[Histonet] OCT vs. isopentane


Hello all,

I am quite new in the field of histochemistry. I'm doing mouse  
intestine cryostat section right now. I'm so confused about the method  
for cryostat section.

First, I knew that most of time people using OCT to embed their  
sample, but I saw some papers they use liquid nitrogen cooled  
isopentane to embed their sample. Do these two methods cause any  
difference? Because I saw some background flurorescence in the OCT  
embedded slide.

And if the sample embedded in isopentane, does the cryostat section  
take place in the same kind of machine? (with -20oC cryosection)

Second, for the 4% Paraformaldehyde following by sucrose-PBS for  
cryoprotection, does it is really important to wash out the previous  
medium before entering the next step until the embedding.

Third, does the thickness of the slide affect the autofluorescence?

I knew those questions might some kind of vague.
But thank you very much for any answers ahead.


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