Innovative technologies to support reproductive medicine: High-resolution synchrotron radiation nanotomography to explore cryopreserved oocytes ultrastructures

• Oocyte vitrification: a fertility preservation option in young cancer patients. • X-ray nanoimaging has been used for visualizing intact human cryopreserved oocytes. • The 3D approach enables to identify morphological markers of cold shock injuries. • This cutting-edge technology proved as a powerful tool to evaluate oocyte quality. Oocyte vitrification represents a tangible option for preserving fertility in patients of reproductive age undergoing oncological treatments. Evaluation of oocyte quality, especially the detection of potential ultrastructural damage induced by cryostorage and subsequent thawing, is crucial to safeguard successful in vitro fertilization. In this work, synchrotron radiation-based X-ray phase-contrast nanotomography (nanoCT) is used for the first time to assess the quality of intact human cryopreserved oocytes. This novel experimental pipeline has been implemented i) at room temperature following the oocytes’ cooling-warming procedure and ii) in cryogenic conditions simulating real-time oocyte freezing by in-situ cryo-nanoCT. Both three-dimensional (3D) explorations provide morphological insights of relevance for the overall gamete review, even allowing the identification of structural defects attributable to cryo-injuries. Here, X-ray nanoCT results are compared with images obtained by standard techniques routinely used to observe oocytes integrity, such as light microscopy and transmission electron microscopy (TEM). This proof-of-concept evidences that X-ray nanoimaging can effectively complement the well-established approaches thanks to its non-destructive nature and its capacity to deliver 3D information. Thus, our study introduces a novel methodological framework aiming to integrate cutting-edge imaging with cryotechnology in order to drive advances in assisted reproduction research.