Cryopreservation of Semen from Domestic Livestock: Bovine, Equine, and Porcine Sperm

Part of the Methods in Molecular Biology book series (MIMB, volume 2180)


In modern livestock breeding, cryopreserved semen is routinely used for artificial insemination. Sperm cryopreservation allows for long-term storage of insemination doses and secures reproduction at a desired time point. In order to cryopreserve semen, it needs to be carefully processed to preserve its vital functions after thawing. In this chapter, we describe the processes involved in cryopreservation of bull, stallion, and boar sperm. These include preparation of diluents, dilution of sperm in primary and freezing extender, slow cooling from room temperature to 5 °C, packaging of insemination doses in straws, freezing at a defined cooling rate in liquid nitrogen vapor, cryogenic storage, and thawing. Two-step dilution approaches, with commonly used diluents, are presented, namely, TRIS-egg yolk (TEY) extender for bull sperm, skim milk (INRA-82) extender for stallion sperm, and lactose-egg yolk (LEY) extender for boar sperm. Furthermore, simple methods are presented for cooling and freezing of sperm at defined cooling rates.

Key words

Bull Cryopreservation Lactose-egg yolk extender Skim milk extender Pig Semen Spermatozoa Stallion TRIS-egg yolk extender 



Employees of the Unit for Reproductive Medicine of the University of Veterinary Medicine Hannover (Hannover, Germany), National Stud of Lower Saxony (Celle, Germany), and Masterrind GmbH (Verden, Germany) are acknowledged for sharing protocols.


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2021

Authors and Affiliations

  1. 1.Unit for Reproductive Medicine—Clinic for HorsesUniversity of Veterinary Medicine HannoverHannoverGermany
  2. 2.Biostabilization Laboratory—Lower Saxony Centre for Biomedical Engineering, Implant Research and DevelopmentUniversity of Veterinary Medicine HannoverHannoverGermany

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