Cryopreservation of Bovine Blastocysts and Blastocoel Fluid Aspiration for Large Embryos

The aspiration of blastocoele fluid (BF) from bovine blastocysts is a minimally invasive technique that allows artificial collapse of large embryos before cryopreservation, and, optionally, the collection of the BF from the blastocoelic cavity for its further analysis. The procedure aims to remove most of the fluid from the blastocoelic cavity to mitigate the severity of osmotic changes when embryos are exposed to cryoprotectants and to decrease the chance of intracellular ice crystal formation during cryopreservation. This chapter provides a step-by-step methodology for BF aspiration prior to cryopreserve in vitro-produced (IVP) bovine blastocysts by vitrification. Vitrification using the Cryotop ® method and following a short equilibration, as well as warming and step-wise dilution, are also described.