Human Reproduction, Vol. 16, No. 9, 1965-1969,
September 2001
© 2001 European Society of Human Reproduction and Embryology
Cryoloop vitrification yields superior survival of Rhesus monkey blastocysts
1 Andrology/Embryology Laboratory, Departments of Obstetrics/Gynecology and 2 Physiology and Pharmacology, Oregon Health Sciences University, Portland, Oregon 97201, and 3 Division of Reproductive Sciences, Oregon Regional Primate Research Center, Beaverton, Oregon 97006, USA.
BACKGROUND: Vitrification using the cryoloop procedure was evaluated for preservation of non-human primate blastocysts by comparing survival results from two different cryoprotectant mixtures with prior results from controlled rate cooling. METHODS: Rhesus monkey blastocysts were produced by intracytoplasmic sperm injection of mature oocytes from cycling females stimulated with recombinant human hormones. Morphologically well-formed blastocysts were divided between Procedure A (2.8 mol/l dimethylsulphoxide and 3.6 mol/l ethylene glycol with 0.65 mol/l sucrose and 25 µmol/l Ficoll in TALP-HEPES with 20% fetal bovine serum (TH20)) and Procedure B (3.4 mol/l glycerol and 4.5 mol/l ethylene glycol in TH20). After >48 h in liquid nitrogen, the removal of cryoprotectants was accomplished in the presence of a 3-step series of decreasing sucrose concentrations in TH20. Surviving embryos were co-cultured on buffalo rat liver cells. RESULTS: Of 16 blastocysts vitrified via Procedure A, 38% survived with minimal lysis and only one hatched in culture; in contrast, of 33 blastocysts vitrified by Procedure B, 85% survived and 71% hatched. Of 22 blastocysts cryopreserved by conventional slow cooling, 36% survived and 6% hatched. Transfer into three recipients, each with two embryos vitrified with Procedure B, resulted in a successful twin-term pregnancy. CONCLUSION: Modified cryoloop vitrification with a final solution of 3.4 mol/l glycerol and 4.5 mol/l ethylene glycol is a promising procedure for preserving Rhesus monkey blastocysts that is simple, rapid, and inexpensive.
Key words: blastocysts/cryoloop/cryopreservation/monkey/vitrification
4 To whom correspondence should be addressed at: Oregon Primate Research Center, 505 NW 185th Avenue, Beaverton,OR 97006-3448, USA. Email: yeomanr{at}ohsu.edu
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