An-Najah Staff

Effects of first equilibration medium and co-culture with
oviduct epithelial cells on the vitrification of sheep
embryos derived in vitro
H Atalla1, S Pabuccuoglu2 and S Birler2
1Faculty of Veterinary Medicine, Nablus-West Bank, Palestine,
2Istanbul University, Avcilar-Istanbul, Turkey
Effects of two different vitrification protocols on the survival
of sheep embryos were examined. Blastocyst stage embryosproduced in vitro with (C) or without co-culture (CC) with
sheep oviduct epithelial cells were used in this study. Oocytes
were collected from slaughtered ewes, matured in medium 199
supplemented with sodium pyruvate, FSH, LH and 10% FCS
for 24 h, fertilized with fresh ram semen in bicarbonate
buffered synthetic oviduct fluid with 2% sheep oestrous serum
for 20 h and cultured in SOF medium. Glucose was added to
culture medium on the 4th day of culture. Blastocysts were
assigned two equilibration groups randomly; 20% ethylen
glycol (EG) or 10% glycerol (G) for the first equilibration.
After 5 min, all were kept in 20% ethylen glycol plus 10%
glycerol for 5 min as the second equilibration. After 30 s in
vitrification solution (25% ethylen glycol plus 25% glycerol),
they were immersed into liquid nitrogen. Thawing was carried
out in a water bath at 20C for 15–20 s and blastocysts were
transferred into 0.25 M sucrose for 5 min, washed in hepes
buffered synthetic oviduct fluid, and cultured in synthetic
oviduct fluid for 24 h. Survival rates of vitrified-thawed and
cultured blastocysts were 62.10% in C-EG, 38.40% in CC-EG,
30.20% in C-G and 39.30% in CC-G groups. This study shows
that vitrification of sheep embryos using ethylene glycol
instead of glycerol as a first equilibration cryoprotectant could
give reasonable survival rates and that co-culture of embryos
with sheep oviduct epithelial cells could not improve survival
rates.