ABSTRACT
We developed new techniques to improve freezing and vitrification of sperm, oocytes and embryos. Our novel freezing technology is based on 'Multi-Thermal-Gradient' (MTG) freezing that is used for sperm. The freezing apparatus has the ability to control ice crystals propagation by changing thermal gradient or the liquid-ice interface velocity which optimizes ice crystals morphology during freezing of cells and tissue. Using this apparatus we were able to freeze bull, stallion, boar, ram, fowl and human sperm with normal post-thaw motility/pre-freezing motility of 70-100%. The vitrification method includes the cooling of nanoliter sample (the 'Minimum Drop Size' technique) in 'super-cooled' liquid nitrogen (-210 degrees C), which maximized cooling rate to the highest physically possible (24-130000 degrees C/min). Using this method we achieved very high survival of bovine oocytes and embryos. Vitrification of oocytes at the MII stage resulted with cleavage and blastocyst rate of 50 and 20%, respectively. The vitrification of in-vitro production (IVP) of bovine embryos allowed the production of a healthy calf after embryo-transfer carrying the name 'Zegugit' (in Hebrew: made from glass).
