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Hello, we have purchased ab16123 and tested on human cells lines Calu-6 and MCF-10A at 20μg and 70 μg per well. We have detected a 40kDa band for all our western blots but could not see a band at 16kDa as predicted. We have already tried to optimize the results with your suggestions but we have not seen a difference. One of your antibodies against CDKN2A/p16INK4a antibody (ab54210) is giving a band at 40 kDa. On the scientific support page, you’re saying that : << Ab54210 recognizes isoform 4 of the protein, at ˜40KDa>>. Could it be possible for you to tell us where you did get that information? Do you have any literature reference that says the isoform 4 is giving a band at 40kDa? Could it be possible that the ab16123 also recognizes the isoform 4 of the protein?
Asked on Jul 16 2012
Thank you for taking time tocontact us.
For ab16123, have you tried running a positive control with HeLa cell lysate? It may be possible that your cell line is expressing a different isoform. In our testing, we do see the predominant band at 16kDa with HeLa cells, but we also see a faint band at 37 kDa which may be the isoform that is predominant in your samples. Or, you may want to do a cellular fractionation with your samples. It does seem that the different isoforms have different locations (cytoplasm, nucleus).
For ab54210, the 40 kDa band is what has been detected in the lab. I don't see that we have any specific references for this antibody in WB, but we do have some background references from the lab:
1. Hunter, T. 1993. Cell 75: 839-841.
2. Sherr, C.J. 1993. Cell 73: 1059-1065.
3. El-Deiry, W.S., et al. 1993. Cell 75: 817-825.
I hope this information helps. Please contact us with any other questions.
Answered on Jul 16 2012