[Histonet] RE: permeabilization needed in IHC of viral protein?

[C.M. van der Loos]

   Hi Carla,

   Theoretically you don't need permeabilization. Since you have been
   cutting sections, the cell interior is fully exposed to the
   antibodies. On top of that you applied a protease step. My advise
   would be testing the staining under similar conditions without and
   with saponin 0.1% in all steps (including washing buffers). Again, to
   my opinion a different staining pattern is not very likely.

   Chris van der Loos, PhD

   Dept. of Pathology

   Academical Medical Center

   Amsterdam - The Netherlands


   ----- Original Message -----
                                                         From 
   Carla M Conway <cmconway <@t> usgs.gov>
                                                         Date 
   Wed, 02 Mar 2005 14:37:50 -0800
                                                           To 
   Histonet <@t> lists.utsouthwestern.edu
                                                      Subject 
   [Histonet] permeabilization needed in IHC of viral protein?

   Hello,
   I am interested in using IHC to detect nucleocapsid (N) protein within
   rhabdovirus. I have had success in detecting the glycoprotein on the
   envelope surface, however the N protein is located within the virion.
   Sections (FFPE) will be enzymatically digested with 0.05% protease XIV
   in
   TBS according to our standard protocol. Would a permeabilization agent
   increase  access  to  the interior proteins?  Any suggestions/comments
   will be
   greatly appreciated.
   Thank you,
   Carla Conway
   Western Fisheries Research Center
   6505 NE 65th St
   Seattle, WA 98115
   ph: 206-526-6282 ext. 242
   fax: 206-526-6654