The objective of the current study was to investigate and compare the role of buffalo oviductal epithelial cell (BOEC) co-culture and conditioned culture media, on in vitro development of Nili Ravi buffalo embryos. Oocytes were obtained from the ovaries of slaughtered buffaloes within two hours after slaughter and brought to laboratory. After in vitro maturation (IVM) in Tissue Culture Medium-199 (TCM199) for 24 h and fertilization (IVF) in Tyrode’s Albumin Lactate Pyruvate (TALP) medium for about 20 h, the presumptive zygotes were randomly distributed into 6 culture groups; Group I: Synthetic Oviductal Fluid (SOF) medium alone, Group II: SOF + co-culture, Group III: Conditioned SOF, Group IV: M199 alone, Group V: M199 + co-culture and Group VI: Conditioned M199 for in vitro culture (IVC). The percentage of embryos capable of crossing 8-16 cell block and reaching morula stage were higher (P < 0.05) in Group II and Group III compared to other treatments. Both treatment groups were equally effective (P > 0.05). Results also indicated that synthetic oviductal fluid (SOF) medium was found better than M199 when co-cultured and conditioned with buffalo oviductal epithelial cells for culturing buffalo embryos. Conditioned SOF is recommended to be used as IVC media in future studies as it eliminates contamination problem, remove batch to batch variation, long-lived and is readily available.