Sperm Cryopreservation

Sperm cryopreservation involves the isolation of mature sperm for the caudal epididymis followed by the cooling, then rapid freezing of sperm in liquid nitrogen. Sperm can be thawed at a future date and incubated with oocytes to generate embryos that are then surgically transferred into recipient females to recover the line.

Sperm cryopreservation is a fast and inexpensive method for preserving germplasm as compared to preserving embryos. Recent modifications to the sperm cryopreservation procedure have vastly improved the quality of thawed sperm, particularly for C57BL/6 and Balb/c strains. Even with these improvements, the fertilization capasity of thawed sperm can vary significantly amoung mouse lines. However, a substantial amount of material (ie, ~30 x 10⁶ sperm/male) can easily and quickly be isolated and preserved. Sperm from either homozygous or hetero/hemi-zygous males can be cryopreserved. Bear in mind that when cryopreserving sperm, of course, only half the genome is preserved.

Project Initiation

To initiate an sperm cryopreservation project, completed a Sperm Cryopreservation Request Form and submit it to TTML@northwestern.edu.

• The male should be between 3-5 months of age and singly housed for two weeks for optimal sperm production.
• TTML staff will contact you ~2 weeks prior to beginning your project to confirm the schedule date and have you separate a male.
• Typically, sperm from two males of each line is pooled and frozen into 20 straws.

Quality control:

• Test straws from each freeze are thawed, incubated with oocytes to generate embryos (using IVF). The viability and developmental capacity of the thawed sperm is assessed by culturing embryos in vitro to the blastocyst stage.

• If the test thaw is inconclusive, another test straw will be thawed and IVF generated embryos will be transferred into recipient female to confirm development capacity of the thawed sperm.

• Depending on these results, we will contact you to either schedule another session or confirm that the project is complete.

Expected results:

• A minimum of 17-19 straws are frozen for each line.
• A test straw from each freeze is thawed to determine the success of the procedure.

Long-term storage and access to cryopreserved embryos:

• Cryopreserved sperm from each line is stored in two separate liquid nitrogen dewars in two different buildings.
• Currently, there is no additional cost for storage.
• Sperm can be cryorecovered at any time simply by contacting our lab.
• Sperm can also be shipped world-wide from our lab.