Successful cryopreservation of expanded equine blastocysts.

This research explores the successful cryopreservation of expanded equine blastocysts, potentially due to the collapse of blastocoel fluid and using a dimethylsulfoxide-containing medium or ethylene glycol-containing medium.

Objective of the Research

• The study focuses on better understanding the process and effectiveness of cryopreservation for expanded equine blastocysts which have previously been difficult to cryopreserve successfully. The researchers hypothesize that this difficulty may be due to the volume of blastocoele fluid or due to the presence of the embryonic capsule in equines.

Methodology and Techniques Used in the Study

• The first part of the study was to vitrify non-biopsied small embryos using dimethylsulfoxide-containing medium (DM) or an ethylene glycol-containing medium (EG).
• A third group of small embryos was vitrified using EG, but was warmed using sucrose (EG/s).
• The second part of the study was to observe the survival of expanded (larger than 300 μm in diameter) blastocysts when they were biopsied and vitrified using DM or EG/s, resulting in the collapse of capsule and blastocyst.
• A variation of the biopsy technique, Central biopsy technique, was used to aspirate blastocoele fluid before vitrification of expanded blastocysts.
• In the final experiment, expanded blastocysts of size ranging from 407 to 565 μm in diameter were biopsied from the periphery, and the blastocoele fluid was removed with gentle suction. Then, these were vitrified with EG/s.

Findings of the Study

• Small embryos in DM and EG/s were able to grow in culture after vitrification, and established pregnancies after transfer.
• Expanded blastocysts that were biopsied and vitrified using DM and EG/s treatments resulted in normal pregnancies after warming and transfer.
• The study indicated that the more blastocoele fluid that was lost during biopsy, the higher the survival rate after vitrification. This discovery led to changing the biopsy technique to the central biopsy method.
• Finally, expanded blastocysts of specific sizes that were biopsied from the periphery and had their blastocoele fluid removed resulted in a high rate of normal pregnancies after vitrification with EG/s.
• Overall, the research highlighted that the collapse of blastocoel and vitrification in fine-diameter pipettes facilitated successful cryopreservation of expanded equine blastocysts.