Vitrification is a common method which is successfully used for cryopreservation of gametes
and embryosin in several animal species, though subsequent progress is still limited,especially in buffalo.
The present study aimed to evaluate the effect of vitrification process on nuclear status in matured
buffalo oocytes. Morphology, viability, state of nuclear maturation in meiotic stages and maturation rate (matured oocyte/survived oocyte) were assessed in vitrified matured buffalo oocytes.Immature buffalo oocytes were matured in-vitro for 22 hrs. and vitrified at room temperature (25°C) in a mixture of ethylene glycol (EG)and dimethyl sulfoxide (DMSO) with 0.25 ml straw. Vitrification solutions (VS) were VS (1.5M EG+ 1.5M DMSO) 1 and VS (3M EG+ 3M DMSO). Cryoprotectants were added in two steps, with the first step concentration half 2 that of the second (and final) step concentration. After warming, normal oocytes were cultured for further 2 hrs., stained with trypan blue for viability evaluation, then fixed and stained with orcein 1% stain for detection of meiotic stages. Oocytes that reached Telophase I (TI) or Metaphase II (MII) stages were considered matured.
Results showed that morphologically normal and viable oocytes were significantly decreased in vitrified oocytes than control. The percentage of oocytes reached TI and MII in un-vitrified (control) group was significantly (P<0.05) higher than those vitrified after maturation. The maturation rate was higher in control than
vitrified oocytes (76.04±2.29 vs. 66.46±2.25). In conclusion, vitrification is a successful method for cryopreservation of matured buffalo oocytes. However, inspite of higher rates of viability and maturation obtained in current study, it is still lower than un-vitrified oocytes. |
