Anti-CBR1抗体(ab4148)

Thank you for your enquiry. We do not seem to have an exact match where the lysate has been extracted in RIPA buffer. My understanding is that the protein is cytoplasmic, therefore a lysate extracted in a gentler buffer should be ok. I would recommend looking at the following product: ab29889 https://www.abcam.com/index.html?datasheet=29889 I hope this information helps. Please do not hesitate to contact us if you need any more advice or information.

Thank you for your enquiry. I am sorry to hear that you are experiencing difficulties with this product ab4148 in western blot. Often it is possible to make suggestions that help resolve problems. We will happily offer technical support and in the event that a product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 120 days of purchase), and if it appears that the antibody is at fault, a replacement/credit note/refund will be offered. I have looked through the protocols you used and have a few questions and suggestions that might help you resolve the problem. If the suggestions do not prove to be helpful, would you please be so kind to confirm the following items in order to help me better understand the cause of the problem. In most cases, the cause of high background is because the primary and/or secondary antibody concentration is too high and causes non-specific binding. The dilution that we have on the datasheet is a recommended starting dilution and customers are encouraged to determine the optimal concentration/dilution. You can try decreasing the primary (1:2000 ~ 1:5000) and secondary antibody concentration. Another possible cause is that the amount of protein is too much. We normally recommend using 20-40 micrograms per well but please try 20ug if you have not already done so. Increasing the washing frequency and duration would also help eliminate high background. Can you confirm that you have used Tween in your washing solution and have washed the primary antibody at least 15min x 3 times? Please try this if you have not already done so. Can you confirm you have reduced and denatured the sample in buffer containing SDS and mercaptoethanol? This will ensure the protein is in the correct conformation to run at the correct molecular weight and be detected by the antibody. I am sorry for the string of questions but I hope the above recommendations may already help you. If you have already tried the above suggestions and still experience problems, please do not hesitate to contact me with the answers to the above. Also, please advice me on how you would like to proceed with your enquiry, so that I can immediately arrange for a replacement or refund to you.

Thank you for your enquiry. I am sorry to hear that you are experiencing difficulties with this product ab4148 in western blot. Often it is possible to make suggestions that help resolve problems. We will happily offer technical support and in the event that a product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 120 days of purchase), and if it appears that the antibody is at fault, a replacement/credit note/refund will be offered. I have looked through the protocols you used and have a few questions and suggestions that might help you resolve the problem. In most cases, the cause of multiple bands is because the amount of protein is too much. We normally recommend using 20-40 micrograms per well. Loading more than this may overload the gel, which will result in many non-specific bands. Please try this amount if you have not already done so. Another possible cause is that the primary and/or secondary antibody concentration is too high and causes non-specific binding. The dilution that we have on the datasheet is a recommended starting dilution and customers are encouraged to determine the optimal concentration/dilution. You can try decreasing the primary (1:2000 ~ 1:5000) and secondary antibody concentration or run a no-primary control (without the primary antibody). Running a no-primary control will be able to eliminate the possibility that your secondary antibody is binding non-specifically. Regarding the bands that you obtained, the bands located above the expected molecular weight band could be multimers. Please try boiling your protein in SDS-PAGE for 10 minutes rather than 5 minutes to ensure proper disruption of multimers. This will ensure the protein is in the correct conformation to run at the correct molecular weight and be detected by the antibody. Meanwhile, bands located below the expected molecular weight may indicate that your target protein has been digested. Please make sure that you incorporated sufficient protease inhibitors and have kept your samples on ice the whole time. Increasing the washing frequency and duration would also help eliminate high background. Can you confirm that you have used Tween in your washing solution and have washed at least 15min x 3 times? I hope the above recommendations may already help you. If you still experience problems please do not hesitate to contact me with the answers to the above questions. Also, please advice on how you would to proceed with this enquiry so that I can immediately arrange for a replacement or refund to you.

Yes I would recommend your using ab6740 (Rabbit polyclonal to Goat IgG H&L (biotin)) to detect ab4148. This secondary antibody works very well in IHC - please see the two customer reviews accessible via the online datasheet. As you have seen, ab4148 has only been tested in WB and we do not yet know if it will work in IHC. Should you go ahead and test this antibody, please let us know how you get on.