Elevated incidence of monozygotic twinning is associated with extended embryo culture, but not with zona pellucida manipulation or freeze-thaw procedure

Specific in vitro fertilization techniques, such as the number of embryos replaced, zona pellucida manipulation, and freeze-thaw procedure, did not significantly increase the rate of monozygotic twinning, except for the day of embryo transfer.

Volume 109, Issue 6, Pages 1044–1050


Hanyan Liu, M.M., Jianqiao Liu, M.D., Shiping Chen, Ph.D., Xiangjin Kang, Ph.D., Hongzi Du, M.D., Lei Li, Ph.D.



To identify the incidence and risk factors associated with IVF-conceived monozygotic twinning (MZT).


Retrospective study.


Academic hospital.


A total of 3,463 women with clinical pregnancies between January 2014 and February 2015 were analyzed.



Main Outcome Measure(s)

The measures were the incidence of MZT based on the number of embryos that were replaced, type of insemination method (conventional IVF or intracytoplasmic sperm injection [ICSI]), with or without the use of assisted hatching (AH), and day of embryo transferred in fresh and frozen cycles.


Ninety-three women (2.69%) with MZT were observed. No statistically significant differences were observed in the cycle parameters of fresh or frozen cycles between MZT and other non-MZT pregnancies. Specific IVF procedures or techniques, such as the number of embryo replaced, zona pellucida manipulation (ICSI and AH), and freeze-thaw procedure, did not significantly increase the rate of MZT, except for the day of embryo transferred. Compared with day 3 transferred, day 4 and 5/6 transferred showed an increased probability of MZT (odds ratio [OR], 2.73; 95% confidence interval [CI], 1.16–6.42 for day 4 transferred and OR, 3.68; 95% CI, 2.29–5.93 for day 5/6 transferred).


Extended culture (advanced embryo stage) in fresh and frozen cycles appeared to be associated with increased rates of MZT. The effect of the number of embryos transferred, ICSI and AH, and freeze-thaw procedures on the risk for MZT was not demonstrated.

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