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Theriogenology. 2018 Feb;107:41-49. doi: 10.1016/j.theriogenology.2017.10.036. Epub 2017 Oct 31.

Long term maintenance of frozen mouse spermatozoa at -80 °C.

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National Research Council (IBCN), CNR-Campus International Development (EMMA-INFRAFRONTIER-IMPC), Monterotondo Scalo, Rome, Italy.
Mary Lyon Centre, MRC Harwell Institute, Harwell Campus, Oxfordshire, OX11 0RD, United Kingdom.
Allevamenti Plaisant srl., Rome, Italy.
National Centre for Biotechnology (CNB-CSIC), Department of Molecular and Cellular Biology, Campus de Cantoblanco, Darwin 3, 28049 Madrid, Spain; CIBERER-ISCIII, Madrid, Spain.
Istituto Superiore Sanita', Roma, Italy.
National Research Council (IBCN), CNR-Campus International Development (EMMA-INFRAFRONTIER-IMPC), Monterotondo Scalo, Rome, Italy. Electronic address:


Maintaining mouse stocks as frozen materials offers both ethical and economical advantages over live animal breeding if the lines are not actively used. The European Mouse Mutant Archive (EMMA) promotes the archiving and distribution of important mouse models for biomedical research through the cryopreservation of their embryos and gametes. Embryo freezing in liquid nitrogen (LN2) at -196 °C has traditionally been the method of choice for archiving mouse lines. However, sperm freezing is emerging as a more convenient alternative due to the application of innovative cryopreservation and recovery protocols. In addition, frozen spermatozoa are less sensitive to post-freezing temperature fluctuations. We have previously reported that spermatozoa frozen using standard laboratory protocols can be safely stored in a -80 °C freezer or in dry ice (-79 °C) for at least seven days. We now report the extension of this period of maintenance at -80 °C up to two 2 years both for wild type and mutant strains, indicating that once frozen, mouse spermatozoa are quite resistant and can be transported, shared and stored at -80 °C for a long time without a significant loss of viability. The importance of this finding will be especially relevant for small laboratories with no constant access to liquid nitrogen and for labs generating many mouse mutant lines by CRISPR/Cas9 who do not want to saturate the limited space of a LN2 tank, using a more accessible -80 °C freezer.


-80 °C freezer; Biorepository; Cryopreservation; In vitro fertilisation (IVF); Laboratory mouse; Spermatozoa

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