Comparison of the effect of cryopreservation protocols on the histology of bioengineered tissues.

The purpose of this study was to compare the effects of five different cryopreservation protocols on the histology of bioengineered tissues. Although several artificial tissues have been developed to the date by tissue engineering, classical histological analysis methods and techniques must be optimized for these new tissues with special properties. The results of this study showed that the use of volatile solutions (formaldehyde, glutaraldehyde, glacial acetic acid and acetone) was not able to prevent the formation of large ice crystals that, in turn, can alter the structure of the artificial tissues. However, preincubation of the tissues in different concentrations of a carbon hydrate (glucose, maltose or trehalose) resulted in a better preservation of the tissue structure. We conclude that the best protocol that allows for an efficient analysis of the bioengineered tissues with very few artifacts is preincubation of the tissues in 0.300M or 0.400M trehalose for 30 or 120 min prior to OCT (optimal cutting temperature) embedding and cryosectioning. For all those reasons, we recommend the use of a cryoprotective agent before OCT embedding of human artificial tissues.