Fertilization failure is a common problem in assisted reproductive Technology (ART). The main reason for fertilization failure of conventional IVF fertilization is sperm penetration failure, and the main reason of ICSI is insufficient oocyte activation. Artificial assisted activation may provide an effective technique to rescue fertilization failure. In this study, standard ICSI procedures were applied to save fertilization failure of unfertilized mature oocytes in IVF cycles. The unfertilized mature oocytes after ICSI were activated by calcium ion, or injected with calcium chloride/activated with mechanical stimulated and then transfer to calcium ion to improve fertilization. In this study, different artificial assisted activation methods were used to save the fertilization failure and assess its effective and subsequent embryo development potential.

A variety of mechanical, electrical, and chemical methods has been used to trigger the calcium oscillations to activate oocytes. Mechanical and chemical activation are the most commonly used methods for artificial oocyte activation, which can mimic calcium oscillations saving fertilization failure.

Control group, a single spermatozoon was injected into the failed fertilization MII oocyte in conventional IVF cycle. What to do when ICSI fails? The investigators collected the unfertilized MII oocytes and divided them into 3 groups to perform different activation methods, including chemical calcium ionophore activation (experiment group 1), CaCl2 injected combined with calcium ionophore activation (experiment group 2), and mechanical stimulation combined with calcium ionophore activation (experiment group 3). Calcium ionophore A23187 (Sigma) was used for assisted activation. The final solution was 10 μmol/L. The oocytes were exposed to the calcium ionophore A23187 for 10 min at 37°C in 5% CO2. The oocytes were checked for pronucleus formation at 16-20 hours after activation. Fertilized oocytes were cultured in vitro for 3-5 days, the developmental potential of the activated embryos were observed.

This study want to explore the effective of different artificial assisted activation methods, to improve the fertilization outcome of unfertilized oocytes after ICSI or IVF.

Inclusion Criteria:

• 8 ≤ Number of retrieved oocytes ≤ 20;
• 18.5< Body Mass Index (BMI) <25;
• The patients who suffered complete fertilization failure or low fertilization after performing standard ICSI/IVF cycle. Low fertilization was defined as less than 33% fertilization rate.

Exclusion Criteria:

• Normal fertilization IVF/ICSI cycle.