[Histonet] morphology on frozen tissues

[Sun Zhon]

Dear All,
These were the steps I processed my frozen tissues,
after sectioning, air dry tissues for 30 mins
fix in acetone for 10 mins at RT
air dry for 30 mins
store in -80 freezer
 
The next morning before I did my IHC, I took out the fixed frozen tissues from the -80 freezer and put them directly in PBS without letting them come to room temperature.
 
My tissues looked like spider webs, especially around the edge. However, in another experiment I used exactly the same tissues and the tissues looked beautiful. The only difference was that while quenching the peroxidase, 0.03% H2O2 in PBS was used on the 1st exp (spider web morphology); 0.03% in 10% horse serum in 3% BSA was used on the 2nd exp (beautiful morphology).
 
Do you think the H2O2 will make such a big difference on the morphology or there are some other reasons for the spider web effect? How do you fix and store your frozen tissues? How do you treat the tissues before you do your IHC exps?
 
Thank you very much for your help.
 
Sun Zhon


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