Molina I.1*, Duque C.C.1, Alfonso J.2, Cervera R.P3 and Romeu A.1
1Human Assisted Reproduction Unit. University Hospital la Fe of Valencia, 46009, Valencia, Spain.
2Reproductive Medicine Institute (IMER), 46009, Valencia, Spain.
3Laboratory of Animal Reproduction and Biotechnology. Polytechnic University of Valencia, Spain.
*To whom correspondence should be addressed at: Human Assisted Reproduction Unit. University Hospital la Fe of Valencia, 46009, Valencia, Spain.
The present study was undertaken to compare the developmental capacity of human embryos derived from abnormally fertilised zygotes (1PN, ≥3 PN; 16-18 hours after ICSI) cryopreserved using two techniques: ultra rapid freezing and vitrification. At 2-4 cell stage, (48 hours after ICSI), these abnormally fertilised embryos were then distributed in three groups: a) embryos that were cryopreserved by ultra rapid freezing (URF Group), b) embryos cryopreserved by vitrification (V Group) and c) embryos that were not cryopreserved (Control group). Survival rates and embryo development after 24 hours of in vitro culture (72 hours after ICSI) were compared. 42 embryos were cryopreserved by ultra rapid freezing in 0.5mL straws, using a mixture of dimethyl sulphoxide (3M) and sucrose (0.25M) in a base solution consisting of IVF medium plus 20% (v/v) of Human Serum Albumin (HSA), and 24 embryos were vitrified in 0.25 ml straws, using a two step protocol with an equilibration solution consisting of 10% ethylene glycol (1.79M) and 10% dimethyl sulphoxide (1.41M) in a base solution of modified phosphate buffered saline (PBS) with 20% of HSA and a vitrification solution consisting of 20% ethylene glycol (3.58M), 20% dimethyl sulphoxide (2.82M) and 0.5M sucrose in base solution. The recovery rate after thawing/warming was lower for the vitrification group (75% V; 83% URF). The number of embryos with less than 50% of intact blastomeres after cryopreservation was significantly higher for the URF group (0% V; 34% URF). After in vitro culture, the rate of embryos not cryopreserved (Control group) that developed in vitro (72 hours after ICSI) was the highest (86%), followed by group V (50%), while group URF was the lowest (13%). These differences were statistically significant. This straw method of vitrification is successful and safe.
Keywords: cryopreservation, embryo, vitrification, freezing
Source: CryoLetters 27 (1), 59-64 (2006).