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Background: The present study was undertaken to examine the usefulness of both vitrification and assisted hatching (AH) on blastocysts that originate from embryos showing different qualities during their cleavage stage., Methods: A total of 281 blastocysts were vitrified (93 vitrification-warming cycles) in a mixture of ethylene glycol-dimethylsulphoxide-Ficoll and sucrose using the Hemi-Straw (HS) carrier system. After warming, AH using the partial dissection technique was performed in 36 cycles., Results: After warming and culture for 24 h, a total of 168 blastocysts (60%) was suitable for embryo transfers and a total of 25 ongoing pregnancies (27%) was obtained. Forty-nine transfers of 96 no-AH blastocysts and 36 transfers of 72 AH blastocysts resulted in an implantation rate of 13 and 22% respectively (P < 0.05). The percentage of transfers with at least one hatching blastocyst was significantly higher after application of AH (69 versus 33%) (P < 0.001). In all, 73 and 38% of blastocysts showing respectively optimal and non-optimal embryo development during the early stage were available for transfer (P < 0.001). Consequently, implantation rates of 19 and 6% were obtained after transfers of blastocysts showing respectively optimal and poor embryo development., Conclusions: Artificial opening of the zona pellucida after warming of vitrified blastocysts significantly improved the rate of transfers with hatched blastocysts and the implantation and pregnancy rates. The percentage of blastocysts that survived the HS vitrification procedure and were available for embryo transfer is related to their previous developmental quality.

Background: In 1996, with the introduction of sequential media, we set up a programme of cryopreservation of supernumerary morulae (day 4) and blastocysts (day 5) using a vitrification procedure. Our results showed that the efficiency of the vitrification method was dependent on the stage of embryo development and was negatively correlated with the expansion of the blastocoele. We postulated that a large blastocoele might disturb cryopreservative potential due to ice crystal formation during the cooling step. We analysed therefore the effectiveness of reducing before vitrification the volume of the blastocoelic cavity., Method: Day 4 and day 5 embryos were vitrified in 40% ethylene glycol-18% Ficoll and 0.3 mol/l sucrose before plunging the straws directly into liquid nitrogen. Artificial shrinkage of the blastocyst was achieved after pushing a needle into the blastocoele cavity until it contracted., Results: The survival rate post-thawing of day 4 and intact day 5 embryos was correlated with the volume of the blastocoele. In the control group only 20.3% blastocysts or expanded blastocysts survived as compared with 54.5 and 58.5% with morulae and early blastocyst respectively. After puncturing the blastocoelic cavity, an increase in the survival rate of up to 70.6% was noted. The pregnancy rates were improved after the artificial shrinkage procedure (20.5%) compared with the control intact blastocyst group (4.5%) (not significant). After reduction of the blastocoelic cavity, a significant increase in the implantation rate per vitrified blastocyst was observed (12.0 versus 1.4% P < 0.01)., Conclusions: Our results showed that survival rates in cryopreserved expanded blastocysts could be improved by reducing the fluid content. This was presumably because mechanical damage caused by ice crystal formation was avoided. These observations should be considered when establishing a strategy and a protocol for cryopreservation of day 5 embryos.

The banked sperm of 5 of 13 Hodgkin patients was thawed for assisted reproduction. The post-thaw sperm characteristics were extremely impaired. Nevertheless, in 80% of all trials fertilization in vitro was obtained and conceptus could be transferred. All spouses became pregnant and seven healthy infants were born. One could recommend every Hodgkin patient to bank his sperm before chemotherapy. In vitro fertilization techniques with this frozen-thawed sperm reveal promising results in comparison with artificial insemination to maintain the reproductive capacity of male Hodgkin patients.