**1. Introduction**

The cryopreservation technique is widely used in reproductive biology because it facilitates the application of assisted reproduction techniques such as artificial insemination and *in vitro* fertilization (IVF). Besides that it allows long-term storage of gametes, facilitating transport over long distances and the formation of a genetic bank, essential for the conservation of species. It is an important complementary tool for conservation to avoid excessive losses of genetic variation or races of extinction. Cryopreservation is considered to be the most effective way for long-term preservation of genetic materials, cells, tissues and microorganisms. It

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

allows to maintain biological material at very low temperatures, indefinitely. The expansion of genetic material from high-value breeders became possible with the advent of semen cryopreservation. The main advantages of the technique are: portability for world trade because it is not necessary to purchase or move the males, and it also acts in the prevention of diseases capable of transmission through natural breeding.

Semen constitutes the union of sperm with seminal plasma, secreted by the male accessory glands. Sperms are produced in the testicles (gametogenesis), and maturation occurs in the epididymis when cells enter in the caput of epididymis, progress to the corpus, and finally reach the cauda region, where they are stored until the moment of ejaculation. Spermatozoa stored in the cauda region are generally of good quality and have a high level of maturity, being able to fertilize oocytes. The epididymis provides a favorable environment to retain the sperm with fertilizing capacity for several weeks. Therefore, the cauda epididymis is a major source of gametes of a breeder and in special cases they can be retrieved directly from it.

The sperm recovery from the epididymis is the last chance to use gametes from dead breeders, in addition to enabling the maintenance of a germplasm bank of animals of commercial interest or threatened with extinction [1]. A factor to be considered is the difficulty in collecting semen from wild species and the unexpected death of animals of zoological interest. In this case, the technic can be used to ensure the rescue of male gametes and the preservation by cryopreservation process for maintaining germplasm banks [2]. For animal production as livestock, the sperm recovery directly from the epididymis is a viable alternative when there is a sudden death of breeder of high commercial value, to increase their genetic stock or in cases of bulls unable to ejaculate for some reason [3]. It is the last alternative for future use of gametes from a breeder in assisted reproduction programs. In equine, this technique is important because there is a high incidence of accidental and unexpected death due to high incidence of colic or severe traumatic accidents that compromise the reproductive life of the stallion.

The first important point is the recovery of still viable sperm with good parameters of motility, concentration, and morphological defects as soon as possible after the male death. There is a time limit for this to successfully occur, usually dependent on temperature. The first step is getting the epididymis in the field, after the death of the animal. For handling ease, due to their anatomical location, the testes can be removed with them. Then both should be sent to the laboratory in suitable containers, with or without cooling. In the lab, the trained technicians perform gonads cleaning, epididymis isolation and spermatozoa recovery. Then it can be used in three different ways: first, shortly after harvesting; second, chilled, and third, after cryopreservation process. The last increases the availability time of gametes for application of assisted reproduction techniques. Next we will focus on the key topics relevant to the understanding of the epididymis spermatozoa cryopreservation technique.
