中文摘要：

在卵母细胞低温保存过程中,低温保护剂的添加与去除是必不可少的步骤,但此过程会对细胞造成致命的渗透损伤和毒性损伤。为了研究保护剂添加和去除联合过程对猪MII期细胞的损伤程度,本文设计并制作了一个适合卵母细胞低温保护剂连续添加与去除的微流体装置,分别采用微流控线性法和传统的分步法加载和去除低温保护剂,分析了两种方法对卵母细胞的体积变化以及存活率与发育潜能的影响。结果表明：微流控线性法加载和去除低温保护剂时,卵母细胞的体积变化明显小于分步法;线性法处理后细胞的存活率、卵裂率、囊胚率分别为95.3%、64.4%、19.4%,都显著高于传统的分步法（79.4%、43.6%、9.7%）（P〈0.05）。因此,微流体装置用于卵母细胞低温保护剂添加与去除是有效的,能够显著减小细胞的渗透损伤,为卵母细胞低温保存技术提供了新思路。

英文摘要：

Oocyte cryopreservation has become a practical tool in assisted reproductive technology and fertility preservation. Cryoprotective agents （CPAs） addition and removal are essential steps during oocytes cryopreservation, however, the processes of CPAs addition and removal may cause fatal osmotic and toxic injuries to oocytes. In order to study the injuries of porcine MII-stage oocytes during CPAs addition and removal combined processes and minimize osmotic and toxic injuries to oocytes, primarily, a microiluidic device for continuous loading and unloading of CPAs was designed and fabricated in this study. CPAs were loaded and unloaded with the microfluidic linear method and the conventional step-wise method, respectively. Then, the cell volume changes and the effects on the survival rate and developmental potential of oocytes were investigated. The results showed that the oocyte volume changes with the microfluidic device were obviously less than step-wise method. The survival, cleavage and blastocyst rate of oocytes were 95.3% , 64.4% , and 19.4% , respectively, which were significantly higher than the traditional step-wise method （ 79.4% , 43.6% , and 9. 7% ） （ P 〈 0. 05 ）. In conclusion, microfluidic device can be used efficiently to load and unload of CPAs, and significantly reduce the osmotic shock to oocytes, which may open up a new path for oocyte cryopreservation.