Miralles A, Sacristán J C, Martínez E, García J
Department of Cryobiology and Cell Therapy, Hospital Duran i Reynals, Barcelona, Spain.
Cryobiology. 1994 Apr;31(2):171-9. doi: 10.1006/cryo.1994.1020.
Embryo cryopreservation procedures have been highly developed to support in vitro fertilization techniques, but the clinical results have not met initial expectations. Many variables influence the outcome of the embryo cryopreservation procedure. Exposure to classical freezing media does not affect further embryo development. Therefore, the first crucial step seems to be embryo thermal behavior during the freezing/thawing procedure. With the aim of avoiding thermal oscillations to the embryos due to liquid nitrogen injection into the freezing chamber, we have developed a straw thermal stabilizer (STS) cryopreservation accessory that when adapted to a programmable biological freezer offers a more exact control of the straws temperature and also a greater reproducibility of the freezing process. The STS incorporates a seeding mechanism controlled by specially designed software. For the biological assays, 2-cell mouse embryos were cooled until seeding at -7 degrees C or frozen to -120 degrees C using 1.5 M 1,2-propanediol and 0.1 M sucrose. As control groups, embryos were either untreated or were exposed to cryoprotectants, developing to blastocyst at rates of 90.8 and 88.4%, respectively. The embryos cooled until seedling or subjected to the complete freezing procedure developed into blastocysts at rates of 95.7 and 82.8%, respectively, disregarding the effect of the cryoprotectant exposure. These results show a substantial improvement in cryopreservation with no loss in embryo development due to the seeding procedure.
胚胎冷冻保存技术已高度发展以支持体外受精技术,但临床结果未达最初预期。许多变量会影响胚胎冷冻保存程序的结果。暴露于传统冷冻培养基中不会影响胚胎的进一步发育。因此,关键的第一步似乎是冷冻/解冻过程中胚胎的热行为。为避免因向冷冻室注入液氮而使胚胎受到热振荡影响,我们开发了一种麦管热稳定器(STS)冷冻保存附件,将其适配于可编程生物冷冻机时,能更精确地控制麦管温度,且冷冻过程的可重复性更高。STS包含一个由专门设计的软件控制的播种机制。在生物学试验中,将2细胞期小鼠胚胎冷却至-7℃进行播种,或使用1.5M 1,2 - 丙二醇和0.1M蔗糖冷冻至-120℃。作为对照组,胚胎要么不做处理,要么暴露于冷冻保护剂中,发育成囊胚的比率分别为90.8%和88.4%。冷却至播种阶段或经历完整冷冻程序的胚胎发育成囊胚的比率分别为95.7%和82.8%,不受冷冻保护剂暴露影响。这些结果表明,冷冻保存有显著改善,且播种程序不会导致胚胎发育受损。