Use of Antifreeze Protein from (TmAFP) in Vitrification of -Produced Bovine Embryos: An Ultrastructural Study.

The objective of this study was to evaluate the effects of different concentrations of antifreeze protein (AFP) extracted from the larva of the beetle, (AFP), on vitrification of -produced bovine embryos. -produced blastocysts were divided into three experimental groups and vitrified using a cryotop. AFP was added to the equilibrium solution (ES) and vitrification solution (VS) at a concentration of 0 ng/mL (control), 500 ng/mL (500AFP), or 1000 ng/mL (1000AFP). Vitrification was carried out by first placing the blastocysts in ES for 2 minutes (7.5% ethylene glycol [EG] and 7.5% dimethyl sulfoxide [DMSO]). The blastocysts were then transferred to VS (15% EG and 15% DMSO) and promptly deposited on a cryotop stem and submerged in liquid nitrogen. Warming was carried out in three steps with decreasing sucrose concentrations. After warming, the blast cells were cultured for 24 hours for subsequent survival analysis and ultrastructural evaluation. There was a significant difference in the survival rate and expansion in the 500AFP group compared with the other groups. The ultrastructural analysis revealed intracellular lesions in all vitrified embryos; however, the embryos of the 500AFP and 1000AFP groups showed fewer cytoplasmic lesions compared with the control group. Taken together, addition of AFP can mitigate cellular changes that involve organelles and cellular components essential for proper functioning and improve the viability of warmed and vitrified -produced bovine embryos.