Human Reproduction, Vol. 19, No. 2, 300-305,
February 2004
© 2004 European Society of Human Reproduction and Embryology
Experimental vitrification of human compacted morulae and early blastocysts using fine diameter plastic micropipettes
1 Unidad de Reproduccion, Servicio de Ginecologia y Obstetricia, Hospital General Universitario de Alicante, Alicante, Spain, 2 Department of Medical Genetics, Faculty of Medicine, University of Porto, 3 Centre for Reproductive Genetics Alberto Barros, Porto and 4 Laboratory of Cell Biology, Institute of Biomedical Sciences Abel Salazar, University of Porto, Lg Prof Abel Salazar 2, 4099-003 Porto, Portugal
5 To whom correspondence should be addressed at: Laboratory of Cell Biology, Institute of Biomedical Sciences Abel Salazar, University of Porto, Lg Prof Abel Salazar 2, 4099-003 Porto, Portugal e-mail: msousa{at}icbas.up.pt
BACKGROUND: Vitrification of human blastocysts has been successfully applied using grids, straws and cryoloops. We assessed the survival rate of human compacted morulae and early blastocysts vitrified in pipette tips with a smaller inner diameter and solution volume than the previously described open pulled straw (OPS) method. METHODS: Excess day 5 human embryos (n = 63) were experimentally vitrified in vessels. Embryos were incubated at 37°C with sperm preparation medium (SPM) for 1 min, SPM + 7.5% ethylene glycol (EG)/dimethylsulphoxide (DMSO) for 3 min, and SPM + 16.5% EG + 16.5% DMSO + 0.67 mol/l sucrose for 25 s. They were then aspirated (0.5 µl) into a plastic micropipette tip (0.36 mm inner diameter), exposed to liquid nitrogen (LN2) vapour for 2 min before being placed into a pre-cooled cryotube, which was then closed and plunged into LN2. Embryos were warmed and diluted using 0.33 mol/l and 0.2 mol/l sucrose. RESULTS: The survival rate for compacted morulae was 73% (22/30) and 82% (27/33) for early blastocysts. CONCLUSIONS: The survival rates of human compacted morulae and early blastocysts after vitrification with this simple technique are similar to those reported in the literature achieved by slow cooling and other vitrification protocols.
Key words: blastocyst/human/morula/open pulled straws/vitrification
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