UCI > Research > TMF > Freezing/Thawing of Embryos

Embryo Cryopreservation and Embryo Thawing Services

• Introduction
• Ordering
• Turnaround Times
• Performance Guarantees
• Service Description

Introduction:

Mouse strains that are no longer needed for experimental purposes are most economically stored as frozen embryos or sperm at liquid nitrogen temperature. Embryos can be cryopreserved at all stages of pre-implantation development except blastocysts. Current methods of cryopreservation do not work with unfertilized oocytes.

Traditionally, the most common method of cryopreservation has used multi-cell, pre-blastocyst stage embryos, infused with a cryoprotectant such as glycerol, slowly lowered to –35C, and then rapidly submerged in, and stored in, liquid nitrogen. Methods are also available for freezing fertilized eggs.

In terms of recovery of live mice, the efficiency of every method of cryopreservation is strain-dependent. Efficiencies have traditionally been much more strain-dependent for sperm cryopreservation than for embryos, although a new technique recently announced by the Jackson Laboratory appears to make sperm cryopreservation almost as reliable as for embryos.

One advantage of cryopreserving embryos rather than sperm is that the diploid genome is preserved, meaning genotypes such as homozygous knockouts can be reconstituted directly, rather than requiring further breeding.

Currently we do not offer sperm cryopreservation as a routine service. However, we can reconstitute mice from frozen sperm obtained from outside sources. Reconstitution from frozen sperm requires our in vitro fertilization service and a source of oocyte donors.

Ordering:

To order our embryo cryopreservation or embryo thawing services, go to our on-line order page. After filling out the on-line Service Request Form, print a hard copy, have it signed by the Principle Investigator, and mail or fax it to Tom Fielder.

In addition to the Service Request Form, the client must provide the embryo donors and stud males. We recommend providing at least 6-8 donors per strain, depending on how many embryos the client wishes to freeze and assuming an average response to superovulation and normal fertility.

If enough stud males are available, it may be to the client’s advantage to order wildtype females of the same background for use as embryo donors. This is a convenient way of ensuring all females are of the optimum age for superovulation. Superovulation is generally most efficient using females that are either 3-4 weeks of age, or 8-10 weeks of age. Females older than 10 weeks can be used, but the embryo yield does diminish gradually with age.

Any number of strains may be cryopreserved in a single session. However, the maximum number of embryo donors per session is 30, and each donor should be mated with a separate male. A flat fee is charged for each cryopreservation session, regardless of how many donors are supplied.

Normally, the embryo donors will be superovulated and mated by TMF staff in the client’s mouse room. Therefore, we prefer to have someone from the client’s lab present during the superovulation process to help identify the mice.

Turnaround Times:

The cryopreservation process usually requires about a week, once the embryo donors and stud males are designated by the client. Thawing embryos stored by the TMF requires about 6-7 weeks from the day of thawing to produce weaned pups ready for transfer to the client. If the frozen embryos are from an outside institution, the foster mothers will have to be tested for pathogens before the weaned pups can be released, adding 1-2 weeks to the turnaround time.

Performance Guarantees:

The number of embryos cryopreserved depends on the number of embryo donors supplied by the client, their age and response to superovulation, and the fertility of the stud males. Therefore, we cannot offer any guarantees as to how many embryos will be frozen in any given session.

Generally about 20-40% of thawed embryos develop into live pups, reflecting strain variation in embryo survival and developmental potential.

Service Description:

Our embryo cryopreservation service includes

• superovulation and mating of embryo donors
• harvesting fertilized embryos from donors
• freezing embryos
• long-term storage in liquid nitrogen

Our embryo thawing service includes

• generating pseudopregnant foster mothers
• Transferring thawed embryos to pseudopregnant foster mothers
• Monitoring pregnancies
• Weaning pups

 

University of California, Irvine
Transgenic Mouse Facility
B123 Gillespie Bldg. • Irvine, CA 92697-1140
© 2008 The Regents of the University of California, All Rights Reserved.
Last Updated: 02.08.2007

Comments & Questions: Privacy & Legal Notice
Copyright Inquiries