32 Bovine embryo cryopreservation in a chemically defined medium

Abstract

Embryo cryopreservation media often contain an animal-derived component, such as bovine serum albumin (BSA) or fetal calf serum. However, the industry is faced with the issue of composition variability between batches and, most importantly, the risk of pathogen transmission. The aim of this study was to compare the effectiveness of two embryo cryopreservation ethylene glycol-based media: IMV's freezing medium with BSA (IMV Technologies) and a chemically defined medium containing STEMALPHA.CRYO3, called CRYO3 (Ref 5617, Stem Alpha). CRYO3 is produced according to EU good manufacturing practice, ensuring the composition and quality of the product. Bovine morulae were collected in vivo and split into two groups to be slow frozen in both media. All results were analysed with chi-squared tests with Yates's correction. For the in vitro approach of this study, frozen morulae (n = 86) were thawed, cultured in medium (1:1:1 mixture of RPMI, Dulbecco's modified Eagle's medium, and Ham's F10) at 38.5°C with 5% CO₂, and monitored every 24 h for 72 h to evaluate survival and expansion rates. Survival rate was higher in the CRYO3 group than in the BSA group (74 and 50%, respectively; P < 0.05). Expansion and hatching rates followed the same trend: 38 and 14%, respectively, at 48 h (P < 0.05) and 40 and 18% at 72 h (P < 0.05). For the in vivo approach of this study, frozen morulae (n = 123) were thawed and transferred on the field to evaluate pregnancy and calving rates. Field results showed no significant difference between CRYO3 and the BSA-based medium for pregnancy rates (73 and 63%; P = 0.3006) nor for the delivery rates (64 and 55%; P = 0.3614).The trial was conducted with different AI teams and in 10 commercial herds, thus adding robustness to the data. This study has shown that CRYO3 can replace BSA in an ethylene glycol cryopreservation medium for in vivo-produced bovine embryos. Therefore, this product eliminates some sanitary risk in embryo commerce, which is a concern in an international context.