Quick Tips to Vitrification
Cryopreservation of oocytes or embryos is a critical step in maximizing the efficiency of any IVF cycle. To achieve clinical success with cryopreservation is highly variable from laboratory to laboratory, and may depend on many factors including patient age and stimulation protocol, quality of embryos selected for freezing, developmental stage at freezing, media formulation including type of method used, parameters of cooling and warming.
All through our time in and outside of the laboratory, during discussions, we have heard vitrification is so difficult. Believe us, vitrification, is not difficult. Having said this, yes, we agree that there are certain critical steps you need to focus on and keep in mind and the rest just follows through.
Principle:
Vitrification can be defined as an extreme elevation in viscosity, i.e. solidification of solutions without ice crystal formation at low temperature. This phenomenon can be induced by either applying an extreme cooling rate or by using high concentrations of cryoprotectant solutions. Although human embryo and oocyte vitrification was slow to evolve, it has become an invaluable technology in the field of reproductive medicine.
Tip no 1 - The ratio of freezing:
If you have not yet noticed, it is important to notice the difference in size as the egg becomes an embryo and an embryo becomes a blastocyst. As the transition takes place the size keeps increasing and thus we use different sized denupets to handle each of these. However, at the same time, it is important to remember that the size of the device (cryotop/cryolock or cryo-loop) does not change. Thus, this means, as the size goes up we need to reduce the number we freeze on 1 device.
Tip 2 – Media left on the device:
The number of oocytes/embryos loaded on the device is also associated with addition of more media. The technique of vitrification demands leaving as less amount of media as possible surrounding the oocytes/embryos.
Tip 3 – Start with Day 3, Day5 and then oocyte
The principle of vitrification demands, removing maximum water content from the oocytes/embryos. We may not be able to see this lysis until we thaw, and hence it is necessary to observe the same while vitrifying. Oocytes have the highest water content followed by blastocyst and then Day 3.
Tip 4 – Immediate immersion in Liquid nitrogen
As suggested earlier, it is important to adhere to the protocol stringently. Thus as per each protocol, you will get approximately 60 seconds to load the material on the device. Once this step is done, you have to ensure device either open or close has to be immediately dipped in liquid nitrogen.
Tip 5 – Document your observations:
Along with following the timeline and technique of vitrifying, it is extremely important to document your observations of the process. For e.g.; if you took more than usual time to load the oocytes/embryos or the shrinkage did not happen and the reason you think could cause it. These may not be important at the time of vitrifying, storing but extremely important to analyse if the oocyte/embryo does not survive.
If you are beginning your training in vitrification, then all the above mentioned points are applicable. If you have been performing vitrification for a long time, this could serve as a checklist for you to see if you have not missed any of the above. We are sure adopting the above mentioned tips will bring a positive change in your technique and its outcome.
Comments
Post a Comment