Sperm cryopreservation, in vitro fertilization, and embryo freezing

The cryopreservation of gametes and tissues is one of the most efficient tools for managing both large and small collections of valuable genetic resources. Cryopreservation minimizes the expenses associated with animal and facility maintenance, such as personnel, water, power, and space. It extends the time that offspring can be produced from individual organisms, reduces the need to maintain live populations, provides flexibility for planning future experiments and research projects, and can prevent the catastrophic loss of irreplaceable research lines. For zebrafish, the cryopreservation of sperm has been routinely practiced since the 1980s, whereas the cryopreservation of embryos has not been possible until recently. In this chapter, we present the procedures for sperm collection, dilution, cryopreservation, thawing, and in vitro fertilization used at the Zebrafish International Resource Center to successfully bank fish lines. In the second part, we present the latest protocol developed at the University of Minnesota to cryopreserve fish embryos using microinjection, rapid cooling, and laser warming.