Patent ID: 7943293

Claim:
A method of freezing or vitrifying an embryo or oocyte specimen or specimens for storage in liquid nitrogen, said method comprising; a) removing said specimen or specimens from a culture medium and placing said specimen or specimens in a first rejuvenation medium comprising effective amounts of glucose, pyruvate, amino acids, vitamins K5 and C, antioxidants, and fatty acids sufficient to supply the specimen or specimens with chemical ingredients and uptake requirements required to recover and prosper during the cryopreservation process, said first rejuvenation medium being operative to supply nutrients which will protect from depletion and damage to said specimen or specimens and its or their mitochondria, spindles, and structural features during cryopreservation, said first rejuvenation medium being devoid of HEPES or MOPS; b) removing said specimen or specimens from said first rejuvenation medium and placing said specimen or specimens into a second rejuvenation medium comprising effective amounts of glucose, pyruvate, amino acids, vitamins K5 and C, antioxidants, and fatty acids sufficient to supply the specimen or specimens with chemical ingredients and uptake requirements required to recover and prosper during the cryopreservation process, said second rejuvenation medium being operative to supply nutrients which will protect from depletion and damage to said specimen or specimens and its or their mitochondria, spindles, and structural features during cryopreservation, said second rejuvenation medium including HEPES or MOPS; c) removing said specimen or specimens from said second rejuvenation medium and placing said specimen or specimens in a first solution comprising a first amount of one or more cryoprotectants; d) removing said specimen or specimens from said first solution and placing said specimen or specimens in a second solution comprising a second amount of one or more cryoprotectants which is greater than said first amount of cryoprotectant in step c); and e) removing said specimen or specimens from said second solution and placing said specimen or specimens in liquid nitrogen.