Abstract

Mammalian spermatozoa were among the very first cells to be successfully cryopreserved and over the last five decades the use of frozen-thawed semen for artificial insemination has come to play an important role in domestic livestock production. More recently, semen freezing has increasingly been utilized in the establishment of genetic resource banks for endangered species. Semen is collected, most commonly either by use of an artificial vagina or by electroejaculation of an anaesthetized animal, and basic sperm parameters assessed. Semen is extended using a TEST-egg yolk-glycerol diluent, packaged in 0.25-mL plastic straws and slowly cooled to 5°C over a period of 1–2 h. Cooled straws are frozen by suspending within liquid nitrogen vapor above the liquid nitrogen surface before plunging into the liquid phase. Straws are thawed briefly in air before immersing in a 35°C water bath for 15 s, and often are used directly for insemination without any further processing.