Development of a synthetic medium for the in vitro culture of bovine embryos

In the majority of media for embryo culture, 2 of typical components used are FCS or BSA; however, the presence of FCS in the culture medium has been shown to have a negative effect on embryo quality and the use of animal-derived proteins in culture media increases the risks of disease transmission through in vitro embryo production. The aim of this study was to develop an in vitro embryo culture medium free from FCS and BSA, but with the addition of various growth factors and cytokines (GF-CYK: IGF-I, IGF-II, bFGF, LIF, GM-CSF) 50ngmL(-1) and (TGF-β1) 100ngmL(-1) supplemented with hyaluronan (HA) and recombinant albumin (RA). Bovine oocytes (n=1043, 6 replicates) from abattoir ovaries were matured in TCM-199 medium with 60μgmL(-1) penicillin, 60μgmL(-1) streptomycin, and 10ngmL(-1) EGF for 24h at 39°C and 5% CO2 in humidified air. Afterward, the oocytes were fertilized in IVF-TALP medium with 6mgmL(-1) fatty acid-free BSA and 1.7IUmL(-1) heparin for 18h under the same conditions. After fertilization, presumptive zygotes were divided into two groups and cultured in 30μL droplets of SOF supplemented with (1) 0.4% BSA+5μgmL(-1) insulin, 5μgmL(-1) transferrin, and 5ngmL(-1) selenium (ITS) as a control; or (2) GF-CYK+0.5mgmL(-1) HA+0.15% RA (M1). Droplets were preserved under mineral oil in a humidified atmosphere of 5% CO2, 5% O2, and 90% N2 at 39°C. Blastocyst development and blastocyst diameter was observed at 7 and 8 days post-fertilization (dpf). Developmental and diameter data were analysed using the Wilcoxon test by using R software. The blastocyst rates were not significantly different between the control and M1 medium: at 7dpf (22.9%±4.8 and 30.2%±3.0), and at 8dpf (29.6%±5.1 and 37.4%±2.0 respectively; P>0.05). The blastocyst diameter obtained with the M1 medium was significantly greater (P