This study was carried out to investigate the effect of culture media, gonadotropins, proteins, growth factors and hyaluronic acid (HA) on the IVM rate of buffalo oocytes. Cumulus oocyte complexes were aspirated from ovaries and classified into 4 Grades, excellent and good quality COCs. Oocyte maturation was tested using TCM199, CR1aa or DMEM (Experiment 1); 10 µg/mL FSH + 50 IU/mL eCG, 10µg/mL FSH or 50 IU/mL eCG (Experiment 2): 10% FCS, 4mg/ml BSA or 1 µg/mL PVA (Experiment 3). Control group;10 µg/mL IGF-1 or 10 µg/mL EGF (Experiment 4) or 0, 2.5, 5 and 10 µg HA (Experiment 5). In all experiments, COCs were cultured for 252-24 h, then cytoplasmic and nuclear maturation was conducted. Results indicated that TCM199 achieved the best cytoplasmic and nuclear maturation. TCM199 supplemented with FSH and/or eCG enhanced in-vitro cytoplasmic and nuclear maturation of buffalo oocytes. Adding FCS, using growth factors, or 2.5 µg/mL HA toTCM199 lead to improving maturation rate. the results indicated that in-vitro maturation of buffalo oocytes is improved in TCM-199 supplemented with 50 IU eCG, 10μg/mL FSH, 10% FCS, 10 μg/ml EGF or 2.5 μg/mL HA.