An improved vitrification protocol for the fast and safe storage of mouse oocytes More about Open Access at the Crick
Authors list
Tina Hodgson Hollie Lane Helen Horsler Juliette Horwood Laura Denti Fabio Delaqua Katharine Crawley Ben DaviesAbstract
Cryopreservation methods for archiving and distributing mouse strains mostly focus on freezing embryos or sperm. Although protocols for the cryopreservation of wild-type mouse oocytes are available, these methods have not been widely adopted in large biomedical research facilities. This is partly due to a lack of validation of the available methods on a large scale using a range of genetically modified oocytes. Furthermore, some of the existing methods report a relatively low rate of fertilisation, requiring either the zona pellucida to be physically breached or the inclusion of cumulus cells to enable efficient fertilisation, interventions which might be incompatible with maintaining hygiene barriers. Existing methods also often use cryovials rather than straws or slimline vitrification devices, which are more practical for storage and handling. Here, we present a robust vitrification protocol for large-scale oocyte cryopreservation, achieving high viability and fertilisation rates comparable to fresh oocytes. We have extensively tested the protocol for in vitro fertilisation of many genetically altered strains, using both genetically altered and wild-type C57BL/6 J oocytes and sperm. Providing an archive of cryopreserved oocytes harbouring genetically modified alleles separately from archives of cryopreserved sperm allows multiple allele combinations to be generated during the rederivation process. This reduces subsequent breeding steps and the need to maintain live stocks of mice. Furthermore, cryopreserving oocytes for later use enables them to be obtained from females at the optimal age, thereby reducing the number of mice required and providing greater scheduling flexibility for subsequent genetic modification and rederivation work.
Journal details
Journal
Biology of Reproduction
Volume
113
Issue number
6
Pages
1355-1363
Available online
Publication date
Full text links
Publisher website (DOI)
10.1093/biolre/ioaf215
Figshare
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Europe PubMed Central
40981676
Pubmed
40981676
