wetern for membrane associated protein - (Nov/13/2007 )

I am trying to do the westerns for membrane bound protein from RAW cells and am unable to get it to work. It might be that I am loosing the protein with nuclear pellet after the lysis.

need help to sort this out? Do I need a different protocol which works for these proteins (maybe different lysis buffer recipe???)

any help will be greatly appreciated.

cheers
Rits

Membrane bound proteins require stronger detergents to separate them from the membranes. Try stronger detergents. You could also sonicate the nuclear pellet with a stronger pellet if you think it has the bulk of your protein.