Basic study in cryopreservation and chromosomal analysis of single blastomeres.

着床前胚胎活检作为一种研究着床前胚胎发育潜力的工具被广泛应用，也为产前诊断提供了可能。本研究的目的首先是研究4细胞和8细胞小鼠胚胎单个卵裂球的发育潜力，其次是探讨单个卵裂球成功冷冻保存和核型的可能性。(1)通过挤压技术从4细胞和8细胞小鼠胚胎中去除单个卵裂球对体外囊胚期发育没有影响（4细胞和8细胞期分别占对照组的80%、活检胚胎的75%和78.3%）。然而，在移植到假妊娠受体小鼠后，活检胚胎的着床率明显低于完整对照组（4细胞组24.3%，8细胞组34.8%，对照组50%）。(2)大多数单卵裂球在体外经历了多次细胞分裂。单卵裂球短时间培养16小时后，4细胞活检的27.3%和8细胞活检的52.2%获得核型。然而，由于细胞死亡和技术问题，20 - 30%的活检囊胚丢失。(3)当冷冻介质为1.5M DMSO+0.25M蔗糖，采用5步稀释法时，活检的单个卵裂球不能成功冷冻保存。本研究开发的技术可能为产前诊断的研究提供有用的工具。

Biopsy of the preimplantation embryo has wide application as a tool to investigate the developmental potential of the preimplantation embryo and also offered a possibility for prenatal diagnosis. The purpose of this study were first, to examine the developmental potential of single blastomeres of four-and eight-cell mouse embryos, and second to investigate the possibility of successful cryopreservation and karyotyping of single blastomeres.(1) Removal of a single blastomere from 4- and 8-cell mouse embryos by extrusion technique did not affect development to the blastocyst stage in vitro (80% of control,75% and 78.3% of biopsied embryo at 4- and 8-cell stage, respectively). However,after transfer to pseudopregnant recipient mice, the implantation rates of biopsied embryos were significantly lower than intact-controls (24.3% at 4-cell, 34.8% at 8-cell and 50% at control, respectively).(2) Most single blastomeres had undergone several cell divisions in vitro. After a short culture of single blastomeres with subsequent exposure to colcemid for 16 hours, 27.3% of 4-cell and 52.2% of 8-cell biopsies were karyotyped. However, 20 to 30% of biopsied blstomeres were lost through cell death and technical problems.(3) Biopsied single blastomeres could not be successfully cryopreserved when the freezing medium contained 1.5M DMSO+0.25M sucrose and 5-step dilution method was used. The techniques developed in this study may provide the useful tool for the study of prenatal diagnosis.