The Vitrification Of Oocytes And Early Embryos In Mouse

The study is to filtrate the best cryoprotectant of vitrification of mature oocytes from mouse, and tentatively verify damage mechanisms during freezing of mature oocytes from mouse. We collected mature oocytes from 6-8-week-old female mice (ICR), and then cryopreserved the oocytes above with cryoprotectant EFS,VS1 and A10-10 respectively. To investigate the change of the fertilized and developmental rate and of the morphology of the cytoskeleton of the post-thaw oocytes, we fertilized and stained the post-thaw oocytes. Then in order to investigate the best cryoprotectant for mouse early embryos, we vitrified the embryos in different stages (2cell,4-8cell and morula) with EFS40, EFS20-40, DAP213 and GP25.Main results and conclusion were showed as follows:1. The results of the rate of normal morphology and fertilizing after vitrification with cryoprotectant EFS,VS1 and A10-10 shows that, compared with cryoprotectant EFS and VS1, the cryoprotectant A10-10 was obtained a higher normal morphology rate and fertilizing rate. The above indicated that A10-10 was the best cryoprotectant for oocytes of mouse.2. The results of fertilizing rate of post-thaw oocytes after cutting zona pellucida shows that the fertilizing rate of post-thaw oocytes after cutting zona pellucida is higer (54.8%) than with whole zona pellucida (11.8%). This indicated that the freezing-thawing process changed the structure of zona pellucida, and blocked sperm penetration, and then resulted in declinig fertilizing rate.3. The results of changes of survival and cytoskeleton of post-thaw oocytes after staining showed that not only the freezing maked oocytes damages, but also hyperisotonic cryoprotectant changed survival and cytoskeleton of oocytes.4. The resulits of developmental competence of post-thaw embryos in different stages (2- cell,4-8-cell, morula) after cryopreserving with cryoprotectant EFS40, EFS20-40, DAP213 and GP25 showed that compared with cryoprotectant EFS20-40 and EFS40 with EG, cryoprotectant GP25 and DAP213 with GL and DMSO had more toxicity for embryo.5. EFS20-40 was the best cryoprotectant for 2-cell and morula stage embryos of mouse, and EFS40 was the best cryoprotectant for 4-8-cell stage embryos.