Is optimal cutting temperature compound essential embedding solution treatment to cryo-sectioning of brain tissue?

We tested a set of conditions for obtaining optimal tissue quality in preparation for histology in samples of mouse brain. C57BL/6J mice were sacrificed and perfused with 4% paraformaldehyde, after which the brains were removed and dehydrated in 30% sucrose solution. The brains were then divided into four groups according to freezing temperature and usage of optimal cutting temperature (OCT) compound. Next, we stained the sectioned brain tissues with Harris hematoxylin and eosin Y and immunohistochemistry was performed for doublecortin. The best quality tissue was obtained at −25oC and by not embedding with the OCT compound. When frozen at −25oC, the embedded tissue was significantly damaged by crystals, while at −80oC there were no meaningful differences between qualities of embeddedand non-embedded tissues. Overall, we identified a set of conditions to obtain quality frozen brain sections. Our developed protocol will help resolve matters associated with damage caused to sectioned brain tissue by crystal formation during freezing.