Bath Mary L
Division of Molecular Genetics of Cancer, The Walter and Eliza Hall Institute of Medical Research, Victoria, Australia.
J Reprod Dev. 2011 Feb;57(1):92-8. doi: 10.1262/jrd.10-039n. Epub 2010 Nov 10.
Although procedures for in vitro fertilization with cryopreserved sperm have been published there is a lack of data indicating that the cryoprotectant and cryopreservation procedures used for those procedures were optimal. To redress this, fertilization rate of eggs exposed to sperm in vitro was used as the outcome in the optimization of raffinose concentration in the cryoprotectant (raffinose in water), volume of cryoprotectant, and freezing conditions for C57BL/6J mouse sperm. Sperm were frozen in a cylindrical Dewar with an internal diameter and height of 14.0 cm and 36.0 cm respectively. The optimal concentration of raffinose was 23-24% (510-540 mOsm/kg). The optimal volume of cryoprotectant used to prepare the sperm suspension from a single mouse was 180-400 µl, and sperm proved most fertile when frozen 13-25 mm above liquid nitrogen. Raffinose in the fertilization medium did not inhibit fertilization. Fertilized eggs transferred to oviducts of recipient mice developed into viable offspring.
尽管已经发表了关于使用冷冻精子进行体外受精的程序，但缺乏数据表明用于这些程序的冷冻保护剂和冷冻保存程序是最佳的。为了解决这个问题，将体外暴露于精子的卵子的受精率用作优化冷冻保护剂（水中的棉子糖）中棉子糖浓度、冷冻保护剂体积以及C57BL/6J小鼠精子冷冻条件的结果。精子在分别具有14.0厘米内径和36.0厘米高度的圆柱形杜瓦瓶中冷冻。棉子糖的最佳浓度为23-24%（510-540毫渗摩尔/千克）。用于从单只小鼠制备精子悬液的冷冻保护剂的最佳体积为180-400微升，并且当在液氮上方13-25毫米处冷冻时精子的受精能力最强。受精培养基中的棉子糖不抑制受精。转移到受体小鼠输卵管中的受精卵发育成有活力的后代。
