Knowing about Vitrification

Knowing about Vitrification

Knowing about Vitrification

Vitrification is a rapid freeze technique that transforms liquid state media with embryos into glass state without affecting the integrity of the embryo. This technique overcomes the most important issue that was seen during slow freezing which was ice crystal formations inside an embryos or blastocyst which damaged the cells and reduced their survival rates.

Vitrification is a process that involves faster dehydrating of the embryo/blast during freezing and a quicker rehydrating during thaw.

Vitrification involves use of cryoprotectants such as dimethyl sulphoxide (DMSO) ethylene glycol(EG) and glycerol. These cryoprotectants upon cooling become viscous and form solid state like glass without forming ice crystals. These solutes mixed along with a dehydrating agent such as sucrose that has a higher permeability to penetrate inside an embryo helps in protecting an embryo to the osmotic shock and toxicity.

Procedure:

Initially vitrification was One step procedure where embryos were exposed a higher concentration of CPA’s( cryoprotecting agents) for 1-2 mins and later dipped into liquid nitrogen. This technique shows reduced survival rates of embryos upon thawing as a result the technique was modified into a two step procedure as described below:

Step 1:embryos are introduced into equilibrium solution(ES solution) that contains relatively low concentrations of CPA’s (7.5% DMSO + 7.5%EG) for 7-15 mins. This step allows exchange of solutes between media and the embryos. Today most of the commercial kits state that this process happen at room temperature allowing smoother exchange of solutes.

Step 2: this step is a rather shorter step whereby the equilibrated embryos are now dipped into vitrification solution(VS). VS solution contains a higher concentration of CAP’s (usually 15%DMSO + 15%EG) as well as a dehydrating agent sucrose. During this process the embryo undergoes a quick dehydration of all the water contents inside the embryo and later gets rehydrated with the CPA’s from the VS solution. Embryos are kept in VS solution for about 30-90 seconds and then loaded into a cryotop or any cryodevice and quickly dipped into liquid nitrogen. The logical reasoning behind such a short exposer to high levels of CPA’s is that the cell size and water content play an important role in the rate of embryo dehydration. minimal exposer to higher concentration of CPA’s helps in achieving higher CPA equilibration.

Vitrification vs slow freezing:

Slow freezing is a step-wise performed technique that allows embryos to be exposed to lower concentrations of CPA’s initially followed by gradually increasing the concentrations of CPA and dehydrating agents. Later the embryos are frozen in a slow controlled rates of cooling(0.3c/min) starting from 25c to -125c during a time period of 60-80 mins. slower dehydrating rate causes ice crystal formations inside an embryo/blast that causes cell damages resulting in lower survival rates. Vitrification is a quicker method performed in less than 15 mins where embryos gets exposed to higher CPA’s in a two-step manner and gets frozen rapidly (cooling rate of ~20,000c/min). Rapid dehydrating rates and the solutes used in vitrification technique can convert media from liquid state into solid glass state. This provides a protective environment around the embryos which upon thaw results in higher survival rates. Numerous Studies have shown that vitrified embryos have higher the implantation rates in comparison to slow-cooled embryos.

With the introduction of PGD and single embryo transfer methods into the Morden day IVF vitrification has shown more promising results then slow freezing. Vitrification is a simple less time consuming and cost effective technique. Embryos stored in a thin cryodevice has helped embryologist to store it in a smaller space solving the biggest problem of cryostorage. The Morden day IVF is more effective in generating higher pregnancies and lower complications thanks to vitrification.