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Mouse ovarian follicle cryopreservation using vitrification or slow programmed cooling: Assessment of in vitro development, maturation, ultra‐structure and meiotic spindle organization
Author(s) -
Desai Nina,
AbdelHafez Faten,
Ali Mansour Y.,
Sayed Ezzat H.,
AbuAlhassan Ahmed M.,
Falcone Tomasso,
Goldfarb James
Publication year - 2011
Publication title -
journal of obstetrics and gynaecology research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.597
H-Index - 50
eISSN - 1447-0756
pISSN - 1341-8076
DOI - 10.1111/j.1447-0756.2010.01215.x
Subject(s) - vitrification , cryopreservation , antral follicle , andrology , meiosis , oocyte , in vitro maturation , follicle , ovarian follicle , biology , ovary , medicine , embryo , endocrinology , microbiology and biotechnology , biochemistry , gene
Aim: To compare different outcomes of vitrification and slow freezing of isolated pre‐antral follicles and to evaluate different cryo‐devices for vitrification of isolated follicles. Methods: Pre‐antral follicles were isolated from mouse ovaries and cryopreserved using vitrification and slow freezing. A preliminary experiment was carried out to select the optimal cryo‐device for vitrification of isolated follicles. A total of 414 follicles were randomly distributed among four groups: control (CT) fresh ( n = 100), nylon mesh ( n = 96), electron microscopy grid ( n = 102), and micro‐capillary tips ( n = 116). Subsequently, a total of 979 follicles were randomly assigned to three different groups: CT fresh ( n = 256), vitrification ( n = 399) and slow freezing ( n = 324). CT and cryopreserved/thawed follicles were cultured in vitro and examined daily for development. Final maturation was triggered with human chorionic gonadotrophin and rates of oocyte maturation were calculated. The ultra‐structure of cryopreserved/thawed follicles was studied using electron microscopy. Meiotic spindle presence and organization in mature oocytes were examined using the Oosight imaging system. Results: Micro‐capillary tips resulted in poor immediate post‐warming survival but no differences were observed in the subsequent in vitro development characteristics between different cryo‐devices. Nylon mesh proved to be the easiest carrier, particularly when large numbers of follicles were to be vitrified. Compared to vitrification, slow freezing resulted in a significantly lower number of intact follicles at the end of the culture period ( P < 0.0001). However all other outcome measures were comparable between both techniques. Conclusions: Isolated follicles were more vulnerable to cryodamage after slow freezing as compared to vitrification.