Immunofluores and cell permeability

[Richard Mandel]

We are trying to detect a protein on the cell surface while 
the protein is mostly found intracellularly.  We are fixing
with 4% paraformaldehyde but find that there is some 
permeabilization of the cells.  Has anyone found washing and fixing 
conditions that cause very minimal damage to the cell membrane?
Does glutaraldehyde or formaldehyde leave a more intact cell 
membrane than paraformaldehyde?  Should washing be done with
PBS or BSS?  The cells are Chinese hamster ovary cells attached
to a slide.  What is the minimum % of paraformaldehyde which can
be used in fixation and will this improve the situation?  

Thanks for your input.  E-mail or post here.

--

Ciao,
Rich 

---------------------------------------------------------------------------
Richard Mandel			|	E-mail address: rmandel at acs.bu.edu
Boston Univ. School of Medicine	|	Phone No. 617-638-4512	
Boston, MA 02118		|	Fax No. 617-638-4085
-----------------------------------------------------------------------------