Our service entails flushing preimplantation embryos, selecting 225 healthy embryos and freezing them in groups of 25. We use a stepwise slow freezing protocol that maximizes viability.
How many mice are expected from each group of 25 embryos?
We expect that each group of 25 embryos will yield 10 live born mice. Thus, 200 cryopreserved embryos should provide more than sufficient backup for a strain or mutant. However, if cryopreserved embryos will be the only source of this strain or mutant in the world, we recommend freezing two or three times as many.
How many mice are required to get 225 healthy embryos?
Ten to twenty-five plugged, superovulated females are required, depending on the genetic background. A robust, highly fertile outbred strain like CD1 might be frozen in one session from 12 females and six stud males. Typical inbred strains might be cryopreserved in two sessions, mating two sets of 12 superovulated females to 6 stud males. Fewer mice and reduced fecundity will require a larger number of freeze sessions.
The stud males must be older than 2 months of age, and less than a year old. Males should be housed individually for two weeks prior to mating. If it has been some time since they have mated or if it is their first time, put a female in their cage 2 weeks before mating for cryopreservation, and then remove the female one week later.
Females should be 6 to 10 weeks old. We will provede hormones to the investigator to superovulate the females.
Superovulation is the forcing of ovulation through timed injection of hormones, allowing us to synchronize and schedule matings. Superovulation also can lead to greater numbers of eggs in some strains.
What level of confidence can be placed in the frozen samples?
We chose the parameters that we did with the belief that the typical need of investigators would be for backup of strains or mutants that are available elsewhere. If your intention is to cryopreserve as the sole source of a mutant or strain, we recommend that you cryopreserve twice and split the frozen samples between two liquid nitrogen freezers at two sites.