The use of a liquid-phase controlled rate freezing system for the successful cryopreservation of mouse pre-embryos.
OBJECTIVE
To investigate the ability of a liquid-phase, controlled-rate freezing machine to generate reproducible freezing gradients with a constant rate of change; temperature fluctuations and heat dissipation during seeding; to assess the viability of mouse pre-embryos exposed to the silicone liquid cooling phase and the rates of survival and viability of mouse pre-embryos cryopreserved using this system.


METHODS
Freezing gradients were generated from the bath or a sample and compared for reproducibility and slope. Temperature fluctuations and gradients around the freezing chamber and the temperature rises and dissipation during seeding were measured. The toxicity of the silicone polymer freezing-phase was tested with mouse pronuclear pre-embryos. Different developmental stages of mouse pre-embryos were frozen and thawed and survival recorded in vitro and in vivo.


METHODS
Research Laboratories, Sinai Hospital of Baltimore, Baltimore, Maryland.


METHODS
The reproducibility of freezing gradients; temperature fluctuations during seeding; and the in vitro and in vivo viability of mouse pre-embryos exposed to the silicone polymer or frozen and thawed.


RESULTS
The freezing gradients generated were constant and reproducible with a constant rate of change; no temperature differences were recorded around the freezing chamber; temperature changes at seeding are minimal and rapidly dispersed. The silicone polymer was nontoxic to mouse pre-embryos and mouse pre-embryos frozen with the system and subsequently thawed are viable both in vitro and in vivo.


CONCLUSIONS
This liquid-phase cryopreservation system is an attractive option for assisted reproductive technologies laboratories because liquid nitrogen is not required for operation, it is reliable, there are small temperature fluctuations during seeding, and it can be kept constantly running.
Scott
L F
LF
Division of Reproductive Endocrinology, Sinai Hospital of Baltimore, Maryland 21215.
Sundaram
S G
SG
Smith
S
S
D000818:Animals; D015925:Cryopreservation / Q000295:instrumentation / Q000379:methods*; D004622:Embryo, Mammalian / Q000187:drug effects / Q000502:physiology; D005314:Embryonic and Fetal Development; D004864:Equipment and Supplies; D005069:Evaluation Studies as Topic; D051379:Mice / Q000196:embryology; D008815:Mice, Inbred Strains; D011108:Polymers / Q000494:pharmacology; D015203:Reproducibility of Results; D012828:Silicones / Q000494:pharmacology; D013696:Temperature
D011108:Polymers; D012828:Silicones