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

[Tan, MinHan]

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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