[Histonet] RE: Low temp Antigen Retrieval for Bone Sections

[C.M. van der Loos]

   Dear Kim and others,

   We  have  been  testing  (not  for  bone  unfortunately) the overnight
   antigen  retrieval  at  60-70C  with  both citrate pH6.0 and Tris-EDTA
   pH9.0  and compared the staining results with the short boiling method
   (15  min  + 20 min cool-down). After overnight antigen retrieval using
   citrate6.0  the staining intensity was much less than with the boiling
   method.  However, after overnight antigen retrieval using Tris-EDTA9.0
   compared  with  the short boiling procedure the staining intensity was
   sometimes  similar,  sometimes less, sometimes even stronger. As ever,
   it varied  per  antigen. Interestingly, during the overnight procedure
   the  "fatty  tissue  parts" stayed much better on the glass than after
   the  short  boiling  procedure.  Nearly the same was described by Junn
   Chavez and Tim Morken in their NSH-poster.

   Overnight  antigen  retrieval  at  60-70C  is  to my opinion something
   worthwhile  to  investigate  further.  It's definitely interesting for
   those  working  with  tissues  that  tends  to fall off during antigen
   retrieval using the short boiling procedure.

   Chris van der Loos, PhD
   Dept. of Pathology
   Academic Medical Center M2-230
   Meibergdreef 9
   NL-1105 AZ Amsterdam
   The Netherlands

   phone:  +31 20 5665631
   fax:    +31 20 6960389



   Date: Mon, 26 Sep 2005 11:34:06 -0700 (PDT)
   From: Kim Merriam <kmerriam2003 <@t> yahoo.com>
   Subject: [Histonet] Low temp Antigen Retrieval for Bone Sections
   To: Histonet <histonet <@t> lists.utsouthwestern.edu>
   Hello All,
   I  will  be  performing  some  IHC  on  mouse  femurs  (about 12 or so
   different  antibodies).   In  the  past,  I  have had a lot of trouble
   keeping  these  sections  on  the  slides  during HIER.  I did a quick
   search on the archives and there were lots of suggestions, but nothing
   definitive.
   I  was reading an article about "low temp AR", overnight at 60C in TEG
   buffer,  pH  9.0.  Has anyone tried this method?  Also - what does TEG
   buffer stand for?
   Any help would be greatly appreciated.
   Kim
   Kim Merriam
   Novartis
   Cambridge, MA