[Histonet] Frozen section fixing problems - but paraffin works!
Message
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=20frozen
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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