摘要
本文研究不同类型的防冻剂及其浓度对快速冷冻牛胚胎的效果。试验Ⅰ：７日龄囊胚先分别放入含１０％（Ｖ／Ｖ）甘油或１０％（Ｖ／Ｖ）乙二醇改良的磷酸盐缓冲液（ＰＢＬ）中，室温下平衡１０ｍｉｎ。然后，分别移入１．５ｍｏｌ／Ｌ，２．０ｍｏｌ／Ｌ甘油或２．０ｍｏｌ／Ｌ，３．０ｍｏｌ／Ｌ乙二醇＋０．２５ｍｏｌ／Ｌ蔗糖ＰＢＬ液中，１０ｍｉｎ后将胚胎放进冰箱的结冰层（约─２３℃），３５ｍｉｎ后直接投入液氮。冻后孵化率最高的处理组为１．５ｍｏｌ／Ｌ甘油＋０．２５ｍｏｌ／Ｌ蔗糖（８２．４％），其极显著（Ｐ＜０．０１）地高于２．０ｍｏｌ／Ｌ甘油＋０．２５ｍｏｌ／Ｌ蔗糖（１５．４％），２．０ｍｏｌ／Ｌ乙二醇＋０．２５ｍｏｌ／Ｌ蔗糖（６３．９％），３．０ｍｏｌ／Ｌ乙二醇＋０．２５ｍｏｌ／Ｌ蔗糖（３８．７％）。试验Ⅱ：７日龄胚胎先在１０％（Ｖ／Ｖ）甘油中平衡１０ｍｉｎ，而后，分别移入１．５ｍｏｌ／Ｌ甘油＋０，０．２５，０．５ｍｏｌ／Ｌ蔗糖，停留１０ｍｉｎ后快速冷冻。解冻后，胚胎分别用０．５，０．２５，０ｍｏｌ／Ｌ蔗糖除甘油。结果表明：解冻液中的蔗糖浓度对胚胎的存活力无显著影响。当冷冻处理为１．５ｍｏｌ／Ｌ甘油＋０．２５ｍｏｌ／Ｌ蔗糖时，胚?
A simple method for rapid freezing of bovine IVF embryos was investigated in this study. In Exp.Ⅰ, Day 7 embryos were firstly exposed to equilibration medium of either 10% glycerol (G) or 10% Ethylene glycol (EG) in PBS for 10 min, and secondary correspondently transferred to 1. 5, 2. 0 mol/L G or 2. 0, 3. 0 mol/L EG + 0. 25 mol/L sucrose (S) in PBS for 10 min, and held in a domestic freezer at -23℃ for 35 min, before plunging into liquid nitrogen. The results indicated that the hatching rate of embryos frozen in medium with 1. 5 mol/L G + 0. 25 mol/L S was significantly higher (P<0. 01)than that in media with 2. 0 mol/L G +0. 25 mol/L S (15. 4% ), 2. 0 mol/L EG (63. 9%)and 3. 0 mol/L EG (38. 7% ), respectively. Embryos frozen in 2. 0 mol/L EG showed significantly higher (P<0. 01) hatching rate than those frozen in 3. 0 mol/L EG. In Exp.Ⅱ, Day 7 blastocysts were quickly frozen with the same method as Exp Ⅰ. after equilibration in 1. 5 mol/L G + 0,0. 25 or 0. 5 mol/L S at 25℃ for 10 min. After thawing, glycerol was removed in PBS puls 0, 0. 25 or 0. 5 mol/L S, respectively. The sucrose concentration in the thawing medium had no effect on embryo viability. The freezing of 1. 5 mol/L G +0. 25 mol/L S resulted in a significantly higher (P<0. 01) hatching rate (73. 0%) than that of 1. 5 mol/L G (49. 2%) and 1. 5 mol/L G + 0. 5 mol/L S (56. 2%).
关键词
快速冷冻
牛
体外受精
胚胎
rapid freezing
cryoprotection
bovine
in vitro fertilization (IVF)
blastocyst
embryo
hatching rate
