Vitrification of porcine oocytes and zygotes in microdrops on a solid metal surface or liquid nitrogen

Abstract

Oocyte cryopreservation is a potent approach to keep female germplasm safe from epidemic diseases. In the last decade, we developed simple, cheap, and robust vitrification protocols which enable quick cryopreservation of immature porcine oocytes and zygotes in large numbers. In this chapter, we describe vitrification procedures for porcine oocytes and zygotes where they are vitrified in 1–2 μL aliquots of a defined (protein-free) vitrification medium and dropped either on a metal surface pre-cooled from the bottom with liquid nitrogen (solid surface vitrification) or directly into liquid nitrogen. Vitrified microdrops can be stored in cryo-vials in liquid nitrogen. Low concentrations of permeating cryoprotectants during equilibration and proper temperatures during equilibration and warming are crucial for achieving high survival rates. The device used for cooling does not seem to affect system efficacy as vitrification of oocytes or zygotes either on Cryotop® sheets or in microdrops were equally effective.