IF showed predominant localization of this protein in nucleus (z-stack confirmed), however, when I performed nuclear fractionation and westernblot, my protein was barely detected in the nucleus but most in cytoplasmic fraction(and all fractionation controls were good; GAPDH only detected in the cytoplasm and sp1 only in the nucleus,loading control shows equal loading).
I don't understand how come with such a strong nuclear signal by IF, but I cannot detect it by western blot. Which result is more convincing? I am frustrated with those results and I will appreciate if any of you can give me an explaination.
Edited by tianna, 28 November 2010 - 09:46 AM.