I am trying to sectioning fresh frozen mouse testis, but the H&E doesnt look good. I am enbedding in OCT and cutting in cryostat at 10 um. The temperature of the blade is -15C and the sample is -10. After that, I tryied to fix in bouins solution or formalin for 1h. But the morphology of the tissue doesnt look good.
Any tip?
Cryosectioning does not go well with H&E staining. You can try parraffin embeding.
I already have the samples frozen and is a long treatment so not possible to change it now.
Hi Paula,
Well.., It's always good to use paraffin sections for H&E staining. Since you have no choice I would say give a try.
Try to cut the cryosections at least to a thickness of 5-8um. thinner the section better the staining (paraffin sections are cut at 5um, so it has only one layer of cells most of the time which gives a clear morphology).
The other drawback of cryosectioning is tissue degradation. Try to avoid dramatic temperature fluctuations; i.e: keep the OTC embedded blocks at -20C. I think most of the cryostats works at -15-20C.
Fix the slide as soon as sectioned it. get a help from someone else to fix the slide while you section the blocks.
Do not freeze thaw the slides that you sectioned.
Cheers...!