Cryopreservation of Bovine, Equine, and Porcine Sperm

In livestock breeding, cryopreserved semen is routinely used for artificial insemination. Sperm cryopreservation enables long-term storage and transport of insemination doses, thereby securing genetic material for later use. Although specifics may vary, commonly used sperm cryopreservation protocols for domestic species feature similar processing steps as outlined in this chapter. We here describe common procedures for cryopreservation of bull, stallion and boar sperm. This includes preparation and use of the following diluents: TRIS egg yolk extenders (TEY) for bull sperm, ordinary and freezing skim milk extenders (INRA-82) for stallion sperm, and Beltsville thawing solution (BTS) and lactose egg yolk extenders (LEY/GO) for use with boar sperm. Details and schematic presentations are given for dilution with cryoprotective agents, using a one-step versus a two-step approach. Furthermore, simple methods are presented for packaging of insemination doses in straws, slow cooling of sperm samples to 5°C, controlled freezing in a polystyrene box with liquid nitrogen vapor, and rapid warming at 37°C.