Blastocoele collapse by micropipetting prior to vitrification gives excellent survival and pregnancy outcomes for human day 5 and 6 expanded blastocysts

Hiraoka, Kenichiro; Kinutani, Masayuki; Kinutani, Kazuo

doi:10.1093/humrep/deh504

Cited by 176 publications

(117 citation statements)

References 12 publications

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“…Since then, vitrification has been widely used for the cryopreservation of human oocytes [4,5], in addition to pronuclear stage [6,7], cleavage stage [4,[8][9][10][11], and blastocyst-stage [4,[12][13][14] embryos. Human embryo vitrification has been attempted with a variety of vessels such as electron microscope grids [15,16], open pulled and hemi-straws [17], the Flexipet [18], the Cryotop [4] and the CryoLoop [19,20].…”

Section: Introductionmentioning

confidence: 99%

Vitrification of mouse embryos at 2-cell, 4-cell and 8-cell stages by cryotop method

Zhang

Cui

Ling

et al. 2009

J Assist Reprod Genet

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Purpose The objective of this study was to investigate the effects of vitrification on the preimplantation developmental competence of mouse 2-cell, 4-cell and 8-cell stage embryos. Methods Mouse 2-cell, 4-cell and 8-cell stage embryos were cryopreserved using the cryotop vitrification method and subsequently warmed on a later date. The embryos were then assessed by their morphology, blastocyst formation and hatching rates. Additionally, trophectoderm (TE) and inner cell mass (ICM) cell numbers were compared in hatched blastocysts from the control and experimental groups. Results Vitrified embryos at the 2-cell, 4-cell and 8-cell stages appeared morphologically normal after warming. The overall survival rate of vitrified embryos at various stages after warming was 96.7% and there were no significant differences among 2-cell stage (96.0%), 4-cell stage (96.8%) and 8-cell stage (97.1%) embryos (P>0.05). The blastocyst formation rate (69.4%) and hatching rate (52.6%) of vitrified 2-cell embryos were significantly lower than that from the control group and vitrified 8-cell embryos (P<0.05). In the vitrified 4-cell embryo group, the blastocyst formation rate (90.3%) was similar to the 8-cell group (91.2%), but the hatching rate (60.0%) was significantly lower than that of the non-vitrified control ( 84.1%) and vitrified 8-cell embryo (78.4%) groups (P<0.05). When further development to the fully hatched blastocyst stage was compared, hatched blastocysts derived from vitrified 2-cell, 4-cell and 8-cell embryos had significantly lower cell counts both in the ICM and TE, as compared to fresh blastocysts (P<0.05). Among the vitrified 2-cell, 4-cell and 8-cell embryo groups, there were no significant differences in the cell counts of ICM and TE (P>0.05). Conclusions Although cryotop vitrification was suitable for the cryopreservation of mouse embryos from the 2-cell stage, 4-cell stage and 8-cell stage without significant loss of survival, vitrification had an adverse effect on the development of 2-cell embryos. Mouse embryos at the 8-cell stage had the best tolerance for vitrification and would yield the highest level of post-vitrification developmental competence among early cleavage stage embryos. Nevertheless, it is unclear how these findings can be extrapolated to human embryos.

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Section: Introductionmentioning

confidence: 99%

Vitrification of mouse embryos at 2-cell, 4-cell and 8-cell stages by cryotop method

Zhang

Cui

Ling

et al. 2009

J Assist Reprod Genet

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“…In all the three cases, most surplus embryos were at morula stage on day 6 and expanded blastocysts were available for cryopreservation only on day 7. It is our policy to cryopreserve not morula stage but expanded blastocyst stage in order to cryopreserve better quality embryo [4]. Therefore, re-vitrification was done on day 7 rather than day 6 in these three cases of the present report.…”

Section: Patient Treatmentmentioning

confidence: 96%

“…The embryos were vitrified by the method developed by Kuwayama et al [3] using a cryotop (Kitazato Supply Co., Fujinomiya, Japan), albeit with slight modifications, and has been described previously [4]. The cryotop consists of a 0.4 mm wide×20 mm long×0.1 mm thick polyethylene strip attached to a plastic handle and equipped with a cover straw.…”

Section: Vitrification Of Embryomentioning

confidence: 99%

“…In fresh blastocyst transfer, previous investigators have found superior implantation rates with fresh transfer occurring at day 5 as compared with day 6 [13,14] and at days 5 and 6 as compared with day 7 [15,16]. On the other hand, blastocysts vitrified on day 5 have a pregnancy potential superior [5,6] or similar [4,8] as compared with that of day 6 blastocysts. However, to the best of our knowledge, there are no reports on pregnancies following the transfer of day 7 vitrified and warmed blastocysts.…”

Section: Introductionmentioning

confidence: 97%

“…This greatly increases the cooling and warming rate, while reducing chilling injuries and ice crystal formation. The efficacy of vitrification in small volumes is demonstrated by good survival rates of human blastocysts with the cryotop [3][4][5][6], the cryoloop [7,8], electron microscope grids [9,10] or the hemi-straw [11,12].…”

Section: Introductionmentioning

confidence: 99%

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Case report: Two successful pregnancies following the transfer of re-vitrified human day 7 blastocysts developed from vitrified cleaved embryos

Fujimoto

Tateaki

Hiraoka

et al. 2008

J Assist Reprod Genet

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Purpose To report successful pregnancies after the transfer of re-vitrified human day 7 blastocysts developed from vitrified cleaved embryos. Methods and results A total of five day 7 blastocysts developed from vitrified cleaved embryos were re-vitrified and re-warmed. All of five re-vitrified day 7 blastocysts (100%) survived after warming and were transferred to three patients. Two of the women became clinically pregnant. Of these women, one woman delivered a healthy baby and the other pregnancy is ongoing at 26 weeks of gestation. Conclusions This is the first report of successful pregnancies after the transfer of re-vitrified human day 7 blastocysts developed from vitrified cleaved embryos.

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Cryopreservation of mammalian embryos: Advancement of putting life on hold

Tsang

Chow

2010

Birth Defects Research Pt C

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Rodent transgenesis and human-assisted reproductive programs involve multistep handling of preimplantation embryos. The efficacy of production and quality of results from conventionally scheduled programs are limited by temporal constraints other than the quality and quantities of embryos per se. The emergence of vitrification, a water ice-free cryopreservation technique, as a reliable way to arrest further growth of preimplantation embryos, provides an option to eliminate the time constraint. In this article, current and potential applications of cryopreservation to facilitate laboratory animal experiments, colony management, and human-assisted reproductive programs are reviewed. Carrier devices developed for vitrification in the last two decades are compared with an emphasis on their physical properties that infer cooling rate of samples and sterility assurance. Biological impacts of improved cryopreservation on preimplantation embryos are also discussed.

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Blastocoele collapse by micropipetting prior to vitrification gives excellent survival and pregnancy outcomes for human day 5 and 6 expanded blastocysts

Abstract: The results suggest that artificial shrinkage with pipetting is a simple and effective technique to assist successful cryopreservation of expanded blastocysts by vitrification.

Cited by 176 publications

References 12 publications

Vitrification of mouse embryos at 2-cell, 4-cell and 8-cell stages by cryotop method

Vitrification of mouse embryos at 2-cell, 4-cell and 8-cell stages by cryotop method

Case report: Two successful pregnancies following the transfer of re-vitrified human day 7 blastocysts developed from vitrified cleaved embryos

Cryopreservation of mammalian embryos: Advancement of putting life on hold

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