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The efficacy of cryopreservation by direct plunging into liquid nitrogen (vitrification) of human pronuclear oocytes using open pulled straws with a super-finely pulled tip, as well as the ultrastructural changes caused by cooling and vitrification, were evaluated. Clinical and electron microscopic studies of cooled and vitrified oocytes were performed. Oocytes were cooled to 4°C in the presence and absence of cryoprotectants, vitrified, warmed, cultured and transferred. Abnormally fertilized oocytes were examined by electron microscopy. Vitrified and warmed 2-pronuclear oocytes showed 71.1% survival rate and 83.3% developmental rate. One- and 3-pronuclear oocytes, after cooling without cryoprotectants (presumably non-viable), showed progressive swelling of mitochondrial smooth endoplasmic reticulum (SER). After vitrification, oocytes (presumably viable) showed the formation of large SER vesicles associated with mitochondria. The described protocol of vitrification of human pronuclear oocytes was shown to be effective in producing pregnancy. Normal ultrastructure after undergoing the described vitrification protocol was confirmed. |
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