大野 伸彦

The quick-freezing method often used for physical fixation also contributed enormously to the advancement of morphology, but it could not provide enough information about the dynamic morphological changes in vivo in living animal organs. Therefore, we developed the in vivo cryotechnique in 1995, which could directly cryofix organs in vivo under anesthetized conditions without stopping blood circulation or producing effects of anoxia. We have already reported new findings about the in vivo ultrastructures of living animal organs with the cryotechnique followed by freeze-substitution or replica preparation. Recently, it has also been applied to other morphological analyses in vivo, including immunohistochemistry and FISH, for obtaining dynamically functioning structures of cells and tissues at a light microscopic level. In such experimental processes, the in vivo cryotechnique has been shown to have the added benefit of direct antigen-retrieval effects since it reduces several steps of retrieval treatments which are required in common paraffin-embedded samples prepared by the conventional fixation and dehydration. In conclusion, the in vivo cryotechnique allows us to investigate the functioning real morphology of living animals, which was technically difficult to be observed before, and perform dynamic immunohistochemistry of cells and tissues in various animal organs in vivo at ultimate time-resolution.