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Cryopreservation and Cryobiology of Mammalian Oocytes
Kasai, Magosaburo,Song, Hai-Bum 대구대학교 생명과학연구소 2003 생명과학연구 Vol.1 No.3
The cryopreservation of embryos has become a powerful tool in assisted reproduction in several mammalian species and a more important technology in bioscience, agriculture and medicine. Embryos are cryopreserved by slow freezing or by vitrification. However, consistently high survival has not been obtained in most oocytes and in some embryos. The main reasons for the low survival would be sensitivity to low temperatures, which leads to chilling injury, and low permeability of the cell membrane, which leads to the formation of intracellular ice. As a stratefy aiming to overcome these injuries, modified vitrification methods have been devised in which the cooling and warming rate is markedly increased by minimizing the volume of the solution and the container. The modified methods use electron microscope grids, open-pulled straws, cryoloops, or container-less microdrops. Ultrarapid vitrification is a promising approach to overcoming the problems of the chilling sensitivity and lower permeability of oocytes/embryos. However, to make this method more reliable, defining the optimal combinations of cooling rate and the composition of the vitrification solution for each type of oocyte/embryo will be necessary. Refinement of the procedures for practical use, including labeling and stable preservation, would also be important.
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Han Myung-Sook,Koji Niwa,Magosaburo Kasai 한국발생생물학회 2003 한국발생생물학회 학술발표대회 Vol.2003 No.1
In cryopreserved rat embryos, survival rates obtained in vitro are not always consistent with the rates obtained in vivo. To determine the optimal conditions for in vivo development to term, rat embryos at the 4-cell, 8-cell and morula stages were vitrified in EFS40 by a 1-step method and transferred into oviducts or uterine horns of recipients at various times during pseudopregnancy. Vitrified and fresh 4-cell embryos only developed after transfer into oviducts of asynchronous recipients on Day -1 to -2 of synchrony, i.e., at a point in pseudopregnancy that was 1-2 days earlier than the embryos. However, although about half the vitrified embryos transferred into oviducts on Day -1 developed to term, only a minority of embryos transferred at later times did so, whether vitrified (10-34%) or fresh (24-33%), suggesting that this may not be the most suitable stage for cryopreservation. Very few 8-cell embryos, either vitrified or fresh, developed when transferred into oviducts on Day 0 to -0.5. However, when transferred into uterine horns, high proportions of vitrified 8-cell embryos (-63%) developed to term in reasonably synchronous recipients (Day 0 to -0.5) but not in more asynchronous ones (6%; Day-1). A majority of vitrified morulae also developed to term (52-68%) in a wider range of recipients (Day 0 to -1), the greatest success occurring with recipients on Day -0.5. Similar proportions of vitrified and fresh 4-cell embryos, 8-cell embryos and morulae developed to term when there was appropriate synchronization between embryo and recipient. Thus vitrification of preimplantation stage rat embryos does not appear to impair their developmental potential in vivo.
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