Abstract

The aim of the study was to determine the optimum concentration of the mare serum (MS) for sheep in vitro oocyte maturation. Sheep ovaries were collected from a local abattoir and transported within 1 h to the laboratory in a warm saline solution (30 – 35^oC), supplemented with 100 IU penicillin G and 100 μg streptomycin sulfate/ml. Cumulus-oocyte complexes (COC’s) were obtained by slicing of follicles, washed in TCM-199 modification with NaHCO₃ and supplemented with 50 μg/ml gentamycin, and 0.25 mM sodium pyruvate without any serum supplementation. The COC’s were randomly divided into four groups. Group 1 (n = 105) COC’s were fresh control and cultured in TCM-199 medium without serum supplementation. Group 2 (n = 108) COC’s were washed five times and cultured in TCM-199 medium supplemented with 10% MS. Group 3 (n = 112) COC’s were washed five times and cultured in TCM-199 medium supplemented with 15% MS. Group 4 (n = 114) COC’s were washed five times and cultured in TCM-199 medium supplemented with 20% MS. After 38 - 42 h of IVM, oocytes were denuded with the aid of 0.1% hyaluronidase and passing them through a fine pipette, fixed for 24 – 48 h in a mixture of acetic acid and alcohol (1:3) at room temperature, stained for 10 min with 1% (w/v) orcein in 45% acetic acid and examined for the evidence of different stages of maturation. Significantly higher (p < 0.05) maturation rates of oocytes (69 – 72%) were observed in all concentrations of mare serum compared to those without serum supplementation. However, no significant difference was observed between the 10, 15 and 20% serum supplemented group.

Key words: Sheep; Mare serum; Follicle slicing; Oocyte maturation; Ovary.