FREEZING OF EMBRYOS
The basic knowledge of the main events that occur during the freezing-thawing process reduces the lethal effects that can harm the embryos. Embryos are permeable to cryoprotectant. DMSO, 1.2 propanediol or glycerol are among the cryoprotectants used in the various stages to dilute the non-frozen parts. This is done at a certain temperature to reduce the concentration of solutes.
The main objective during the freezing process is to prevent the formation of ice inside the cell. The formation of ice should start at a temperature close to the melting point in order to avoid over freezing. This is achieved through seeding, which is the process that starts the formation of ice through mechanical means.
Freezing process: PROH is the most commonly used cryoprotectant for freezing embryos at early stages (zygote or cells), because it’s more permeable and less toxic.
Balance of embryos in solution at room temperature, loading of embryos in straws, decreasing temperature and balancing before seeding, seeding, quick freezing, direct immersion of the straws in liquid nitrogen.
Success of the transfer of frozen-thawed embryos depends on: the freezing process, clinical parameters (ovarian response influences the outcome of cryopreservation) and embryo quality. Embryos of regular appearance and normal aspect survive significantly better compared to those of irregular Blastomeres and with the presence of fragments.
Another aspect that is advisable is the embryonic transfer if only more than 50% of the frozen Blastomeres survive; the survival rate and the number of intact Blastomeres are also linked to the rates of implantation and gestation.
The results of embryo freezing could significantly increase clinical pregnancies if the embryos to be transferred are selected not only based on morphological characteristics after defrosting, but also in its ability to divide for an additional period of 24 hours, as this additional embryonic division indicates not only the morphological survival, but metabolic as well. On the other hand, a large proportion of embryos that are not divided after defrosting show an increase of chromosomal abnormalities.
The pronuclear stage may be the best for freezing because the oocyte is in the process of fertilization, but has not reached syngamy. It must be made when the pronuclei and the nucleolus are aligned in the equatorial plate. It is very useful in cases where, for various reasons, the transfer is not fresh, in cases of ovarian hyperstimulation syndrome, donated oocytes, bleeding from the recipient, in cases where the patient can not show up to the transfer center, in cases where the proper amount of embryos are not obtained though diagnostic preimplantation and it's necessary to accumulate them for several cycles to ensure the success of the embryo biopsy.
Some the advantages of freeze oocytes include: