Vitrification of zona‐free rabbit expanded or hatching blastocysts: a possible model for human blastocysts

• Published in: Human reproduction (Oxford) Vol. 18; no. 10; pp. 2151 - 2156
• Main Authors: Cervera, R.P., Garcia‐Ximénez, F.
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.10.2003; Oxford Publishing Limited (England)
• Subjects: Animals; Blastocyst - physiology; Cryopreservation - methods; Embryonic and Fetal Development; Female; Histological Techniques; Hot Temperature; Humans; Key words: blastocyst size/in‐vitro implantation/rabbit/standard sealed straws/vitrification; Models, Animal; Rabbits; Tissue Survival; Zona Pellucida; blastocyst size/in‐vitro implantation/rabbit/standard sealed straws/vitrification
• ISSN: 0268-1161; 1460-2350; 1460-2350
• DOI: 10.1093/humrep/deg428

BACKGROUND: The purpose of this study was to test the effectiveness of one two‐step (A) and two one‐step (B1 and B2) vitrification procedures on denuded expanded or hatching rabbit blastocysts held in standard sealed plastic straws as a possible model for human blastocysts. The effect of blastocyst size was also studied on the basis of three size categories (I: diameter <200 µm; II: diameter 200–299 µm; III: diameter ≥300 µm). METHODS: Rabbit expanded or hatching blastocysts were vitrified at day 4 or 5. Before vitrification, the zona pellucida was removed using acidic phosphate buffered saline. For the two‐step procedure, prior to vitrification, blastocysts were pre‐ equilibrated in a solution containing 10% dimethyl sulphoxide (DMSO) and 10% ethylene glycol (EG) for 1 min. Different final vitrification solutions were compared: 20% DMSO and 20% EG with (A and B1) or without (B2) 0.5 mol/l sucrose. RESULTS: Of 198 vitrified blastocysts, 181 (91%) survived, regardless of the vitrification procedure applied. Vitrification procedure A showed significantly higher re‐expansion (88%), attachment (86%) and trophectoderm outgrowth (80%) rates than the two one‐step vitrification procedures, B1 and B2 (46 and 21%, 20 and 33%, and 18 and 23%, respectively). After warming, blastocysts of greater size (II and III) showed significantly higher attachment (54 and 64%) and trophectoderm outgrowth (44 and 58%) rates than smaller blastocysts (I, attachment: 29%; trophectoderm outgrowth: 25%). CONCLUSIONS: These result demonstrate that denuded expanded or hatching rabbit blastocysts of greater size can be satisfactorily vitrified by use of a two‐step procedure. The similarity of vitrification solutions used in humans could make it feasible to test such a procedure on human denuded blastocysts of different sizes.