Cryostorage and grafting of frozen-thawed (F/T) isolated ovarian follicles ovarian tissue fragments or whole ovaries are all possible therapeutic approaches now available to preserve fertility in young patients undergoing potentially gonadotoxic radio/chemotherapic treatments for cancer or other systemic pathologies. Our aim was to evaluate through a morphological approach the impact of enzymatic isolation cryopreservation and grafting on ovarian tissue integrity and viability. Human ovarian tissue was analyzed after the following treatments: 1- Follicle isolation by collagenase or Liberase enzymatic digestion; 2 - Xenotransplantation and orthotopic autotransplantation of F/T ovarian cortical strips; 3 - Cryopreservation of intact ovaries with their vascular pedicles. Observations were carried out by transmission electron microscopy vital fluorescent staining immunohistochemistry and DNA strand breaks analysis by TUNEL. In treatment 1 a higher proportion of follicles were viable after Liberase isolation in respect to those isolated with collagenase and most of Liberase-treated follicles were of good ultrastructural morphology. These data imply that the treatment with Liberase - a purified endotoxin-free enzyme blend - is a promising alternative to impure collagenase preparation for the isolation of intact ovarian follicles for culture and grafting purposes. In treatment 2 cryopreservation and transplantation do not appear to greatly affect the morphology of human primordial/primary follicles which even grow in the grafts; however follicular density was reduced after transplantation and follicular development appeared initially impaired. Thus further studies are needed to extend follicular life span and to improve follicular growth in the graft. In treatment 3 cryopreservation was not associated with any particular sign of apopotosis or ultrastructural alteration in all ovarian (follicular vascular and stromal) compartments. This result suggests that whole-organ sampling and transplantation may be a viable option in the future since vascular pedicle anastomosis allows immediate revascularization and greatly reduces ischemic injuries in the transplanted tissue. Funds were provided by Italian Ministry of Education University and Research (university grants) and by Italian Ministry of Foreign Affairs.