Matured (MII) oocytes were selected and manipulated in TCM-Hank's supplemented with 10% FBS, 1% antibiotic/antimycotic solution and 0.2 mM sodium pyruvate. Oocytes were incubated in groups of eight for 20 to 30 min in 10 μg/mL cytochalasin B and 5 μg/mL Hoechst 33342 for enucleation by micromanipulation