This study was conducted to evaluate the protective effects of cholesterol pre-incubation on ovine oocytes exposed to cold stress and vitrification. In the first experiment, cumulus oocyte complexes harvested from ovaries of slaughtered ewes were exposed to 2 mg/ml cholesterol loaded methyl-ßcyclodextrin (CLC) in maturation medium for 2, 4 or 24 h. Oocytes were then exposed to cold stress at 4°C for 30 min, and the viability and chromosome abnormality of oocytes were evaluated. Our results showed no significant differences between treatment groups in regarding to oocyte viability or ooplasm integrity. However, oocytes exposed to cold stress without preincubation with CLC had significantly higher chromatin dispersion (P<0.05) than pre-incubated oocytes with CLC. In second experiment, oocytes were exposed to 2 mg/ml CLC in maturation medium for 2 h and vitrified after maturation in conventional straws (0.25 ml; CS) or in open pulled straws (OPS). Osmotic stress of vitrification media to oocytes was also evaluated. Oocytes pre-treated with CLC had a significantly higher viability rate and ooplasm integrity than non-treated oocytes after vitrification in OPS (P < 0.05). The findings of this study also show that CLC pre-incubation of ovine oocytes reduce the chromosome degeneration after vitrification in both OPS and CS.