I'm working on senescence, and I'm trying to observe senescence markers. I have performed western blots for p21 but it seems that the bands are at a much higher molecular weight than expected. The expected molecular weight is aprox. 18KDa but I see bands around 50KDa. Has it ever happened to anyone? why does it happen? (I am using a monoclonal antibody 1:2000 from proteintech 67362-1-Ig). thanks :)
Hello, Camilla!
What is the gel concentration you are working with? Perhaps it is too concentrated... I' ve had some trouble like yours and we have found the problem was with acrylamide that it was too concentrated in the stock solution. So, every time we did the gel, it was too concentrated and bands were appearing in the wrong molecular weight.
I have had non specific bands in western blots before. You can try to contact the vendor that makes the antibody and ask if they have mapped the epitope on p21. Looking at their website, they only show immunofluorescent analysis. It may work better for that application more than for western blots. Maybe try a different antibody where the vendor shows an actual western blot in their catalog.
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