Multiple vitrification-warming and biopsy procedures on human embryos: clinical outcome and neonatal follow-up of children

Abstract STUDY QUESTION Does double vitrification and warming of human blastocysts having undergone biopsy once or twice have an impact on the clinical outcome? SUMMARY ANSWER The clinical pregnancy rate obtained with double vitrification single biopsy blastocysts was comparable to that obtained with single vitrification single biopsy blastocysts in our center in the same time period (46%; 2016–2018), whereas that obtained with double-vitrified double-biopsied blastocysts seemed lower and will need further study. WHAT IS KNOWN ALREADY Genetic testing on cryopreserved unbiopsied embryos involves two cryopreservation procedures. Retesting of failed/inconclusive-diagnosed blastocysts inevitably involves a second round of biopsy and a second round of vitrification as well. To what extent this practice impacts on the developmental potential of blastocysts has been studied to a limited extent so far and holds controversy. Additionally, the obstetrical/perinatal outcome after the transfer of double-vitrified/single or double-biopsied blastocysts is poorly documented. STUDY DESIGN, SIZE, DURATION This retrospective observational study included 97 cycles of trophectoderm biopsy and preimplantation genetic testing (PGT) on vitrified-warmed embryos followed by a second round of vitrification between March 2015 and December 2019. PARTICIPANTS/MATERIALS, SETTING, METHODS In 36 warming cycles, no biopsy was performed on the embryos before the first vitrification (single biopsy group). In 61 warming cycles, the embryos had been biopsied on Day 3 (n = 4) or on Day 5/6 (n = 57) before the first vitrification (double biopsy group). A second biopsy was mostly indicated in cycles of failed or inconclusive diagnosis at the first biopsy. Two cycles involved a more specific mutation test for X-linked diseases on male embryos and one cycle involved testing for a second monogenic indication supplementary to a previously tested reciprocal translocation. Post-warming suitability for biopsy, availability of genetically transferable embryos and clinical outcome of subsequent frozen-thawed embryo transfer (FET) cycles were reported. Neonatal follow-up of the children was included. MAIN RESULTS AND THE ROLE OF CHANCE In total, 91 cleavage-stage embryos and 154 blastocysts were warmed, of which 34 (37.4%) and 126 (81.8%), respectively, were of sufficient quality to undergo trophectoderm biopsy and were subsequently vitrified for a second time. Out of these, 92 underwent biopsy for the fi