[Histonet] Frozen section fixing problems - but paraffin works!

Wed Nov 19 17:09:08 CST 2003

MessageDear Tan
You have a lot of advaices in the wonderfull manual of Technical Immunohistochemistry of Rodney Miller, go to www.propathlab.com and download it free.Or go to http://hjaldanamarcos.bravepages.com/documentos.htm and donwload the documents named INMUNO1.... to INMUNO5.
By

Dr. Hernán J. Aldana Marcos
Facultad de Medicina. Universidad de Morón
Machado 914. B1708JPD. Buenos Aires. Argentina 
e-mail alternativo hernanjavier <@t> yahoo.com
web: http://hjaldanamarcos.bravepages.com
http://histologia.bigthicketdirectory.net/main.html

  ----- Original Message ----- 
  From: Tan, MinHan 
  To: histonet <@t> lists.utsouthwestern.edu 
  Sent: Wednesday, November 19, 2003 7:32 PM
  Subject: [Histonet] Frozen section fixing problems - but paraffin works!

  Hi,

  I am working with a new monoclonal antibody in parathyroid tissue, and I use the ABC-DAB reageant system for immunohistochemistry.

  I am new to frozen sections immunostaining - and I am encountering problems with frozen sections - staining is very weak or absent, in comparison to paraffin embedded tissue, which I have no problems with. 

  My protocol for frozen sections: 

  Unfixed frozen slides with 5 micron sections thawed at room temperature for 5 minutes
  Placed in 70% ethanol x 5 minutes.
  Washed in PBS x 5 minutes.

  ...

  followed with standard protocol: hydrogen peroxide 0.3% x 30 min, donkey serum 5% x 30 min, primary antibody (mouse) 4 deg overnight, secondary (goat anti-mouse), ABC, DAB.

  Does anyone have any advice?

  Thank you!

  Regards,
  Min-Han Tan 

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