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As I told you in my previous mail, I repeated the immunostaining following your suggestion of testing different blocking conditions and changing the antibody diluent to a less concentrated BSA solution.
Unfortunately this only increased my background but the quality of the staining did not increase. It is still not possible to detect RAD51 foci with the ab1837 that we bought. Our secondary antibody is not contributing either to the background staining.
I think it is of interest for you to know that we tested also ab133534 (rabbit monoclonal) on our preparations in the usual conditions (the ones I described in my first enquiry): this antibody works quite well (1:250) although it gives a little bit too much background compared to our homemade rabbit polyclonal.
Taking all this together, I would guess that there is really something wrong with the ab1837 that we received.
You mentioned in your previous reply that you could provide a replacement in case the antibody would still not work. If that is still an option, we would like to proceed that way, also because there are published data showing a quite good staining of RAD51 with ab1837 in testis spread preparations.
I am looking forward to hearing from you soon
Asked on Nov 20 2012
Thank you for your reply and for letting me know how you have been getting on.
I am sorry that my suggestions have not improved the results observed. As you say, especially with the positive results with the rabbit monoclonal ab133534, it may be that the vial you received may have lost its activity. I am sorry for the inconvenience this has caused you.
If you would like I can provide you with a free vial of ab1837 however, we only have the same lot you have been trying in stock. Alternatively I can provide you with a different antibody altogether such as the chicken polyclonal ab63802 or the rabbit polyclonal ab137323. Or if you would prefer, I could provide you with an alternative antibody against any target of your choice (as you already have ab133534 which seems to be working for you) or a credit note or refund.
In order to arrange for any of these I would need confirmation of the order number on which ab1837 was received. Could you provide this, or if you do not have this information, the approximate delivery date and the delivery address used?
Many thanks for your cooperation. I look forward to hearing how you would like to proceed.
Answered on Nov 20 2012