Immunofluorescence - (Oct/27/2005 )
certainly there are proteins that exist in one form in the cytosol and another in the nucleus...NF-kB subunits come to mind. it all depends on what your Ab is recognizing
Regarding your suggestion, this is how I find out about the presence of my protein in the cytosol. in an SDS PAGE I will detect it, right?
Thanks
Yes,it is called as western blot. First do SDS PAGE and then transfer it on PVDF membrane and then block the membrane with milk and add your antibodies ( similar steps like IF, with slight variation). Only detection method will be different. I think this will be best solution to resolve your antibody related problem (if thee is one).
Perhaps the protein associates with the cytosolic side of the nuclear envelope?
Hello,
I have a general question concerning immunofluorescence :
a) when I want to co-stain with Hoechst and an antibody, which should I do first?
when staining my cells with the primary antibody, shall I leave my cells at RT or at 4°C/ice?
a ) antibody first (its far more sensetive to being mucked up)
b ) room temps ok - i only use 4'C if its overnight
dom
Ok, thanks! 
But if I want to see cell surface proteins, doesn't ice make it easier? I mean doesn't ice prevent the antibody from going into the cell? 
not sure about that - you could reduce permeableisation i suppose
i'd let someone else answer that question
dom