323 OPTIMIZATION OF SERUM-FREE IVP PROTOCOL: EFFECT OF SERUM-FREE IVM ON BOVINE EMBRYO DEVELOPMENT
M.E. Räty A , E.H. Ketrja A , K.M. Kananen-Anttila B and J.M.H. Peippo AA MTT Agrifood Research Finland, Animal Production Research, FIN-31600 Jokionen, Finland. email: mervi.raty@mtt.fi;
B University of Kuopio, Institute of Applied Biotechnology, FIN-70211 Kuopio, Finland.
Reproduction, Fertility and Development 16(2) 281-281 https://doi.org/10.1071/RDv16n1Ab323
Submitted: 1 August 2003 Accepted: 1 October 2003 Published: 2 January 2004
Abstract
The use of serum in IVP medium may cause abnormalities, e.g. reduced post-thaw survival, in bovine IVP embryos (Abe H et al., 1999 Mol. Reprod. Dev. 53, 325–335). Moreover, serum may be a source of contamination and its composition is highly variable. Several reports have shown that serum-free IVF–IVC does not compromise embryo development (see, e.g. Keskinteipe L et al., 1996 Biol. Reprod. 55, 333–339). Here we studied the effect of serum-free IVM on bovine IVP embryo development, aiming for production of good quality Day 7 embryos for freezing. In total, 11 734 abattoir-derived bovine oocytes in 13 batches were washed with emCare Complete Medium with BSA (1 mg L−1) and matured for 24 h in TCM-199 with glutamax-I (Gibco, Paisley, UK), 0.25 mM Na-pyruvate, 100 IU mL−1 penicillin, 100 μ g mL−1 streptomycin, 10 μg mL−1 LH, 2 μg mL−1 FSH and 1 μ g mL−1 β-estradiol supplemented either with 1) 10% FBS (Gibco, New Zealand), 2) 4 mg mL−1 fatty acid-free albumin (FAF-BSA), 3) 4 mg mL−1 FAF-BSA + growth factors (GF;; 100 ng mL−1 IGF-I + 100 ng mL−1 EGF), 4) 4 mg mL−1 polyvinylpyrrolidone (PVP), 5) 4 mg mL−1 PVP+GF and 6) PVP + amino acids (10 μL mL−1 MEM + 20 μL mL−1 BME). After 20 h fertilization in FERT-TALP + 2 mg mL−1 BSA with semen of pre-tested IVF-bull, the oocytes were denuded and cultured in modified SOFaaci + 6 mg mL−1 FAF-BSA in 5% O2 (Holm P et al., 1999 Theriogenology 52, 683–700). The statistical analyses are based on logistic regression models with IVP batch and treatment as explanatory variables. The estimated probabilities (P) are shown in Table 1. The upper and lower values of 95% confidence intervals varied within P ± 0.05, P ± 0.03, and P ± 0.02 for cleavage, Day 7 embryo development and development of good quality Day 7 embryos, respectively. PVP+GF-IVM supported cleavage equally well as FBS-IVM (P = 0.49), whereas the remaining serum-free IVM treatments had lower embryo cleavage rate than FBS-IVM (P < 0.01). On Day 7 none of the serum-free IVM treatments supported embryo development and development of good quality embryos as well as FBS-IVM (P < 0.01). Addition of GF in FAF-BSA-IVM reduced embryo cleavage, Day7 embryo development and development of good quality embryos compared to that of FAF-BSA alone (P < 0.03). PVP-IVM resulted in lower embryo cleavage rate than PVP+GF (P < 0.005), whereas according to the two other criteria there were no differences between the treatments (P > 0.13). In conclusion, these preliminary results indicate that replacing the FBS as a protein source in IVM needs more optimization.
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