Smad4 - double band on Western blot - (May/15/2007 )
Hi all,
Does anyone work with Smad proteins?
When running a western and probing with antiSmad4 (Santa cruz) I always receive 2 bands like here:
What is your opinion? Are those active and inactive form? Or maybe my primary antibody is crossreacting with another protein?
Thank you
I don't really work on smad4 but I do work on TGF-B stuff. I imagine it's obviously the correct MW for Smad4? When I run BMP-15 on a WB i also get a simmilar double bands. Apparently one of them is a glycosilated form of the BMP-15 mature protein (BMP-15 produced from transfected 293h). Perhaps your Smad4 is getting modified as well? Glycosilation? Phosphorylation? I do know that Smad4 binds to other Smads and perhaps what you're seeing is Smad4 bound to another Smad? But I could be wrong about the binding part because the size could then be considerably bigger.
Positive controls would possibly give you the best answer? I'm not sure but is it even possible to get purified Smad4?
I would go with "The erotic terrorist" and guess that your protein is post-traductionnaly modified so that its weight is slighlty increased.
Hi there,
i have a recent smad4 WB from SCBT antibody. i had a single defined band in my experiments from leukemic cells. all lanes are smad4 at different conditions.
i don't want to confuse you any further 
Does anyone work with Smad proteins?
When running a western and probing with antiSmad4 (Santa cruz) I always receive 2 bands like here:
What is your opinion? Are those active and inactive form? Or maybe my primary antibody is crossreacting with another protein?
Thank you