Long-term in vitro culture of bovine preantral follicles: Effect of base medium and medium replacement methods

Two culture media and replacement methods were compared during long-term in vitro culture of secondary follicles of cattle using α-MEM+ or TCM-199+ as base media. The medium replacement methods were: Conventional – removal and subsequent addition of the same amount (60μl) in a 100μl aliquot (MEM-C and TCM-C), and Small Supplementation – addition of 5μl of fresh medium to an initial small aliquot (50μl), resulting in a final volume of 125μl on the last day of culture (MEM-S and TCM-S). A total of 207 secondary follicles were cultured individually for 32 days at 38.5°C in 5% CO2 and medium replacement was performed every other day. The MEM-S treatment resulted in a larger (P