I want to prepare cryosections of human heart muscle for immunohistochemistry: The tissue should be as native as possible (to allow antigen-antibody interaction without antigen retrieval) and tissue destruction (due to ice crystals during freezing) should be avoided. Does anyone have experience with that?
1. Quick freezing method: Inmerse the tissue in tissue-tek (OCT) and cover with aluminium paper. Put this in cold isopentane (dry ice) until the tissue tek turns white and cut.
2. Another method: put the tissue in 30% sucrose (in PBS). The tissue will be prepared to freeze when it sinks (a few hours or over night). Embbed in tissue-tek (OCT) and freeze.
Both methods work fine in fixed tissue, if you use unfixed tissue i would prefer the first method.
Usually the observation of ice crystals in cryostat section of tissue is due to a thawing-refreezing of the tissue during the procedure of cryosectioning not during the freezing. See the methodology described in the "Muscle Biopsy: A Practical Approach" Book writing by Pr V. Dubowitz: it shows the best results in cryoconservation during cryosectionning procedure.
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