The effect of multiple cryopreservation procedures and blastomere biopsy on the in-vitro development of mouse embryos

Preimplantation genetic diagnosis is currently used in clinical practice. In experienced hands the biopsy procedure alone does not affect the further in-vitro and in-vivo developmental potential of animal and human embryos. No data exist on the combination of cryopreservation of embryos at the pronuclear and/or 8-cell stage and/or biopsy at the 8-cell stage. Pronuclear stages of mouse F1 hybrids (C57Bl/jxCBA) were harvested and divided into several experimental groups. The developmental rates of zygotes, which were neither biopsied nor cryopreserved were used as data control. Others were only cryopreserved at the pronuclear stage (C-PN), or at the cleavage stage (C-CS), or both. Each of these groups was also combined with or without a biopsy. Only the hatched blastocyst rate (HBR), but not the 'simple' blastocyst rate, showed significant differences between groups. Neither C-PN (HBR = 60.42%), nor C-CS (63.16%), nor a combination of both (59.46%) had an impact on the hatched blastocyst rate when compared with that of the control group (67.46%). The biopsy procedure (55.93%) also proved not to be harmful for the embryos. The embryos, which were C-PN and C-CS, and subsequently biopsied, showed a significantly lower hatched blastocyst rate (39.62%) than that of the control, C-PN, C-CS, and C-PN/C-CS groups (P < 0.05). The combination of C-PN and cleavage-stage biopsy also lead to a lower hatched blastocyst rate (42.22%), compared with that of the control group (P < 0.05). It was concluded that couples must be advised that an effect on embryos which have undergone a combined cryopreservation and micromanipulation procedure cannot be ruled out. However, cryopreservation at the pronuclear or at the 8-cell stage alone, or in combination with a biopsy procedure, does not influence the further development of the embryo.