微型载体无保护剂冷冻微量精子的初步研究

发布时间：2018-05-04 16:18

本文选题：冷冻环 + 无保护剂玻璃化　；参考：《华中科技大学》2006年硕士论文

【摘要】： 第一部分精子冷冻环无保护剂玻璃化冷冻方法的实验研究目的通过探讨冷冻环无保护剂玻璃化冷冻微量精子的可行性,试图寻找一种适合于睾丸或附睾微量精子的冷冻方法。方法正常标本上游处理后分别进行慢速冷冻和冷冻环无保护剂的玻璃化冷冻,复苏后分别从活动率及电镜下超微结构等指标来比较两种冷冻方法的效果。结果两种方法冷冻后其活动率之间差异无显著性(44.5% vs 43.5%, P0.05),但均较未冷冻时明显下降(44.5% vs 43.5% vs 88.5%, P0.001)。超微结构亦较未冷冻时发生了一定的改变,但核结构基本保持完整。结论精子冷冻环无保护剂的玻璃化冷冻是一种简单、方便而有效的冷冻方法;冷冻环有望成为适合于睾丸或附睾微量精子冷冻的一种新型载体。第二部分微滴法无保护剂玻璃化冷冻正常精子的可行性探讨目的探讨在不添加冷冻保护剂的情况下微滴法玻璃化冷冻正常精子的可行性方法收集16份正常精液标本,上游处理后分别进行慢速冷冻和微滴法冷冻,微滴法即用拉细巴氏管将标本形成10ul的液滴,直接投入液氮内冻存,复苏后镜检其活动率。结果两种方法冷冻后其活动率之间差异无显著性(40% vs 37.5%, P0.05),但均较未冷冻时明显下降(40% vs 37.5% vs 85%, P0.001)。结论无保护剂的微滴法玻璃化冷冻正常人类精子可以取得较理想的结果;但需进一步探索以提高其复苏率。第三部分微滴法无保护剂玻璃化冷冻PESA精子的可行性探讨目的探讨微滴法无保护剂玻璃化冷冻PESA精子的可行性方法收集12份PESA精子标本,洗涤处理后进行无保护剂的微滴法冷冻,用拉细巴氏管将标本形成10ul的液滴,直接投入液氮内冻存,复苏后镜检其活动率。结果将复苏后活动精子数大于30个作为复苏成功的指标,12份标本有8份复苏成功,复苏成功率为66.7%。结论微滴法无保护剂玻璃化冷冻PESA精子可以取得一定的成功率;无保护剂的微滴法是微量精子的一种有效的冷冻方法。
[Abstract]:The first part: experimental study on vitrification of sperm cryopreservation ring with unprotected agent Purpose By exploring the feasibility of vitrification of microsperm by vitrification without cryopreservation ring, this paper attempts to find a suitable method for freezing spermatozoa of testis or epididymis. Method After upstream treatment of normal specimens, the cryopreservation and vitrification of cryopreservation without protective agent were carried out respectively. After resuscitation, the effects of the two freezing methods were compared from the activity rate and ultrastructure under electron microscope. Result There was no significant difference in the activity rate between the two methods after freezing (44.5% vs 43.5%, P0.05%, P 0.05), but they were significantly lower than those without freezing (44.5% vs 43.5% vs 88.5%, P 0.001). The ultrastructure also changed, but the nuclear structure remained intact. Conclusion Vitrification of unprotected spermatozoa is a simple, convenient and effective method for cryopreservation of spermatozoa. The cryopreservation ring is expected to be a new type of carrier suitable for spermatozoa cryopreservation of testis or epididymis. The second part: feasibility of vitrification of normal spermatozoa by microdrop method Purpose Study on the feasibility of vitrification of normal spermatozoa by microdrop method without adding cryopreservation agent Method Sixteen normal semen samples were collected and cryopreserved respectively by slow freezing and microdrop method after upstream treatment. Microdrop method was used to form the 10ul droplets from the specimen with a thin pasteurian tube, and was directly frozen in liquid nitrogen. The activity rate was examined by microscope after resuscitation. Result There was no significant difference in activity rate between the two methods after freezing (40% vs 37.5%, P 0.05), but they were significantly lower than those without freezing by 40% vs 37.5% vs 85g, P0.001, respectively. Conclusion Vitrification of normal human spermatozoa without protective agent can obtain ideal results, but further exploration is needed to improve the recovery rate of human spermatozoa. The third part: feasibility of vitrification of PESA spermatozoa by microdrop method Purpose Feasibility of vitrification of PESA spermatozoa by microdrop method Method Twelve PESA spermatozoa samples were collected and frozen by microdrop method without protective agent after washing. The samples were formed into 10ul droplets by pull-down pasteurian tube and directly put into liquid nitrogen cryopreservation. The motility rate was examined by microscope after resuscitation. Result After resuscitation, the number of motile sperm more than 30 was taken as the index of success of resuscitation, and 8 of 12 specimens were successfully resuscitated, the success rate of resuscitation was 66.7%. Conclusion Vitrification of PESA spermatozoa with microdrop method can obtain a certain success rate, and microdrop method without protection agent is an effective freezing method for microsperm.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R321;R318.52

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