I would like to perform immunofluorescence (IF) on testicular biopsy samples using frozen sections. Should I fix the tissues prior to sectioning, or do you recommend fixation after sectioning? What would you suggest as the optimal approach?
Dear Rumeysa
There may be no need for fixation. I never worked with testis tissue in frozen IF, but I do IF on frozen skin sections routinely. We don’t fix the 4 mm sections before IF-staining. Let me tell you how we do this:
The skin biopsies are frozen to -80°C
Then we bring the sample to the -25°C cryostat room. A large drip of OCT-medium is put on a metal carrier, the biopsy is put on top of the freezing OCT. A little while later this is transferred to the cutting position and we cut 4 mm sections that remain stretched on the knife. The slide for microscopy is put on top of the section and it is hence picked up. We let this dry for a short while.
We use l-lysine coated glass slides.
The next is to block for a-specific staining w/ 1% OVA in PBS
IF-staining without fixation
Good luck
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