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The use of a chemically defined medium, of non-biological origin, to freeze embryos
THERIOGENOLOGY
THE
USE
R.J.
OF
A CHEPllCALLY ORIGIN.
DEFINED HEDIUll. OF TO FREEZE EI’IBRYOS
NON-BIOLOGICAL
Mapletoft, B. Bavisterl. J.S. Moker and W.C. Dept. Herd Medicine and Theriogenology WCVM University of Saskatchewan Saskatoon, Sask. SlN OWO CANADA IDept of Vet Sci , University of Wisconsin Madison, WI USA 53706
Hagele
The use of biological products in freezing media can be an impediment to the international movement of embryos. An experiment was designed to evaluate the use of a chemically defined medium containing a macromolecule (polyvinyl alcohol; PVA) of non-biological origin (Bavister, 1981. J. Exp Zool. 217: 45-51) to freeze embryos. A total frozen consisted Img/ml consisted glucose, straws medium. C/minute Embryos bath for placed at room medium. modified and 48 in oi I one and
of 218 mouse embryos and 164 cow embryos were in two different media in a blind trial. Medium A of Dulbecco’s phosphate bufered saline (PBS), PVA, glucose, pyruvate and antibiotics. Hedium B of PBS, 3mg/ml bovine serum albumin (BSA), pyruvate and antibiotics. Embryos were frozen in containing a 1.5M glycerol solution in the chosen Straws were seeded at -70 C, cooled at 0.30 and plunged into liquid nitrogen at -350 C. were thawed by placing straws in a 350 C water 40 seconds. Embryos were recovered from straws, in a I.OH solution of sucrose in PBS for IO minutes temperature and then washed with PBS culture House embryos were placed in culture in a Brinster’s solution under oil and evaluated at 24 Cow embryos were placed in culture under hours. Ham’s F-10 medium with BSA and evaluated at 24 hours.
At 48 hours in culture 88 (41.3%) of 213 mouse embryos in medium A and 102 (52%) of 196 mouse embryos frozen frozen in medi urn 8 had developed to expanded or hatched blastocysts. This difference was significant (p <0.05). There was no significant difference in survival as judged by number of transferrable embryos when cow embryos were frozen in either medium A or B (Table 1). Table freezing
1. in
Survival PBS plus
of PVA
cow embryos (medium A) or Survival
Med i urn A B
1 hour 133 81
62.7% 66.7%
in BSA in
culture (Medium
Culture 24
after B)
hours 27.7% 35.8%
Results indicate that media containing macromolecules of non-biological origin can be used to successfully freeze mammal ian embryos and may be of use in international trade.
JANUARY
1987 VOL. 27 NO. 1
THE
USE
R.J.
OF
A CHEPllCALLY ORIGIN.
DEFINED HEDIUll. OF TO FREEZE EI’IBRYOS
NON-BIOLOGICAL
Mapletoft, B. Bavisterl. J.S. Moker and W.C. Dept. Herd Medicine and Theriogenology WCVM University of Saskatchewan Saskatoon, Sask. SlN OWO CANADA IDept of Vet Sci , University of Wisconsin Madison, WI USA 53706
Hagele
The use of biological products in freezing media can be an impediment to the international movement of embryos. An experiment was designed to evaluate the use of a chemically defined medium containing a macromolecule (polyvinyl alcohol; PVA) of non-biological origin (Bavister, 1981. J. Exp Zool. 217: 45-51) to freeze embryos. A total frozen consisted Img/ml consisted glucose, straws medium. C/minute Embryos bath for placed at room medium. modified and 48 in oi I one and
of 218 mouse embryos and 164 cow embryos were in two different media in a blind trial. Medium A of Dulbecco’s phosphate bufered saline (PBS), PVA, glucose, pyruvate and antibiotics. Hedium B of PBS, 3mg/ml bovine serum albumin (BSA), pyruvate and antibiotics. Embryos were frozen in containing a 1.5M glycerol solution in the chosen Straws were seeded at -70 C, cooled at 0.30 and plunged into liquid nitrogen at -350 C. were thawed by placing straws in a 350 C water 40 seconds. Embryos were recovered from straws, in a I.OH solution of sucrose in PBS for IO minutes temperature and then washed with PBS culture House embryos were placed in culture in a Brinster’s solution under oil and evaluated at 24 Cow embryos were placed in culture under hours. Ham’s F-10 medium with BSA and evaluated at 24 hours.
At 48 hours in culture 88 (41.3%) of 213 mouse embryos in medium A and 102 (52%) of 196 mouse embryos frozen frozen in medi urn 8 had developed to expanded or hatched blastocysts. This difference was significant (p <0.05). There was no significant difference in survival as judged by number of transferrable embryos when cow embryos were frozen in either medium A or B (Table 1). Table freezing
1. in
Survival PBS plus
of PVA
cow embryos (medium A) or Survival
Med i urn A B
1 hour 133 81
62.7% 66.7%
in BSA in
culture (Medium
Culture 24
after B)
hours 27.7% 35.8%
Results indicate that media containing macromolecules of non-biological origin can be used to successfully freeze mammal ian embryos and may be of use in international trade.
JANUARY
1987 VOL. 27 NO. 1