X-Message-Number: 9494
Date: Thu, 16 Apr 1998 00:37:18 -0700 (PDT)
From: Doug Skrecky <>
Subject: ethylene glycol cryopreservation of sheep embryos

Cryobiology 32: 78-91 1995
"In Vitro and In Vivo Survival of Cryopreserved Sheep Embryos"


   Experiments were conducted to devise an efficient method to cryopreserve
ovine embryos for field application. Embryos were surgically collected from
superovulated ewes on Day 6 after natural breeding: oocytes were collected
from ovaries obtained at the abattoir. Osmotic behaviour of oocytes and
embryos was determined by measuring their responces to hypertonic solutions
of CsCL or sucrose. Embryos and oocytes contracted osmotically by
decreasing their volumes proportionally to the reciprocal of the solution's
osmolality. The respective nonosmotic volumes of embryos in CsCl and
sucrose were 13.8 and 13.5% of their isotonic volume, and those of oocytes
were 18.5 and 19.6%. Tests of the permeability of morulae to commonly used
cryoprotectants, ethylene glycol (EG), proplene glycol (PG), dimethyl
sulfoxide (DMSO), and glycerol (Glyc), showed that the order of
permeability was EG>PG>DMSO=Glyc. Comparison of the efficacy of
cryoprotective agents indicated that the respective survivals of embryos
frozen in EG, PG, and DMSO were 76.9, 62.5 and 55.6%, based on their
development into hatched blastocysts in vitro. Therefore, EG appeared to be
superior to the other two cryoprotectants for freezing sheep embryos. To
determine the functional survival of embryos in vivo, 67 embryos frozen in
EG were thawed and directly diluted with phosphate-buffered saline; 47 of
these (70%) appeared morphologically normal and were transferred into 14
recipents. Five of these recipients, which had recieved a total of 16
embryos, became pregnant. Ten lambs were born, showing that the method
employed in this study for cryopreservation of sheep embryos followed by
their dilution out of EG has potential application for practical field use.

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