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

[Tan, MinHan]

Sorry, I neglected to mention - 
 
I have also tried 
 
Sections were cut in cryostat and stored as unfixed slides overnight in
-80. (no dessication though)
Unfixed frozen slides (stored at -80) placed at -20 for 5 minutes
Placed in -20 acetone for 5 minutes.
Air dried at room temperature for 10 minutes

As per standard protocol.
 
 

	-----Original Message-----
	From: Tan, MinHan 
	Sent: Wednesday, November 19, 2003 5:33 PM
	To: histonet <@t> lists.utsouthwestern.edu
	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 
	 
	 
	 
	 
	 
	 
	
  _____  

	
	This email message, including any attachments, is for the sole
use of the intended recipient(s) and may contain confidential
information.  Any unauthorized review, use, disclosure or distribution
is prohibited.  If you are not the intended recipient(s) please contact
the sender by reply email and destroy all copies of the original
message.  Thank you.
  _____  



This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential information.  Any unauthorized review, use, disclosure or distribution is prohibited.  If you are not the intended recipient(s) please contact the sender by reply email and destroy all copies of the original message.  Thank you.
-------------- next part --------------
An HTML attachment was scrubbed...
URL: http://lists.utsouthwestern.edu/pipermail/histonet/attachments/20031119/c8bdf1f4/attachment.htm