[Histonet] Antigen retrieval and rat skin

[Ronald P. Wilson]

Thank you for all the replies so far. As for additional details about our
procedure that some requested, the tissue is formalin-fixed and
paraffin-embedded. Sections are cut at 4 microns, mounted on Superfrost Plus
slides, air-dried and baked in a 55-60ºC oven for 1 hour before starting the
procedure. We use a Black and Decker rice steamer in which we place 3-4
plastic coplin jars containing citrate buffer (pH 10) into which the slides
are loaded back-to-back and standing upright. We steam for 20 min; the
steamer is started ahead of time so that the temperature reaches 96-98ºC.
The slides are then allowed to cool for 20 min before continuing with the
staining procedure. We recently tried some capillary gap slides and used the
reagent reservoirs in place of the coplin jars. There seemed to be a slight
improvement in the resulting slides but still had some disruption of the
dermis.

One reply recommended covering the tissue with about 1/2 ml of 10% formalin
and then placing in a 60ºC oven. Is this for frozen sections or
formalin-fixed paraffin-embed sections? I am willing to give it a try. We
believe that the problem is a result of the antigen retrieval with heating
since our standard H&E sections and BrdU sections which uses 20 min trypsin
digestion have very minimal wrinkling or disruption of the dermis.

Again, any suggestions or comments are much appreciated.

Ronald P. Wilson
Department of Comparative Medicine, HO54
Penn State University College of Medicine
M. S. Hershey Medical Center
500 University Drive
Hershey, PA 17033
phone:  717-531-8460
fax:       717-531-5001
e-mail:  rpw4 <@t> psu.edu