本文選題：冷凍環(huán) + 無保護劑玻璃化　； 參考：《華中科技大學》2006年碩士論文

【摘要】： 第一部分精子冷凍環(huán)無保護劑玻璃化冷凍方法的實驗研究 目的 通過探討冷凍環(huán)無保護劑玻璃化冷凍微量精子的可行性,試圖尋找一種適合于睪丸或附睪微量精子的冷凍方法。 方法 正常標本上游處理后分別進行慢速冷凍和冷凍環(huán)無保護劑的玻璃化冷凍,復蘇后分別從活動率及電鏡下超微結(jié)構(gòu)等指標來比較兩種冷凍方法的效果。 結(jié)果 兩種方法冷凍后其活動率之間差異無顯著性(44.5% vs 43.5%, P0.05),但均較未冷凍時明顯下降(44.5% vs 43.5% vs 88.5%, P0.001)。超微結(jié)構(gòu)亦較未冷凍時發(fā)生了一定的改變,但核結(jié)構(gòu)基本保持完整。 結(jié)論 精子冷凍環(huán)無保護劑的玻璃化冷凍是一種簡單、方便而有效的冷凍方法;冷凍環(huán)有望成為適合于睪丸或附睪微量精子冷凍的一種新型載體。 第二部分微滴法無保護劑玻璃化冷凍正常精子的可行性探討 目的 探討在不添加冷凍保護劑的情況下微滴法玻璃化冷凍正常精子的可行性 方法 收集16份正常精液標本,上游處理后分別進行慢速冷凍和微滴法冷凍,微滴法即用拉細巴氏管將標本形成10ul的液滴,直接投入液氮內(nèi)凍存,復蘇后鏡檢其活動率。 結(jié)果 兩種方法冷凍后其活動率之間差異無顯著性(40% vs 37.5%, P0.05),但均較未冷凍時明顯下降(40% vs 37.5% vs 85%, P0.001)。 結(jié)論 無保護劑的微滴法玻璃化冷凍正常人類精子可以取得較理想的結(jié)果;但需進一步探索以提高其復蘇率。 第三部分微滴法無保護劑玻璃化冷凍PESA精子的可行性探討 目的 探討微滴法無保護劑玻璃化冷凍PESA精子的可行性 方法 收集12份PESA精子標本,洗滌處理后進行無保護劑的微滴法冷凍,用拉細巴氏管將標本形成10ul的液滴,直接投入液氮內(nèi)凍存,復蘇后鏡檢其活動率。 結(jié)果 將復蘇后活動精子數(shù)大于30個作為復蘇成功的指標,12份標本有8份復蘇成功,復蘇成功率為66.7%。 結(jié)論 微滴法無保護劑玻璃化冷凍PESA精子可以取得一定的成功率;無保護劑的微滴法是微量精子的一種有效的冷凍方法。
[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.
【學位授予單位】：華中科技大學
【學位級別】：碩士
【學位授予年份】：2006
【分類號】：R321;R318.52

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