Effect of holding technique and culture drop size in individual or group culture on blastocyst development after ICSI of equine oocytes with low meiotic competence.

The study explored how different techniques of holding oocytes and varying culture drop sizes affect the development of blastocysts after the injection of equine oocytes with low meiotic competence. It found that compact-cumulus oocytes can be effectively stored using the EH treatment before maturation process, and that individual culturing of equine embryos yields acceptable development rate of blastocysts.

Methodology

• The study comprised three experiments. In the first one, oocytes were held using roscovitine for around 16 to 18 hours before maturation, then fertilized through intracytoplasmic sperm injection. Each was cultured individually in droplets of varied sizes: 2.5, 5, 10, or 50 microliters.
• In the second experiment, oocytes were cultured either with roscovitine or held in a modified M199 with a 20% serum at room temperature, also known as the EH treatment, for around 16 to 18 hours. They were then matured, fertilized, and grown in groups using 5 microliters of culture medium for each embryo.
• The third experiment involved holding the oocytes using the EH treatment, after which they were matured and fertilized. There were two studies within this experiment: Study 3.1, where the injected oocytes were cultured individually in droplet sizes as done in Experiment 1 and; Study 3.2, where groups of 2 to 7 oocytes were cultured in fixed droplet sizes of 5 or 50 microliters.

Findings

• The development rates of individually-cultured blastocysts were found to be largely unaffected by the size of the droplet, across both Experiments 1 and 3. The rates ranged between 15% and 29%.
• In Experiment 2, there was no statistically significant difference in the blastocyst development rates between the different oocyte holding treatments, with rates ranging from 17% to 23%.
• Results from Experiment 3 suggested that for group-cultured oocytes, the size of the droplet (whether 5 or 50 microliters) did not significantly impact the rate of blastocyst development, at 39% and 27% respectively.

Conclusion

• The study concluded that compact-cumulus oocytes can be effectively held in the EH treatment before maturation.
• Individual culturing of equine embryos can yield acceptable rates of blastocyst development.
• The highest rate of blastocyst development (39%) was obtained when oocytes were cultured in groups within a 5 microliter droplet.