人植入前胚胎细胞超薄电镜切片的制作及应用TUNEL技术对该细胞凋亡的原位检测

发布时间：2018-01-12 16:27

本文关键词：人植入前胚胎细胞超薄电镜切片的制作及应用TUNEL技术对该细胞凋亡的原位检测　出处：《华中科技大学》2006年硕士论文　论文类型：学位论文

【摘要】： 目的:改进人植入前胚胎细胞超薄切片的制备方法,观察不同时期人早期胚胎细胞超微结构的变化。方法:选取我院生殖中心试管婴儿助孕技术后异常受精的3原核胚胎,于2-8细胞期进行固定,改进透射电镜的常规操作方法,使用3%～4%琼脂进行预包埋,制成超薄切片后在镜下观察植入前胚胎细胞凋亡的超微结构。结果:在镜下可见胚胎细胞正常形态和凋亡的形态变化。结论:使用本方法可以进行少量胚胎的超薄切片。目的:建立原位末端标记法(TdT-mediated nick end labeling TUNEL)检测人植入前胚胎凋亡征象的方法。方法:取试管婴儿助孕技术后异常受精的3原核胚胎,于2-10细胞期进行固定,采用TUNEL法检测3原核胚胎中的凋亡征象。结果:TUNEL阳性信号为绿色强荧光,位于胞核,呈环状或颗粒状。在发育正常和发育停滞胚胎中,未检测到凋亡征象;在有碎片的胚胎中20%可检测到TUNEL阳性信号。结论:采用TUNEL法加荧光共焦显微镜观察凋亡细胞的方法可以有效检测人植入前胚胎中的凋亡征象,其中细胞碎片的出现可能与凋亡有关。
[Abstract]:Aim: to improve the preparation of ultrathin sections of human preimplantation embryonic cells and observe the ultrastructural changes of human early embryonic cells at different stages. Methods: the 3 prokaryotic embryos of IVF in our hospital were fixed at 2-8 cell stage, and the conventional operation of transmission electron microscope was improved. The ultrastructure of apoptosis of preimplantation embryo cells was observed under microscope after ultrathin sections were made by using 3% Agar and 4% Agar. Results: the morphological changes of normal and apoptotic embryonic cells were observed under microscope. Conclusion: this method can be used to carry out ultrathin sections of a small number of embryos. Aim: to establish a TdT-mediated nick end labeling Tunel method for the detection of apoptosis in human preimplantation embryos. Methods: the 3 prokaryotic embryos with abnormal fertilization after IVF were fixed in 2-10 cell phase and the apoptotic signs in 3 prokaryotic embryos were detected by TUNEL method. Results the positive signal of TUNEL was green strong fluorescence, which was located in the nucleus and was annular or granular. No apoptotic sign was detected in normal and stunted embryos. TUNEL positive signals were detected in 20% of the embryos with fragments. Conclusion: the method of TUNEL and fluorescence confocal microscopy can be used to detect apoptosis in human preimplantation embryos, and the appearance of cell fragments may be related to apoptosis.
【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

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