Sperm Cryopreservation

Sperm cryopreservation is a technique used to freeze and store mature spermatozoa for later use. This technique has become a viable alternative to embryo cryopreservation, as it is more cost-effective and less labor-intensive. The process involves obtaining sperm from the epididymides and vas deferentia of a male transgenic or knock-out mouse and then freezing it in straws.

The investigator is responsible for providing a complete description of the mouse line to be cryopreserved, including its strain and genotype. Once the appropriate forms have been completed and online documentation of the mouse line has been provided, the sperm is cryopreserved and stored in liquid nitrogen storage tanks. A yearly storage fee applies to each line stored, unless the lines are donated to the Vanderbilt Cryopreserved Mouse Repository. These tanks are monitored weekly by personnel of the resource and cannot be accessed by other individuals.

Sperm cryopreservation requires two male mice that are proven breeders from the colony. The cryopreservation service includes the quality control check of thawing one or two straws of sperm and performing a small scale in vitro fertilization (IVF) to produce 2-cell stage embryos. Additional breeding steps may be required if more than one mutation is involved when re-deriving a line from cryopreserved sperm.

Timeline:

1. PI submits a service request iLab Organizer (corefacilities.org)
2. Service requests are reviewed and approved by the Director and Senior Program Manager of the facility
3. Experiments are scheduled
4. PI submits a transfer request through DAC for males to be moved on the specified cryopreservation date to Vanderbilt Genome Editing Resource procedure room.
5. Cryopreservation is performed and 21 straws are placed into long term storage
6. A small scale IVF is scheduled
7. One to two straws of sperm is utilized for IVF
8. Confirmation of fertilized embryos at the 2-cell stage
9. PI notified about results of IVF