Do I look at the most prominent bands on the figure? or do I look at bands at level 53kDa, even though they are not the most prominent in the image?
Would i also mention that multiple bands represent different p53 isoforms at that level?
Would appreciate the help :)
Also do i label it as molecular mass or Molecular weight if i am simply looking at a protein ladder of kDa?
Hi Carla,
If you want to check purified p53 protein, I mean pure protein or overexpressed protein, you can use SDS Page. But for p53 protein from cells, you can not simply determine which 53 kDa protein on your SDS gel exactly is p53, even you use a protein ledder. You can not be sure that 53 kDa protein band is really p53 protein, it could be another protein too.
Therefore, to detect p53 protein, which is isolated from cells, you should transfer your protein to Nitrocellulose or similar membran and perform Western blotting by using an antibody against p53 as a primary antibody. There are very good working monoclonal antibodies for p53.
Good luck
Cigdem
Multiple Band most often represent unspecific binding of the antibody (which is a known problem (for ref. see Baker M. Reproducibility crisis Blame it on the antibodies. Nature. 2015_521.p274-6. doi 10.1038521274a).
If the antibody is specific (which should be validated), the band corresponding to the expected MW should be used for interpretation of experimental findings (e.g. induction by stimuli, reduction via other processes etc.).
If your antibody is not isoform specific and all p53 protein isforms have the same MW in kD, you can not analyse protein expression using this tool.
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