the best by far. The transfer buffer I use is 20mM sodium phosphate, pH 6.8, 20% MeOH, and 0.05% SDS. Since there's SDS in the buffer, the transfer is slower than what we are used to nowadays and it is performed at 0.4 Amp for 4-5 hrs (though shorter times are also possible depending on your transfer apparatus size etc so you'll have to titrate a teensy bit); temperature is of the essence so I normally put my transfer apparatus in the cold room and I place it inside an ice bath to ensure that the temperature stays constant. You can find a detailed version of this protocol on Estes et al. 1987. I do not follow the rest of the protocol as described and instead block in milk/tbst and incubate the primary in 5% bsa/tbst but it'll at least give you a good sense of why the phosphate buffer is used and why it tends to work so much better than towbin buffer for this protein. I have tried the clone described in this article and it gave me some non-specific bands however the one I've pointed you to here is perfectly clean. I hope your steroid receptor woes are over soon. Best of luck!

We have used many antibodies from Cell Signaling- all superb". Santa Cruz more hit and miss.

HC-20 from Santa Cruz will detect the 66 KDa form of ER -alpha from MCF-7 cells by WB . I use it like every other week. There aren't many good ones that detects the ER beta form. Santa cruz will issue credit if the antibody does not work.

I have used ER-alpha from Cell Signaling (#8644) and it works quite well.

If I need to perform both IHC and western blot for ER-alfa from MCF7 cell line/ tumor, does anyone have good recommendation for a good antibody that targets both experiment? (HC-20 or Cell signaling?)

Thanks for your help

ER alpha MC-20 from santa cruz fails to detect the protein band in endometrial cells. while ER beta is well detected. can anyone suggest why?