255. In vitro maturation of bovine oocytes in serum-free media
S. Zhang A C , A. J. French A C and R. T. Tecirlioglu BA Centre for Reproduction and Development, Monash Institute of Medical Research, Monash University, 27-31 Wright Street, VIC, Australia
B Monash Immunology and Stem Cell Laboratories, Monash University, Clayton, VIC, Australia
C CRC for Innovative Dairy Products, Melbourne, VIC, Australia
Reproduction, Fertility and Development 17(9) 102-102 https://doi.org/10.1071/SRB05Abs255
Submitted: 26 July 2005 Accepted: 26 July 2005 Published: 5 September 2005
Abstract
Culture medium supplemented with sera is commonly used for the in vitro production (IVP) of livestock embryos. However, serum induced complications including batch variation, the potential risk of virus and mycoplasma contamination and the implication in the large offspring syndrome in domestic animals impels the development of a serum-free culture system. In this study, we investigated whether replacement of fetal bovine serum (FBS) with bovine serum albumin (BSA) in three maturation media, tissue culture medium-199 (TCM-199), a modified synthetic oviduct fluid (mSOF) routinely used in our laboratories and a commercially available SOF-VC (Vitro Cleave, Cook Australia). Harvested oocytes were matured, parthenogenetically activated and in vitro cultured (Day 7) to measure maturation efficiency, embryo development and quality with the aim of developing a simplified and defined culture medium for the in vitro production of bovine embryos. Abattoir derived cumulus oocyte complexes were matured in TCM-199, mSOF and SOF-VC media supplemented with LH and beta-estradiol in the presence of 15% FBS or 0.08% BSA at 39ºC in 5% CO2 in air. Polar body extrusion was assessed twenty-two hour post maturation and selected MII occytes were activated using calcium ionophore/6-dimethylaminopurine and cultured for seven days in SOF medium supplemented with 0.8% BSA. On day seven, blastocyst development was assessed and randomly selected blastocysts were stained to determine inner cell mass (ICM), trophectoderm (TE) and total cell numbers (TCN). Supplementation with either BSA or FCS did not significantly affect the maturation efficiency, blastocyst rates or differential cell numbers within each maturation media tested. However, maturation efficiency and blastocyst rates were significantly lower (P < 0.01) when oocytes were matured in either mSOF or SOF-VC regardless of FBS or BSA supplementation. From this study, we conclude that BSA effectively replaces FCS and TCM-199 is superior to SOF (mSOF or SOF-VC) in terms of oocyte maturation regardless of protein source. Once matured SOF and TCM-199 parthenogenetically blastocysts were equivalent in terms of embryo development and quality.
